TREA

SERPINA-MODULATING COMPOSITIONS AND METHODS

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Information

  • Patent Application
  • 20240084334
  • Publication Number
    20240084334
  • Date Filed
    September 18, 2023
    a year ago
  • Date Published
    March 14, 2024
    6 months ago
Information
Abstract
The disclosure provides, e.g., compositions, systems, and methods for targeting, editing, modifying, or manipulating a host cell's genome at one or more locations in a DNA sequence in a cell, tissue, or subject. Gene modifying systems for treating alpha-1 antitrypsin deficiency (AATD) are described.
Description
SEQUENCE LISTING

The instant application contains a Sequence Listing which has been submitted electronically in XML, format compliant with WIPO Standard ST.26 and is hereby incorporated by reference in its entirety. Said XML copy, created on Sep. 15, 2023, is named V2065-702420FT SL.XML and is 31,115,476 bytes in size.


CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of International Application No. PCT/US2022/076073, filed Sep. 7, 2022, which claims the benefit of U.S. Provisional Application No. 63/241,970, filed Sep. 8, 2021, U.S. Provisional Application No. 63/253,087, filed Oct. 6, 2021, and U.S. Provisional Application No. 63/303,905, filed Jan. 27, 2022. The contents of the aforementioned applications are hereby incorporated by reference in their entirety.


BACKGROUND

Integration of a nucleic acid of interest into a genome occurs at low frequency and with little site specificity, in the absence of a specialized protein to promote the insertion event. Some existing approaches, like CRISPR/Cas9, are more suited for small edits that rely on host repair pathways, and are less effective at integrating longer sequences. Other existing approaches, like Cre/loxP, require a first step of inserting a loxP site into the genome and then a second step of inserting a sequence of interest into the loxP site. There is a need in the art for improved compositions (e.g., proteins and nucleic acids) and methods for inserting, altering, or deleting sequences of interest in a genome.


AATD is characterized by low circulating levels of AAT. AAT is produced primarily in liver cells and secreted into the blood, but it is also made by other cell types including lung epithelial cells and certain white blood cells. AAT inhibits several serine proteases secreted by inflammatory cells (most notably neutrophil elastase [NE], proteinase 3, and cathepsin G) and thus protects organs, such as the lung, from protease-induced damage, especially during periods of inflammation.


The two most common clinical variants of AAT are E264V (PiS) and E342K (PiZ) alleles. The clinical single nucleotide variant E342K (PiZ) leads to structurally unstable and/or inactive AAT protein and, as a consequence, causes toxicity in liver and inactivity in lung. Inheritance is autosomal codominant. More than a half of AATD patients harbor at least one copy of the mutation E342K.


The mutation most commonly associated with AATD involves a substitution of glutamic acid for lysine (E342K) in the SERPINA1 gene that encodes the AAT protein. The E342K mutation is located at the hinge between the beta sheet and the Reactive Center Loop (RCL) of the AAT protein and causes a loop-sheet dimer that later can extend to form long chains of loop-sheet polymers that that aggregate AAT-Z proteins inside the rough Endoplasmic Reticulum (rER) of hepatocytes during biosynthesis. This mutation, known as the Z mutation or the Z allele, leads to misfolding of the translated protein, which is therefore not secreted into the bloodstream and. Consequently, circulating AAT levels in individuals homozygous for the Z allele (PiZZ) are markedly reduced; only approximately 15% of mutant Z-AAT protein folds correctly and is secreted by the cell. An additional consequence of the Z mutation is that the secreted Z-AAT has reduced activity compared to wild-type protein, with 40% to 80% of normal antiprotease activity (American thoracic society/European respiratory society, Am J Respir Crit Care Med. 2003; 168(7):818-900; and Ogushi et al. J Clin Invest. 1987; 80(5):1366-74).


There are two disease phenotypes associated with the PiZZ genotype. The accumulation of polymerized Z-AAT protein within hepatocytes results in a gain-of-function cytotoxicity that can result in cellular stress, inflammation, fibrosis, cirrhosis, and hepatocellular carcinoma (HCC) and neonatal liver disease in 12% of patients. This accumulation may spontaneously remit but can be fatal in a small number of children. A loss-of-function phenotype results from the reduced systemic levels of AAT that lead to increased protease digestion of connective tissue in the lower airway. Excess protease-digestion of the connective tissues and alveoli linings deteriorates lung elasticity and pulmonary functions, leading to emphysema, a hallmark of Chronic Obstructive Pulmonary Disease (COPD). This effect is severe in PiZZ individuals and typically manifests in middle age, resulting in a decline in quality of life and shortened lifespan (mean 68 years of age) (Tanash et al. Int J Chron Obstruct Pulm Dis. 2016; 11:1663-9). The effect is more pronounced in PiZZ individuals who smoke, resulting in an even further shortened lifespan (58 years). Piitulainen and Tanash, COPD 2015; 12(1):36-41. PiZZ individuals account for the majority of those with clinically relevant AATD lung disease.


A milder form of AATD is associated with the SZ genotype in which the Z-allele is combined with an S-allele. The S allele is associated with somewhat reduced levels of circulating AAT, but causes no cytotoxicity in liver cells. The result is clinically significant lung disease but not liver disease. Fregonese and Stolk, Orphanet JRare Dis. 2008; 33:16. As with the ZZ genotype, the deficiency of circulating AAT in subjects with the SZ genotype results in unregulated protease activity that degrades lung tissue over time and can result in emphysema, particularly in smokers.


While limited treatment options for AATD exist, there is currently no cure. A small fraction of newborn patients and patients at the advanced stage of liver disease undergo liver transplant. The current standard of care for AAT deficient individuals who have or show signs of developing significant lung disease is augmentation therapy or protein replacement therapy. Augmentation therapy involves administration of a human AAT protein concentrate purified from pooled donor plasma to augment the missing AAT. This treatment involves weekly infusion of AAT proteins purified from healthy blood donors. Although infusions of the plasma protein have been shown to improve survival or slow the rate of emphysema progression, augmentation therapy is often not sufficient under challenging conditions (e.g., active lung infection). Augmentation therapy also fails to restore the normal physiological regulation of AAT in patients and efficacy has been difficult to demonstrate. In addition, augmentation therapy cannot address liver disease, which is driven by the toxic gain-of-function of the Z allele. Accordingly, there is a need for new and more effective treatments for AATD.


SUMMARY OF THE INVENTION

This disclosure relates to novel compositions, systems and methods for altering a genome at one or more locations in a host cell, tissue or subject, in vivo or in vitro. The disclosure provides gene modifying systems that are capable of modulating (e.g., inserting, altering, or deleting sequences of interest) alpha-1 antitrypsin (AAT) activity and methods of treating alpha-1 antitrypsin deficiency (AATD) by administering one or more such systems to alter a genomic sequence at a single nucleotide to correct the SERPINA1 PiZ mutation causing alpha-1 antitrypsin deficiency.


In one aspect, the disclosure relates to a system for modifying DNA to correct a human SERPINA1 gene mutation causing AATD comprising (a) a nucleic acid encoding a gene modifying polypeptide capable of target primed reverse transcription, the polypeptide comprising (i) a reverse transcriptase domain and (ii) a Cas9 nickase that binds DNA and has endonuclease activity, and (b) a template RNA comprising (i) a gRNA spacer that is complementary to a first portion of the human SERPINA1 gene, (ii) a gRNA scaffold that binds the polypeptide, (iii) a heterologous object sequence comprising a mutation region to correct the mutation, and (iv) a primer binding site (PBS) sequence comprising at least 3, 4, 5, 6, 7, or 8 bases of 100% homology to a target DNA strand at the 3′ end of the template RNA. The SERPINA1 gene may comprise an E342K mutation (also referred to as a PiZ mutation). The template RNA sequence may comprise a sequence described herein, e.g., in Table 1, 3, 4, 5, 6a, 6B, X2, X3, X3a, X5, or XX.


The gRNA spacer may comprise at least 15 bases of 100% homology to the target DNA at the 5′ end of the template RNA. The template RNA may further comprise a PBS sequence comprising at least 5 bases of at least 80% homology to the target DNA strand. The template RNA may comprise one or more chemical modifications.


The domains of the gene modifying polypeptide may be joined by a peptide linker. The polypeptide may comprise one or more peptide linkers. The gene modifying polypeptide may further comprise a nuclear localization signal. The polypeptide may comprise more than one nuclear localization signal, e.g., multiple adjacent nuclear localization signals or one or more nuclear localization signals in different regions of the polypeptide, e.g., one or more nuclear localization signals in the N-terminus of the polypeptide and one or more nuclear localization signals in the C-terminus of the polypeptide. The nucleic acid encoding the gene modifying polypeptide may encode one or more intein domains.


Introduction of the system into a target cell may result in insertion of at least 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 500, or 1000 base pairs of exogenous DNA. Introduction of the system into a target cell may result in deletion, wherein the deletion is less than 2, 3, 4, 5, 10, 50, or 100 base pairs of genomic DNA upstream or downstream of the insertion. Introduction of the system into a target cell may result in substitution, e.g., substitution of 1, 2, or 3 nucleotides, e.g., consecutive nucleotides.


The heterologous object sequence may be at least 5, 10, 25, 50, 100, 150, 200, 250, 300, 400, 500, 600, or 700 base pairs.


In one aspect, the disclosure relates to a pharmaceutical composition comprising the system described above and a pharmaceutically acceptable excipient or carrier, wherein the pharmaceutically acceptable excipient or carrier is selected from the group consisting of a plasmid vector, a viral vector, a vesicle, and a lipid nanoparticle. In one aspect, the disclosure relates to a pharmaceutical composition comprising the system described above and multiple pharmaceutically acceptable excipients or carriers, wherein the pharmaceutically acceptable excipients or carriers are selected from the group consisting of a plasmid vector, a viral vector, a vesicle, and a lipid nanoparticle, e.g., where the system described above is delivered by two distinct excipients or carriers, e.g., two lipid nanoparticles, two viral vectors, or one lipid nanoparticle and one viral vector. The viral vector may be an adeno-associated virus (AAV).


In one aspect, the disclosure relates to a host cell (e.g., a mammalian cell, e.g., a human cell) comprising the system described above.


In one aspect, the disclosure relates to a method of correcting a mutation in the human SERPINA1 gene in a cell, tissue or subject, the method comprising administering the system described above to the cell, tissue or subject, wherein optionally the correction of the mutant SERPINA1 gene comprises an amino acid substitution of K342E (reversing the pathogenic substitution which is E342K). The system may be introduced in vivo, in vitro, ex vivo, or in situ. The nucleic acid of (a) may be integrated into the genome of the host cell. In some embodiments, the nucleic acid of (a) is not integrated into the genome of the host cell. In some embodiments, the heterologous object sequence is inserted at only one target site in the host cell genome. The heterologous object sequence may be inserted at two or more target sites in the host cell genome, e.g., at the same corresponding site in two homologous chromosomes or at two different sites on the same or different chromosomes. The heterologous object sequence may encode a mammalian polypeptide, or a fragment or a variant thereof. The components of the system may be delivered on 1, 2, 3, 4, or more distinct nucleic acid molecules. The system may be introduced into a host cell by electroporation or by using at least one vehicle selected from a plasmid vector, a viral vector, a vesicle, and a lipid nanoparticle.


Features of the compositions or methods can include one or more of the following enumerated embodiments.


Enumerated Embodiments

1. A template RNA comprising, e.g., from 5′ to 3′:

    • (i) a gRNA spacer that is complementary to a first portion of the human SERPINA1 gene, wherein the gRNA spacer has a sequence comprising the core nucleotides of a gRNA spacer sequence of Table 1, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer (e.g., comprises one or more flanking nucleotides that are adjacent to the core nucleotides), or wherein the gRNA spacer has a sequence of a gRNA spacer of Table 6A, 6B, X2, X3, X3a, X5, or XX, or a sequence having 1, 2, or 3 substitutions thereto;
    • (ii) a gRNA scaffold that binds a gene modifying polypeptide (e.g., binds the Cas domain of the gene modifying polypeptide),
    • (iii) a heterologous object sequence comprising a mutation region to introduce a mutation into (e.g., to correct a mutation in) a second portion of the human SERPINA1 gene (wherein optionally the heterologous object sequence comprises, from 5′ to 3′, a post-edit homology region, a mutation region, and a pre-edit homology region), and
    • (iv) a primer binding site (PBS) sequence comprising at least 3, 4, 5, 6, 7, or 8 bases with 100% identity to a third portion of the human SERPINA1 gene.


2. The template RNA of embodiment 1, wherein the heterologous object sequence comprises the core nucleotides of an RT template sequence from Table 3, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence, or wherein the heterologous object sequence comprises a sequence of an RT template sequence from Tables 6A or 6B.


3. The template RNA of embodiment 1, wherein the heterologous object sequence comprises the core nucleotides of the RT template sequence of Table 3 that corresponds to the gRNA spacer sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence (e.g., comprises one or more flanking nucleotides that are adjacent to the core nucleotides), or wherein the heterologous object sequence comprises a sequence of an RT template sequence from Tables 6A or 6B.


4. The template RNA of any of the preceding embodiments, wherein the heterologous object sequence has the sequence of a heterologous object sequence from a template RNA set out in Table X3, or X3a, or a sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto, or a sequence having 1, 2, or 3 substitutions thereto.


5. The template RNA of any of the preceding embodiments, wherein the heterologous object sequence has a length of 6-16 nucletodies (e.g., 6, 8, 10, 12, 14, 15, or 16 nucleotides).


6. The template RNA according to any one of the preceding embodiments wherein the PBS sequence has a sequence comprising the core nucleotides of the PBS sequence from the same row of Table 3 as the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence (e.g., comprises one or more flanking nucleotides that are adjacent to the core nucleotides).


7. The template RNA according to any one of embodiments 1-5, wherein the PBS sequence has a sequence comprising the core nucleotides of a PBS sequence of Table 3 that corresponds to the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, the gRNA spacer sequence, or both, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence, or wherein the PBS sequence has a sequence comprising the a PBS sequence of Tables 6A or 6B, or a sequence having 1, 2, or 3 substitutions thereto, that corresponds to the RT template sequence, the gRNA spacer sequence, or both.


8. The template RNA of any of the preceding embodiments, wherein the PBS sequence has the sequence of a PBS from a template RNA set out in Table X3, or X3a, or a sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto, or a sequence having 1, 2, or 3 substitutions thereto.


9. The template RNA of any of the preceding embodiments, wherein the PBS sequence has a length of 8-12 nucleotides (e.g., 8, 9, 10, 11, or 12 nucleotides).


10. The template RNA according to any of embodiments 1-9, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


11. The template RNA according to any of embodiments 1-9, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12 that corresponds to the RT template sequence, the gRNA spacer sequence, or both, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


12. The template RNA of any of the preceding embodiments, wherein the gRNA scaffold has the sequence of a gRNA scaffold from a template RNA set out in Table X2, X3, or X3a, or a sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto.


13. The template RNA of any of the preceding embodiments, which comprises a sequence of a template RNA set out in Table X2, X3, or X3a, or a sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto.


14. A template RNA comprising, e.g., from 5′ to 3′:

    • (i) a gRNA spacer that is complementary to a first portion of the human SERPINA1 gene,
    • (ii) a gRNA scaffold that binds a gene modifying polypeptide (e.g., binds the Cas domain of the gene modifying polypeptide),
    • (iii) a heterologous object sequence comprising a mutation region to introduce a mutation into (e.g., to correct a mutation in) a second portion of the human SERPINA1 gene, wherein the heterologous object sequence comprises the core nucleotides of an RT template sequence of Table 3, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence, or wherein the heterologous object sequence comprises an RT template sequence of Tables 6A or 6B; and
    • (iv) a PBS sequence comprising at least 3, 4, 5, 6, 7, or 8 bases of 100% identity to a third portion of the human SERPINA1 gene.


15. The template RNA of embodiment 14, wherein the gRNA spacer comprises the core nucleotides of a gRNA spacer sequence of Table 1, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer sequence, or wherein the gRNA spacer comprises a gRNA spacer sequence of Tables 6A or 6B.


16. The template RNA of embodiment 14, wherein the heterologous object sequence comprises the core nucleotides of the gRNA spacer sequence of Table 1 that corresponds to the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer sequence, or wherein the heterologous object sequence comprises the nucleotides of the gRNA spacer sequence of Tables 6A or 6B.


17. The template RNA according to any one of embodiments 14-16, wherein the PBS sequence has a sequence comprising the core nucleotides of the PBS sequence from the same row of Table 3 as the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


18. The template RNA according to any one of embodiments 14-17, wherein the PBS sequence has a sequence comprising the core nucleotides of a PBS sequence of Table 3 that corresponds to the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, the gRNA spacer sequence, or both, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence, or wherein the PBS sequence has a sequence comprising the a PBS sequence of Tables 6A or 6B that corresponds to the RT template sequence, the gRNA spacer sequence, or both.


19. The template RNA according to any of embodiments 14-18, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 6A or 12, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


20. The template RNA according to any of embodiments 14-18, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 6A or 12 that corresponds to the RT template sequence, the gRNA spacer sequence, or both, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


21. A gene modifying system for modifying DNA, comprising:

    • (a) a first RNA comprising, from 5′ to 3, (i) a guide RNA sequence that is complementary to a first portion of the human SERPINA1 gene, wherein the guide RNA sequence has a sequence comprising the core nucleotides of a spacer sequence of Table 1, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the guide RNA sequence; and (ii) a sequence (e.g., a scaffold region) that binds a gene modifying polypeptide (e.g., binds the Cas domain of the gene modifying polypeptide), and
    • (b) a second RNA comprising (iii) a heterologous object sequence comprising a nucleotide substitution to introduce a mutation into a second portion of the human SERPINA1 gene (wherein optionally the heterologous object sequence comprises, from 5′ to 3′, a post-edit homology region, a mutation region, and a pre-edit homology region), (iv) a primer region comprising at least 5, 6, 7, or 8 bases of 100% identity to a third portion of the human SERPINA1 gene, and (v) an RRS (RNA binding protein recognition sequence) that binds a gene modifying protein.


22. The gene modifying system of embodiment 21, wherein the heterologous object sequence comprises the core nucleotides of an RT template sequence from Table 3, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence.


23. The gene modifying system of embodiment 21, wherein the heterologous object sequence comprises the core nucleotides of the RT template sequence of Table 3 that corresponds to the gRNA spacer sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence.


24. The gene modifying system of any one of embodiments 21-23, wherein the PBS sequence has a sequence comprising the core nucleotides of the PBS sequence from the same row of Table 3 as the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


25. The gene modifying system of one of embodiments 21-23, wherein the PBS sequence has a sequence comprising the core nucleotides of a PBS sequence of Table 3 that corresponds to the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, the gRNA spacer sequence, or both, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


26. The gene modifying system of any one of embodiments 21-25, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


27. The gene modifying system of any one of embodiments 21-25, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12 that corresponds to the RT template sequence, the gRNA spacer sequence, or both, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


28. A gene modifying system for modifying DNA, comprising:

    • (a) a first RNA comprising, from 5′ to 3, (i) a guide RNA sequence that is complementary to a first portion of the human SERPINA1 gene, and (ii) a sequence (e.g., a scaffold region) that binds a gene modifying polypeptide (e.g., binds the Cas domain of the gene modifying polypeptide), and
    • (b) a second RNA comprising (iii) a heterologous object sequence comprising a nucleotide substitution to introduce a mutation into a second portion of the human SERPINA1 gene, wherein the heterologous object sequence comprises the core nucleotides of an RT template sequence of Table 3, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence, and (iv) a primer region comprising at least 5, 6, 7, or 8 bases of 100% homology to a third portion of the human SERPINA1 gene, and (v) an RRS (RNA binding protein recognition sequence) that binds a gene modifying protein.


29. The gene modifying system of embodiment 28, wherein the gRNA spacer comprises the core nucleotides of a gRNA spacer sequence of Table 1, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer sequence.


30. The gene modifying system of embodiment 28, wherein the heterologous object sequence comprises the core nucleotides of the gRNA spacer sequence of Table 1 that corresponds to the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer sequence.


31. The gene modifying system of any one of embodiments 28-30, wherein the PBS sequence has a sequence comprising the core nucleotides of the PBS sequence from the same row of Table 3 as the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


32. The gene modifying system of any one of embodiments 28-30, wherein the PBS sequence has a sequence comprising the core nucleotides of a PBS sequence of Table 3 that corresponds to the RT template sequence, the gRNA spacer sequence, or both, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


33. The gene modifying system of any one of embodiments 28-32, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


34. The gene modifying system of any one of embodiments 28-32, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12 that corresponds to the RT template sequence, the gRNA spacer sequence, or both, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


35. A gRNA comprising (i) a gRNA spacer sequence that is complementary to a first portion of the human SERPINA1 gene, wherein the gRNA spacer has a sequence comprising the core nucleotides of a gRNA spacer sequence of Table 1, Table 2, or Table 4, or a sequence having 1, 2, or 3 substitutions thereto and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer sequence; and (ii) a gRNA scaffold.


36. The gRNA of embodiment 35, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


37. The gRNA of embodiment 35, wherein the gRNA scaffold comprises a sequence of a gRNA scaffold of Table 12 that corresponds to the gRNA spacer sequence, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


38. A template RNA comprising: (iii) a heterologous object sequence comprising a mutation region to introduce a mutation into a second portion of the human SERPINA1 gene, wherein the heterologous object sequence comprises the core nucleotides of an RT template sequence of Table 3, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence, and (iv) a PBS sequence comprising at least 5, 6, 7, or 8 bases of 100% homology to a third portion of the human SERPINA1 gene.


39. The template RNA according to embodiment 38, wherein the PBS sequence has a sequence comprising the core nucleotides of the PBS sequence from the same row of Table 3 as the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


40. The template RNA according to embodiment 38, wherein the PBS sequence has a sequence comprising the core nucleotides of a PBS sequence of Table 3 that corresponds to the RT template sequence, or a sequence having 1, 2, or 3 substitutions thereto, and optionally comprises one or more consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the PBS sequence.


41. The template RNA according to any one of embodiments 1-20 or 38-40, the gene modifying system of any one of embodiments 21-34, or the gRNA of any one of embodiments 35-37, wherein the mutation introduced by the system is a K342E mutation (e.g., to correct a pathogenic E342K mutation) of the SERPINA1 gene.


42. The template RNA according to any one of embodiments 1-20 or 38-41 or the gene modifying system of any one of embodiments 21-34 or 41, wherein the pre-edit sequence comprises between about 1 nucleotide to about 35 nucleotides (e.g., comprises about 1-5, 5-10, 10-15, 15-20, 20-25, 25-30, or 30-35 nucleotides) in length.


43. The template RNA according to any one of embodiments 1-20 or 38-42 or the gene modifying system of any one of embodiments 21-34, 41, or 42, wherein the mutation region comprises a single nucleotide.


44. The template RNA according to any one of embodiments 1-20 or 38-42 or the gene modifying system of any one of embodiments 21-34, 41, or 42, wherein the mutation region is at least two nucleotides in length.


45. The template RNA according to any one of embodiments 1-20, 38-42, or 44 or the gene modifying system of any one of embodiments 21-34, 41, 42, or 44, wherein the mutation region is up to 32 (e.g., up to 5, 10, 15, 20, 25, 30, or 32) nucleotides in length and comprises one, two, or three sequence differences relative to a second portion of the human SERPINA1 gene.


46. The template RNA according to any one of embodiments 1-20, 38-42, 44, or 45 or the gene modifying system of any one of embodiments 21-34, 41, 42, 44, or 45, wherein the mutation region comprises two sequences differences relative to a second portion of the human SERPINA1 gene.


47. The template RNA according to any one of embodiments 1-20, 38-42, or 44-46 or the gene modifying system of any one of embodiments 21-34, 41, 42, or 44-46, wherein the mutation region comprises a first region (e.g., a first nucleotide) designed to correct a pathogenic mutation in the SERPINA1 gene and a second region (e.g., a second nucleotide) designed to inactivate a PAM sequence (e.g., a “PAM-kill” mutation as described in Table 5).


48. The template RNA according to any one of embodiments 1-20, 38-46 or the gene modifying system of any one of embodiments 21-34 or 41-46, wherein the mutation region comprises less than 80%, 70%, 60%, 50%, 40%, or 30% identity to corresponding portion of the human SERPINA1 gene.


49. The template RNA of any one of the preceding embodiments, wherein the template RNA comprises one or more silent mutations (e.g., silent substitutions), e.g., as exemplified in Table 7B.


50. The template RNA of any of the preceding embodiments, wherein the mutation region comprises a first region designed to correct a pathogenic mutation in the SERPINA1 gene and a second region designed to introduce a silent substitution.


51. The template RNA of any one of the preceding embodiments, which comprises one or more chemically modified nucleotides.


52. A gene modifying system comprising:

    • a template RNA of any of embodiments 1-20, 38-42, or a system of any of embodiments 21-34 or 41-46, and
    • a gene modifying polypeptide, or a nucleic acid (e.g., RNA) encoding the gene modifying polypeptide.


53. The gene modifying system of embodiment 52, wherein the gene modifying polypeptide comprises:

    • a reverse transcriptase (RT) domain (e.g., an RT domain from a retrovirus, or a polypeptide domain having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acids sequence identity thereto); and
    • a Cas domain that binds to the target DNA molecule and is heterologous to the RT domain (e.g., a Cas9 domain); and
    • optionally, a linker disposed between the RT domain and the Cas domain.


54. The gene modifying system of embodiment 53, wherein the RT domain comprises:

    • (a) an RT domain of Table 6; or
    • (b) an RT domain from a murine leukemia virus (MMLV), a porcine endogenous retrovirus (PERV); Avian reticuloendotheliosis virus (AVIRE), a feline leukemia virus (FLV), simian foamy virus (SFV) (e.g., SFV3L), bovine leukemia virus (BLV), Mason-Pfizer monkey virus (MPMV), human foamy virus (HFV), or bovine foamy/syncytial virus (BFV/BSV).


55. The gene modifying system of embodiment 53 or 54, wherein the Cas domain comprises a Cas domain of Table X1, XX, or X5, or a sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acids sequence identity thereto.


56. The gene modifying system of any of embodiments 53-55, wherein the spacer comprises a spacer of Table XX, or a sequence having 1, 2, or 3 substitutions thereto, and the Cas domain comprises a Cas domain of the same row of Table XX or a sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acids sequence identity thereto.


57. The gene modifying system of any of embodiments 53-56, wherein the spacer comprises a spacer of Table XX, and the Cas domain comprises a Cas domain of the same row of Table XX.


58. The gene modifying system of any of embodiments 53-57, wherein the spacer comprises a spacer of Table X5, or a sequence having 1, 2, or 3 substitutions thereto, and the Cas domain comprises a Cas domain of the same row of Table X5, or a sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acids sequence identity thereto.


59. The gene modifying system of any of embodiments 53-58, wherein the spacer comprises a spacer of Table X5, and the Cas domain comprises a Cas domain of the same row of Table X5.


60. The gene modifying system of any of embodiments 53-59, wherein the spacer comprises a spacer of Table 6A, or a sequence having 1, 2, or 3 substitutions thereto, and the Cas domain comprises a Cas domain of the same row of Table 6A, or a sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acids sequence identity thereto.


61. The gene modifying system of any of embodiments 53-60, wherein the spacer comprises a spacer of Table 6A, and the Cas domain comprises a Cas domain of the same row of Table 6A.


62. The gene modifying system of any of embodiments 53-51, wherein the spacer comprises a spacer of Table 6B, or a sequence having 1, 2, or 3 substitutions thereto, and the Cas domain comprises a Cas domain of the same row of Table 6B, or a sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acids sequence identity thereto.


63. The gene modifying system of any of embodiments 53-62, wherein the spacer comprises a spacer of Table 6B, and the Cas domain comprises a Cas domain of the same row of Table 6B.


64. The gene modifying system of any one of embodiments 53-63, wherein the Cas domain comprises a Cas domain of Table 7 or Table 8.


65. The gene modifying system of any one of embodiments 53-64, wherein the Cas domain:

    • (a) is a Cas9 domain;
    • (b) is a SpCas9 domain, a BlatCas9 domain, a Nme2Cas9 domain, a PnpCas9 domain, a SauCas9 domain, a SauCas9-KKH domain, a SauriCas9 domain, a SauriCas9-KKH domain, a ScaCas9-Sc++domain, a SpyCas9 domain, a SpyCas9-NG domain, a SpyCas9-SpRY domain, or a St1Cas9 domain; and/or
    • (c) is a Cas9 domain comprising an N670A mutation, an N611A mutation, an N605A mutation, an N580A mutation, an N588A mutation, an N872A mutation, an N863 mutation, an N622A mutation, or an H840A mutation.


66. The gene modifying system of embodiment 65, wherein the Cas9 domain binds a PAM sequence listed in Table 7 or Table 12.


67. The gene modifying system of embodiment 66, wherein a second portion of the human SERPINA1 gene overlaps with a PAM recognized by the Cas domain, e.g., wherein the second portion of the human SERPINA1 gene is within the PAM or wherein the PAM is within the second portion of the human SERPINA1 gene).


68. The gene modifying system any one of embodiments 53-67, wherein the gRNA spacer is a gRNA spacer according to Table 1, and the Cas domain comprises a Cas domain listed in the same row of Table 1, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


69. The gene modifying system of any one of the preceding embodiments, wherein the template RNA comprises a sequence of a template RNA sequence of Table 6A or 6B or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


70. The gene modifying system of any one of embodiments 53-69, wherein:

    • (a) the template RNA comprises a sequence of a template RNA sequence of Table 3;
    • (b) the Cas domain comprises a Cas domain of Table 7 or Table 8;
    • (c) the linker comprises a linker sequence of Table 10 (e.g., of any of SEQ ID NOs: 5217, 5106, 5190, and 5218); and
    • (d) the gene modifying polypeptide comprises one or two NLS sequences from Table 11 (e.g., of any of SEQ ID NOs: 5245, 5290, 5323, 5330, 5349, 5350, 5351, and 4001).


71. The gene modifying system of any of embodiments 53-70, which produces a first nick in a first strand of the human SERPINA1 gene.


72. The gene modifying system of embodiment 71, which further comprises a second strand-targeting gRNA spacer that directs a second nick to the second strand of the human SERPINA1 gene.


73. The gene modifying system of embodiment 72, wherein the second strand-targeting gRNA comprises a sequence comprising the core nucleotides of a left gRNA spacer sequence or a right gRNA spacer sequence from Table 2, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the left gRNA spacer sequence or right gRNA spacer sequence.


74. The gene modifying system of embodiment 72, wherein the second strand-targeting gRNA comprises a sequence comprising the core nucleotides of a left gRNA spacer sequence or a right gRNA spacer sequence from Table 2 that corresponds to the gRNA spacer sequence of (i), and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the left gRNA spacer sequence or right gRNA spacer sequence.


75. The gene modifying system of embodiment 72, wherein the second strand-targeting gRNA comprises a sequence comprising the core nucleotides of a second nick gRNA sequence from Table 4, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the second nick gRNA sequence.


76. The gene modifying system of embodiment 72, wherein the second strand-targeting gRNA comprises a sequence comprising the core nucleotides of the second nick gRNA sequence from Table 4 that corresponds to the gRNA spacer sequence of (i), or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto, and optionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the second nick gRNA sequence.


77. The gene modifying system of any one of the preceding embodiments, wherein the second strand-targeting gRNA has a “PAM-in orientation” with the template RNA of the gene modifying system, e.g., as exemplified in Table 4.


78. The gene modifying system of any one of the preceding embodiments, the second strand-targeting gRNA targets a sequence overlapping the target mutation of the template RNA.


79. The gene modifying system of embodiment 78, wherein second strand-targeting gRNA comprises:

    • (i) a sequence (e.g., a spacer sequence) complementary to the SERPINA1 mutation;
    • (ii) a sequence (e.g., a spacer sequence) complementary to the wild-type sequence at the target locus;
    • (iii) a sequence (e.g., a spacer sequence) complementary to a SNP proximal to the target locus, e.g., a SNP contained in the genomic DNA of a subject (e.g., a patient);
    • (iv) a sequence (e.g., spacer sequence) complementary to or comprising one or more silent substitutions proximal to the target locus.


80. The template RNA, gene modifying system, or gRNA, of any one of the preceding embodiments, wherein the gRNA spacer comprises about 1, 2, 3, or more flanking nucleotides of the gRNA spacer.


81. The template RNA or gene modifying system of any one of the preceding embodiments, wherein the heterologous object sequence comprises about 2, 3, 4, 5, 10, 20, 30, 40, or more flanking nucleotides of the RT template sequence.


82. The template RNA or gene modifying system, of any one of the preceding embodiments, wherein the heterologous object sequence comprises between about 8-30, 9-25, 10-20, 11-16, or 12-15 (e.g., about 11-16) nucleotides.


83. The template RNA or gene modifying system, of any one of the preceding embodiments, wherein the mutation region comprises 1, 2, or 3 nucleotide positions of sequence differences relative to the corresponding portion of the human SERPINA1 gene.


84. The template RNA or gene modifying system of any one of the preceding embodiments, wherein the mutation region comprises at least 2 nucleotide positions of sequence difference relative to the corresponding portion of the human SERPINA1 gene.


85. The template RNA or gene modifying system, of any one of the preceding embodiments, wherein the post-edit homology region and/or pre-edit homology region comprises 100% identity to the SERPINA1 gene.


86. The template RNA or gene modifying system of any one of the preceding embodiments, wherein the PBS sequence additionally comprises about 1, 2, 3, 4, 5, 6, 7, or more flanking nucleotides.


87. The template RNA or gene modifying system of any one of the preceding embodiments, wherein the PBS sequence comprises about 5-20, 8-16, 8-14, 8-13, 9-13, 9-12, or 10-12 (e.g., about 9-12) nucleotides.


88. The template RNA or gene modifying system of any one of the preceding embodiments, wherein the PBS sequence binds within 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides of a nick site in the SERPINA1 gene.


89. The gene modifying system of any one of the preceding embodiments, wherein the domains of the gene modifying polypeptide are joined by a peptide linker.


90. The gene modifying system of embodiment 89, wherein the linker comprises a sequence of a linker of Table 10 (e.g., of any of SEQ ID NOs: 5217, 5106, 5190, and 5218).


91. The gene modifying system of any one of the preceding embodiments, wherein the gene modifying polypeptide further comprise one or more nuclear localization sequences (NLS).


92. The gene modifying system of embodiment 91, wherein the gene modifying polypeptide comprises a first NLS and a second NLS.


93. The gene modifying system of embodiment 91 or 92, wherein the NLS comprises a sequence of a NLS of Table 11 (e.g., of any of SEQ ID NOs: 5245, 5290, 5323, 5330, 5349, 5350, 5351, and 4001).


94. A template RNA comprising a sequence of a template RNA of Table 4, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


95. A template RNA comprising a sequence of a template RNA of Table 4.


96. A gene modifying system comprising:

    • (i) a template RNA comprising a sequence of a template RNA of Table 4, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto; and
    • (ii) a second-nick gRNA sequence from the same row of Table 4 as (i), a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


97. A gene modifying system comprising:

    • (i) a template RNA comprising a sequence of a template RNA of Table 4; and
    • (ii) a second-nick gRNA sequence from the same row of Table 4 as (i).


98. A DNA encoding the template RNA of any one of embodiments 1-20, 38-48, 80-88, 94, or 95, or the gRNA of any one of embodiments 35-37.


99. A pharmaceutical composition, comprising the system of any one of embodiments 52-93, 96, or 97, or one or more nucleic acids encoding the same, and a pharmaceutically acceptable excipient or carrier.


100. The pharmaceutical composition of embodiment 99, wherein the pharmaceutically acceptable excipient or carrier is selected from the group consisting of a plasmid vector, a viral vector, a vesicle, and a lipid nanoparticle.


101. The pharmaceutical composition of embodiment 100, wherein the viral vector is an adeno-associated virus.


102. A host cell (e.g., a mammalian cell, e.g., a human cell) comprising the template RNA or gene modifying system of any one of the preceding embodiments.


103. A method of making the template RNA of any one of embodiments 1-20, 38-48, 80-88, 94, or 953, the method comprising synthesizing the template RNA by in vitro transcription (e.g., solid state synthesis) or by introducing a DNA encoding the template RNA into a host cell under conditions that allow for production of the template RNA.


104. A method for modifying a target site in the human SERPINA1 gene in a cell, the method comprising contacting the cell with the gene modifying system of any one of embodiments 52-93, 96, or 97, or DNA encoding the same, thereby modifying the target site in the human SERPINA1 gene in a cell.


105. A method for modifying a target site in the human SERPINA1 gene in a cell, the method comprising contacting the cell with: (i) the template RNA of any one of embodiments 52-93, 96, or 97, or DNA encoding the same; and (ii) a gene modifying polypeptide or a nucleic acid encoding a gene modifying polypeptide, thereby modifying the target site in the human SERPINA1 gene in a cell.


106. A method for treating a subject having a disease or condition associated with a mutation in the human SERPINA1 gene, the method comprising administering to the subject the gene modifying system of any one of embodiments 52-93, 96, or 97, or DNA encoding the same, thereby treating the subject having a disease or condition associated with a mutation in the human SERPINA1 gene.


107. A method for treating a subject having a disease or condition associated with a mutation in the human SERPINA1 gene, the method comprising administering to the subject the template RNA of any one of embodiments 52-93, 96, or 97, or DNA encoding the same; and (ii) a gene modifying polypeptide or a nucleic acid encoding a gene modifying polypeptide, thereby treating the subject having a disease or condition associated with a mutation in the human SERPINA1 gene.


108. The method of embodiment 106 or 107, wherein the disease or condition is alpha-1 antitrypsin deficiency (AATD).


109. The method of any one of embodiments 106-108, wherein the subject has an E342K mutation (i.e., a PiZ mutation).


110. A method for treating a subject having AATD the method comprising administering to the subject the gene modifying system of any one of embodiments 52-93, 96, or 97, or DNA encoding the same, thereby treating the subject having AATD.


111. A method for treating a subject having AATD the method comprising administering to the subject (i) the template RNA of any one of embodiments 52-93, 96, or 97, or DNA encoding the same, and (ii) a gene modifying polypeptide or a nucleic acid encoding a gene modifying polypeptide, thereby treating the subject having AATD.


112. The gene modifying system or method of any one of the preceding embodiments, wherein introduction of the system into a target cell results in a correction of a pathogenic mutation in the SERPINA1 gene.


113. The gene modifying system or method of any one of the preceding embodiments, wherein the pathogenic mutation is a E342K mutation, and wherein the correction comprises an amino acid substitution of K342E.


114. The gene modifying system or method of any of the preceding embodiments, wherein correction of the mutation occurs in at least 30% (e.g., 30%, 40%, 50%, 60%, 70%, or more) of target nucleic acids.


115. The gene modifying system or method of any of the preceding embodiments, wherein correction of the mutation occurs in at least 30% (e.g., 30%, 40%, 50%, 60%, 70%, or more) of target cells.


116. The gene modifying system or method of any of the preceding embodiments, wherein the gene modifying system comprises a second strand-targeting gRNA, and wherein correction of the mutation in a population of target cells is increased relative to a population of target cells treated with a gene modifying system comprising a template RNA without a second strand-targeting gRNA.


117. The gene modifying system or method of any of the preceding embodiments, wherein the template RNA comprises one or more silent substitutions (e.g., as exemplified in Tables 7B), and wherein correction of the mutation in a population of target cells is increased relative to a population of target cells treated with a gene modifying system comprising a template RNA that does not comprise one or more silent substitutions.


118. The method of any of the preceding embodiments, wherein the cell is a mammalian cell, such as a human cell.


119. The method of any one of the preceding embodiments, wherein the subject is a human.


120. The method of any of the preceding embodiments, wherein the contacting occurs ex vivo, e.g., wherein the cell's or subject's DNA is modified ex vivo.


121. The method of any of the preceding embodiments, wherein the contacting occurs in vivo, e.g., wherein the cell's or subject's DNA is modified in vivo.


122. The method of any of the preceding embodiments, wherein contacting the cell or the subject with the system comprises contacting the cell or a cell within the subject with a nucleic acid (e.g., DNA or RNA) encoding the gene modifying polypeptide under conditions that allow for production of the gene modifying polypeptide.





BRIEF DESCRIPTION OF THE DRAWINGS

The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.



FIG. 1 depicts a gene modifying system as described herein. The left hand diagram shows the gene modifying polypeptide, which comprises a Cas nickase domain (e.g., spCas9 N863A) and a reverse transcriptase domain (RT domain) which are linked by a linker. The right hand diagram shows the template RNA which comprises, from 5′ to 3′, a gRNA spacer, a gRNA scaffold, a heterologous object sequence, and a primer binding site sequence (PBS sequence). The heterologous object sequence can comprise a mutation region that comprises one or more sequence differences relative to the target site. The heterologous object sequence can also comprise a pre-edit homology region and a post-edit homology region, which flank the mutation region. Without wishing to be bound by theory, it is thought that the gRNA spacer of the template RNA binds to the second strand of a target site in the genome, and the gRNA scaffold of the template RNA binds to the gene modifying polypeptide, e.g., localizing the gene modifying polypeptide to the target site in the genome. It is thought that the Cas domain of the gene modifying polypeptide nicks the target site (e.g., the first strand of the target site), e.g., allowing the PBS sequence to bind to a sequence adjacent to the site to be altered on the first strand of the target site. It is thought that the RT domain of the gene modifying polypeptide uses the first strand of the target site that is bound to the complementary sequence comprising the PBS sequence of the template RNA as a primer and the heterologous object sequence of the template RNA as a template to, e.g., polymerize a sequence complementary to the heterologous object sequence. Without wishing to be bound by theory, it is thought that reverse transcription can then proceed through the pre-edit homology region, then through the mutation region, and then through the post-edit homology region, thereby producing a DNA strand comprising a mutation specified by the heterologous object sequence.



FIG. 2 is a graph showing the percent rewriting achieved using the RNAV209-013 or RNAV214-040 gene modifying polypeptides with the indicated template RNAs.



FIG. 3 is a graph showing the amount of Fah mRNA relative to wild type when template RNAs are used with the RNAV209-013 or RNAV214-040 gene modifying polypeptides.



FIG. 4 is a graph showing the percentage of Cas9-positive hepatocytes 6 hours following dosing with LNPs containing various gene modifying polypeptides and template RNAs.



FIG. 5 is a graph showing the rewrite levels in liver samples 6 days following dosing with LNPs containing various gene modifying polypeptides and template RNAs.



FIG. 6 is a graph showing wild type Fah mRNA restoration compared to littermate heterozygous mice in liver samples following dosing with LNPs containing various gene modifying polypeptides and template RNAs.



FIG. 7 is a graph showing Fah protein distribution in liver samples following dosing with LNPs containing various gene modifying polypeptides and template RNAs.



FIG. 8 is a series of western blots showing Cas9-RT Expression 6 hours after infusion of Cas9-RT mRNA+TTR guide LNP. Each lane represents an individual animal where 20 μg of tissue homogenate was added per lane. Positive control was from an in vitro cell experiment where Cas9-RT was expressed (described previously). GAPDH was used as a loading control for each sample. n=4 per group, vehicle or treated.



FIG. 9 is a graph showing gene editing of TTR locus after treatment with Cas9-RT mRNA+TTR guide LNP. Level of indels detected at the TTR locus measured by TIDE analysis of Sanger sequencing of the TTR locus where the protospacer targets.



FIG. 10 is a graph showing that TTR Serum levels decrease after treatment with Cas9-RT mRNA+TTR guide LNP. Measurement of circulating TTR levels 5 days after mice were treated with LNPs encapsulating Cas9-RT+TTR guide RNA.



FIG. 11 is a graph showing Cas9-RT Expression after infusion of Cas9-RT mRNA+TTR guide LNP. Relative expression quantified by ProteinSimple Jess capillary electrophoresis Western blot. Numbers in the symbols are animal number in group. Vehicle n=2, Cas9-RT+TTR guide n=3.



FIG. 12 is a graph showing gene editing of TTR locus after infusion of Cas9-RT mRNA+TTR guide LNP. Level of indels detected at the TTR locus were measured by amplicon sequencing of the TTR locus where the protospacer targets. Each animal had 8 different biopsies taken across the liver where amplicon sequencing measured the percentage of reads showing an indel.



FIG. 13 is a graph showing percent indel activity of various gene modifying systems comprising template RNAs comprising 5 SpCas9 spacers, in combination with wild type SpCas9 polypeptide evaluated in HEK293T cells.



FIG. 14 is a graph showing percent indel at the PiZ mutation site in HEK293T landing pad cells after treatment with the gene modifying systems.



FIG. 15 is a graph showing a ranking of active spacer by indel activity and distance from the PiZ mutation following screening evaluation in HEK293T cells.



FIG. 16 is a graph showing percent perfect rewrite activity for various gene modifying systems comprising template RNAs.



FIGS. 17A-17B are heat maps graphing the % rewriting of gene modifying systems comprising various SpRY EDO template RNAs (varying PBS and RT lengths) and an exemplary SpRY Cas9-containing gene modifying polypeptide (FIG. 17A) and gene modifying systems comprising various St1_ED4 template RNAs (varying PBS and RT lengths) and an exemplary St1Cas9-containing gene modifying polypeptide (FIG. 17B).



FIG. 18 is a graph showing top-performing 17 combinations of template RNAs and gene modifying polypeptides comprising Cas9 variants (as ranked by rewriting activity).





DETAILED DESCRIPTION
Definitions

The term “expression cassette,” as used herein, refers to a nucleic acid construct comprising, nucleic acid elements sufficient for the expression of the nucleic acid molecule of the instant invention.


A “gRNA spacer”, as used herein, refers to a portion of a nucleic acid that has complementarity to a target nucleic acid and can, together with a gRNA scaffold, target a Cas protein to the target nucleic acid.


A “gRNA scaffold”, as used herein, refers to a portion of a nucleic acid that can bind a Cas protein and can, together with a gRNA spacer, target the Cas protein to the target nucleic acid. In some embodiments, the gRNA scaffold comprises a crRNA sequence, tetraloop, and tracrRNA sequence.


A “gene modifying polypeptide”, as used herein, refers to a polypeptide comprising a retroviral reverse transcriptase, or a polypeptide comprising an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% amino acid sequence identity to a retroviral reverse transcriptase, which is capable of integrating a nucleic acid sequence (e.g., a sequence provided on a template nucleic acid) into a target DNA molecule (e.g., in a mammalian host cell, such as a genomic DNA molecule in the host cell). In some embodiments, the gene modifying polypeptide is capable of integrating the sequence substantially without relying on host machinery. In some embodiments, the gene modifying polypeptide integrates a sequence into a random position in a genome, and in some embodiments, the gene modifying polypeptide integrates a sequence into a specific target site. In some embodiments, a gene modifying polypeptide includes one or more domains that, collectively, facilitate 1) binding the template nucleic acid, 2) binding the target DNA molecule, and 3) facilitate integration of the at least a portion of the template nucleic acid into the target DNA. Gene modifying polypeptides include both naturally occurring polypeptides as well as engineered variants of the foregoing, e.g., having one or more amino acid substitutions to the naturally occurring sequence. Gene modifying polypeptides also include heterologous constructs, e.g., where one or more of the domains recited above are heterologous to each other, whether through a heterologous fusion (or other conjugate) of otherwise wild-type domains, as well as fusions of modified domains, e.g., by way of replacement or fusion of a heterologous sub-domain or other substituted domain. Exemplary gene modifying polypeptides, and systems comprising them and methods of using them, that can be used in the methods provided herein are described, e.g., in PCT/US2021/020948, which is incorporated herein by reference with respect to gene modifying polypeptides that comprise a retroviral reverse transcriptase domain. In some embodiments, a gene modifying polypeptide integrates a sequence into a gene. In some embodiments, a gene modifying polypeptide integrates a sequence into a sequence outside of a gene. A “gene modifying system,” as used herein, refers to a system comprising a gene modifying polypeptide and a template nucleic acid.


The term “domain” as used herein refers to a structure of a biomolecule that contributes to a specified function of the biomolecule. A domain may comprise a contiguous region (e.g., a contiguous sequence) or distinct, non-contiguous regions (e.g., non-contiguous sequences) of a biomolecule. Examples of protein domains include, but are not limited to, an endonuclease domain, a DNA binding domain, a reverse transcription domain; an example of a domain of a nucleic acid is a regulatory domain, such as a transcription factor binding domain. In some embodiments, a domain (e.g., a Cas domain) can comprise two or more smaller domains (e.g., a DNA binding domain and an endonuclease domain).


As used herein, the term “exogenous”, when used with reference to a biomolecule (such as a nucleic acid sequence or polypeptide) means that the biomolecule was introduced into a host genome, cell or organism by the hand of man. For example, a nucleic acid that is as added into an existing genome, cell, tissue or subject using recombinant DNA techniques or other methods is exogenous to the existing nucleic acid sequence, cell, tissue or subject.


As used herein, “first strand” and “second strand”, as used to describe the individual DNA strands of target DNA, distinguish the two DNA strands based upon which strand the reverse transcriptase domain initiates polymerization, e.g., based upon where target primed synthesis initiates. The first strand refers to the strand of the target DNA upon which the reverse transcriptase domain initiates polymerization, e.g., where target primed synthesis initiates. The second strand refers to the other strand of the target DNA. First and second strand designations do not describe the target site DNA strands in other respects; for example, in some embodiments the first and second strands are nicked by a polypeptide described herein, but the designations ‘first’ and ‘second’ strand have no bearing on the order in which such nicks occur.


The term “heterologous,” as used herein to describe a first element in reference to a second element means that the first element and second element do not exist in nature disposed as described. For example, a heterologous polypeptide, nucleic acid molecule, construct or sequence refers to (a) a polypeptide, nucleic acid molecule or portion of a polypeptide or nucleic acid molecule sequence that is not native to a cell in which it is expressed, (b) a polypeptide or nucleic acid molecule or portion of a polypeptide or nucleic acid molecule that has been altered or mutated relative to its native state, or (c) a polypeptide or nucleic acid molecule with an altered expression as compared to the native expression levels under similar conditions. For example, a heterologous regulatory sequence (e.g., promoter, enhancer) may be used to regulate expression of a gene or a nucleic acid molecule in a way that is different than the gene or a nucleic acid molecule is normally expressed in nature. In another example, a heterologous domain of a polypeptide or nucleic acid sequence (e.g., a DNA binding domain of a polypeptide or nucleic acid encoding a DNA binding domain of a polypeptide) may be disposed relative to other domains or may be a different sequence or from a different source, relative to other domains or portions of a polypeptide or its encoding nucleic acid. In certain embodiments, a heterologous nucleic acid molecule may exist in a native host cell genome, but may have an altered expression level or have a different sequence or both. In other embodiments, heterologous nucleic acid molecules may not be endogenous to a host cell or host genome but instead may have been introduced into a host cell by transformation (e.g., transfection, electroporation), wherein the added molecule may integrate into the host genome or can exist as extra-chromosomal genetic material either transiently (e.g., mRNA) or semi-stably for more than one generation (e.g., episomal viral vector, plasmid or other self-replicating vector).


As used herein, “insertion” of a sequence into a target site refers to the net addition of DNA sequence at the target site, e.g., where there are new nucleotides in the heterologous object sequence with no cognate positions in the unedited target site. In some embodiments, a nucleotide alignment of the PBS sequence and heterologous object sequence to the target nucleic acid sequence would result in an alignment gap in the target nucleic acid sequence.


As used herein, a “deletion” generated by a heterologous object sequence in a target site refers to the net deletion of DNA sequence at the target site, e.g., where there are nucleotides in the unedited target site with no cognate positions in the heterologous object sequence. In some embodiments, a nucleotide alignment of the PBS sequence and heterologous object sequence to the target nucleic acid sequence would result in an alignment gap in the molecule comprising the PBS sequence and heterologous object sequence.


The term “inverted terminal repeats” or “ITRs” as used herein refers to AAV viral cis-elements named so because of their symmetry. These elements promote efficient multiplication of an AAV genome. It is hypothesized that the minimal elements for ITR function are a Rep-binding site (RBS; 5′-GCGCGCTCGCTCGCTC-3′ for AAV2; SEQ ID NO: 4601) and a terminal resolution site (TRS; 5′-AGTTGG-3′ for AAV2) plus a variable palindromic sequence allowing for hairpin formation. According to the present invention, an ITR comprises at least these three elements (RBS, TRS, and sequences allowing the formation of an hairpin). In addition, in the present invention, the term “ITR” refers to ITRs of known natural AAV serotypes (e.g. ITR of a serotype 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11 AAV), to chimeric ITRs formed by the fusion of ITR elements derived from different serotypes, and to functional variants thereof. “Functional variant” refers to a sequence presenting a sequence identity of at least 80%, 85%, 90%, preferably of at least 95% with a known ITR and allowing multiplication of the sequence that includes said ITR in the presence of Rep proteins.


The term “mutation region,” as used herein, refers to a region in a template RNA having one or more sequence difference relative to the corresponding sequence in a target nucleic acid. The sequence difference may comprise, for example, a substitution, insertion, frameshift, or deletion.


The term “mutated” when applied to nucleic acid sequences means that nucleotides in a nucleic acid sequence are inserted, deleted, or changed compared to a reference (e.g., native) nucleic acid sequence. A single alteration may be made at a locus (a point mutation), or multiple nucleotides may be inserted, deleted, or changed at a single locus. In addition, one or more alterations may be made at any number of loci within a nucleic acid sequence. A nucleic acid sequence may be mutated by any method known in the art.


“Nucleic acid molecule” refers to both RNA and DNA molecules including, without limitation, complementary DNA (“cDNA”), genomic DNA (“gDNA”), and messenger RNA (“mRNA”), and also includes synthetic nucleic acid molecules, such as those that are chemically synthesized or recombinantly produced, such as RNA templates, as described herein. The nucleic acid molecule can be double-stranded or single-stranded, circular, or linear. If single-stranded, the nucleic acid molecule can be the sense strand or the antisense strand. Unless otherwise indicated, and as an example for all sequences described herein under the general format “SEQ ID NO:,” or “nucleic acid comprising SEQ ID NO:1” refers to a nucleic acid, at least a portion which has either (i) the sequence of SEQ ID NO:1, or (ii) a sequence complimentary to SEQ ID NO:1. The choice between the two is dictated by the context in which SEQ ID NO:1 is used. For instance, if the nucleic acid is used as a probe, the choice between the two is dictated by the requirement that the probe be complementary to the desired target. Nucleic acid sequences of the present disclosure may be modified chemically or biochemically or may contain non-natural or derivatized nucleotide bases, as will be readily appreciated by those of skill in the art. Such modifications include, for example, labels, methylation, substitution of one or more naturally occurring nucleotides with an analog, inter-nucleotide modifications such as uncharged linkages (for example, methyl phosphonates, phosphotriesters, phosphoramidates, carbamates, etc.), charged linkages (for example, phosphorothioates, phosphorodithioates, etc.), pendant moieties, (for example, polypeptides), intercalators (for example, acridine, psoralen, etc.), chelators, alkylators, and modified linkages (for example, alpha anomeric nucleic acids, etc.). Also included are chemically modified bases (see, for example, Table 13), backbones (see, for example, Table 14), and modified caps (see, for example, Table 15). Also included are synthetic molecules that mimic polynucleotides in their ability to bind to a designated sequence via hydrogen bonding and other chemical interactions. Such molecules are known in the art and include, for example, those in which peptide linkages substitute for phosphate linkages in the backbone of a molecule, e.g., peptide nucleic acids (PNAs). Other modifications can include, for example, analogs in which the ribose ring contains a bridging moiety or other structure such as modifications found in “locked” nucleic acids (LNAs). In various embodiments, the nucleic acids are in operative association with additional genetic elements, such as tissue-specific expression-control sequence(s) (e.g., tissue-specific promoters and tissue-specific microRNA recognition sequences), as well as additional elements, such as inverted repeats (e.g., inverted terminal repeats, such as elements from or derived from viruses, e.g., AAV ITRs) and tandem repeats, inverted repeats/direct repeats, homology regions (segments with various degrees of homology to a target DNA), untranslated regions (UTRs) (5′, 3′, or both 5′ and 3′ UTRs), and various combinations of the foregoing. The nucleic acid elements of the systems provided by the invention can be provided in a variety of topologies, including single-stranded, double-stranded, circular, linear, linear with open ends, linear with closed ends, and particular versions of these, such as doggybone DNA (dbDNA), closed-ended DNA (ceDNA).


As used herein, a “gene expression unit” is a nucleic acid sequence comprising at least one regulatory nucleic acid sequence operably linked to at least one effector sequence. A first nucleic acid sequence is operably linked with a second nucleic acid sequence when the first nucleic acid sequence is placed in a functional relationship with the second nucleic acid sequence. For instance, a promoter or enhancer is operably linked to a coding sequence if the promoter or enhancer affects the transcription or expression of the coding sequence. Operably linked DNA sequences may be contiguous or non-contiguous. Where necessary to join two protein-coding regions, operably linked sequences may be in the same reading frame.


The terms “host genome” or “host cell”, as used herein, refer to a cell and/or its genome into which protein and/or genetic material has been introduced. It should be understood that such terms are intended to refer not only to the particular subject cell and/or genome, but to the progeny of such a cell and/or the genome of the progeny of such a cell. Because certain modifications may occur in succeeding generations due to either mutation or environmental influences, such progeny may not, in fact, be identical to the parent cell, but are still included within the scope of the term “host cell” as used herein. A host genome or host cell may be an isolated cell or cell line grown in culture, or genomic material isolated from such a cell or cell line, or may be a host cell or host genome which composing living tissue or an organism. In some instances, a host cell may be an animal cell or a plant cell, e.g., as described herein. In certain instances, a host cell may be a mammalian cell, a human cell, avian cell, reptilian cell, bovine cell, horse cell, pig cell, goat cell, sheep cell, chicken cell, or turkey cell. In certain instances, a host cell may be a corn cell, soy cell, wheat cell, or rice cell.


As used herein, “operative association” describes a functional relationship between two nucleic acid sequences, such as a 1) promoter and 2) a heterologous object sequence, and means, in such example, the promoter and heterologous object sequence (e.g., a gene of interest) are oriented such that, under suitable conditions, the promoter drives expression of the heterologous object sequence. For instance, a template nucleic acid carrying a promoter and a heterologous object sequence may be single-stranded, e.g., either the (+) or (−) orientation. An “operative association” between the promoter and the heterologous object sequence in this template means that, regardless of whether the template nucleic acid will be transcribed in a particular state, when it is in the suitable state (e.g., is in the (+) orientation, in the presence of required catalytic factors, and NTPs, etc.), it is accurately transcribed. Operative association applies analogously to other pairs of nucleic acids, including other tissue-specific expression control sequences (such as enhancers, repressors and microRNA recognition sequences), IR/DR, ITRs, UTRs, or homology regions and heterologous object sequences or sequences encoding a retroviral RT domain.


The term “primer binding site sequence” or “PBS sequence,” as used herein, refers to a portion of a template RNA capable of binding to a region comprised in a target nucleic acid sequence. In some instances, a PBS sequence is a nucleic acid sequence comprising at least 3, 4, 5, 6, 7, or 8 bases with 100% identity to the region comprised in the target nucleic acid sequence. In some embodiments the primer region comprises at least 5, 6, 7, 8 bases with 100% identity to the region comprised in the target nucleic acid sequence. Without wishing to be bound by theory, in some embodiments when a template RNA comprises a PBS sequence and a heterologous object sequence, the PBS sequence binds to a region comprised in a target nucleic acid sequence, allowing a reverse transcriptase domain to use that region as a primer for reverse transcription, and to use the heterologous object sequence as a template for reverse transcription.


As used herein, a “stem-loop sequence” refers to a nucleic acid sequence (e.g., RNA sequence) with sufficient self-complementarity to form a stem-loop, e.g., having a stem comprising at least two (e.g., 3, 4, 5, 6, 7, 8, 9, or 10) base pairs, and a loop with at least three (e.g., four) base pairs. The stem may comprise mismatches or bulges.


As used herein, a “tissue-specific expression-control sequence” means nucleic acid elements that increase or decrease the level of a transcript comprising the heterologous object sequence in a target tissue in a tissue-specific manner, e.g., preferentially in on-target tissue(s), relative to off-target tissue(s). In some embodiments, a tissue-specific expression-control sequence preferentially drives or represses transcription, activity, or the half-life of a transcript comprising the heterologous object sequence in the target tissue in a tissue-specific manner, e.g., preferentially in an on-target tissue(s), relative to an off-target tissue(s). Exemplary tissue-specific expression-control sequences include tissue-specific promoters, repressors, enhancers, or combinations thereof, as well as tissue-specific microRNA recognition sequences. Tissue specificity refers to on-target (tissue(s) where expression or activity of the template nucleic acid is desired or tolerable) and off-target (tissue(s) where expression or activity of the template nucleic acid is not desired or is not tolerable). For example, a tissue-specific promoter drives expression preferentially in on-target tissues, relative to off-target tissues. In contrast, a microRNA that binds the tissue-specific microRNA recognition sequences is preferentially expressed in off-target tissues, relative to on-target tissues, thereby reducing expression of a template nucleic acid in off-target tissues. Accordingly, a promoter and a microRNA recognition sequence that are specific for the same tissue, such as the target tissue, have contrasting functions (promote and repress, respectively, with concordant expression levels, i.e., high levels of the microRNA in off-target tissues and low levels in on-target tissues, while promoters drive high expression in on-target tissues and low expression in off-target tissues) with regard to the transcription, activity, or half-life of an associated sequence in that tissue.


Table of Contents

1) Introduction


2) Gene modifying systems

    • a) Polypeptide components of gene modifying systems
      • i) Writing domain
      • ii) Endonuclease domains and DNA binding domains
        • (1) Gene modifying polypeptides comprising Cas domains
        • (2) TAL Effectors and Zinc Finger Nucleases
      • iii) Linkers
      • iv) Localization sequences for gene modifying systems
      • v) Evolved Variants of Gene Modifying Polypeptides and Systems
      • vi) Inteins
      • vii) Additional domains
    • b) Template nucleic acids
      • i) gRNA spacer and gRNA scaffold
      • ii) Heterologous object sequence
      • iii) PBS sequence
      • iv) Exemplary Template Sequences
    • c) gRNAs with inducible activity
    • d) Circular RNAs and Ribozymes in Gene Modifying Systems
    • e) Target Nucleic Acid Site
    • f) Second strand nicking


3) Production of Compositions and Systems


4) Therapeutic Applications


5) Administration and Delivery

    • a) Tissue Specific Activity/Administration
      • i) Promoters
      • ii) microRNAs
    • b) Viral vectors and components thereof
    • c) AAV Administration
    • d) Lipid Nanoparticles


6) Kits, Articles of Manufacture, and Pharmaceutical Compositions


7) Chemistry, Manufacturing, and Controls (CMC)


INTRODUCTION

This disclosure relates to methods for treating alpha-1 antitrypsin deficiency (AATD) and compositions for targeting, editing, modifying or manipulating a DNA sequence (e.g., inserting a heterologous object sequence into a target site of a mammalian genome) at one or more locations in a DNA sequence in a cell, tissue or subject, e.g., in vivo or in vitro. The heterologous object DNA sequence may include, e.g., a substitution.


More specifically, the disclosure provides methods for treating AATD using reverse transcriptase-based systems for altering a genomic DNA sequence of interest, e.g., by inserting, deleting, or substituting one or more nucleotides into/from the sequence of interest.


The disclosure provides, in part, methods for treating AATD using a gene modifying system comprising a gene modifying polypeptide component and a template nucleic acid (e.g., template RNA) component. In some embodiments, a gene modifying system can be used to introduce an alteration into a target site in a genome. In some embodiments, the gene modifying polypeptide component comprises a writing domain (e.g., a reverse transcriptase domain), a DNA-binding domain, and an endonuclease domain (e.g., nickase domain). In some embodiments, the template nucleic acid (e.g., template RNA) comprises a sequence (e.g., a gRNA spacer) that binds a target site in the genome (e.g., that binds to a second strand of the target site), a sequence (e.g., a gRNA scaffold) that binds the gene modifying polypeptide component, a heterologous object sequence, and a PBS sequence. Without wishing to be bound by theory, it is thought that the template nucleic acid (e.g., template RNA) binds to the second strand of a target site in the genome, and binds to the gene modifying polypeptide component (e.g., localizing the polypeptide component to the target site in the genome). It is thought that the endonuclease (e.g., nickase) of the gene modifying polypeptide component cuts the target site (e.g., the first strand of the target site), e.g., allowing the PBS sequence to bind to a sequence adjacent to the site to be altered on the first strand of the target site. It is thought that the writing domain (e.g., reverse transcriptase domain) of the polypeptide component uses the first strand of the target site that is bound to the complementary sequence comprising the PBS sequence of the template nucleic acid as a primer and the heterologous object sequence of the template nucleic acid as a template to, e.g., polymerize a sequence complementary to the heterologous object sequence. Without wishing to be bound by theory, it is thought that selection of an appropriate heterologous object sequence can result in substitution, deletion, and/or insertion of one or more nucleotides at the target site.


Gene Modifying Systems

In some embodiments, a gene modifying system described herein comprises: (A) a gene modifying polypeptide or a nucleic acid encoding the gene modifying polypeptide, wherein the gene modifying polypeptide comprises (i) a reverse transcriptase domain, and either (x) an endonuclease domain that contains DNA binding functionality or (y) an endonuclease domain and separate DNA binding domain; and (B) a template RNA. A gene modifying polypeptide, in some embodiments, acts as a substantially autonomous protein machine capable of integrating a template nucleic acid sequence into a target DNA molecule (e.g., in a mammalian host cell, such as a genomic DNA molecule in the host cell), substantially without relying on host machinery. For example, the gene modifying protein may comprise a DNA-binding domain, a reverse transcriptase domain, and an endonuclease domain. In some embodiments, the DNA-binding function may involve an RNA component that directs the protein to a DNA sequence, e.g., a gRNA spacer. In other embodiments, the gene modifying polypeptide may comprise a reverse transcriptase domain and an endonuclease domain. The RNA template element of a gene modifying system is typically heterologous to the gene modifying polypeptide element and provides an object sequence to be inserted (reverse transcribed) into the host genome. In some embodiments, the gene modifying polypeptide is capable of target primed reverse transcription. In some embodiments, the gene modifying polypeptide is capable of second-strand synthesis.


In some embodiments the gene modifying system is combined with a second polypeptide. In some embodiments, the second polypeptide may comprise an endonuclease domain. In some embodiments, the second polypeptide may comprise a polymerase domain, e.g., a reverse transcriptase domain. In some embodiments, the second polypeptide may comprise a DNA-dependent DNA polymerase domain. In some embodiments, the second polypeptide aids in completion of the genome edit, e.g., by contributing to second-strand synthesis or DNA repair resolution.


A functional gene modifying polypeptide can be made up of unrelated DNA binding, reverse transcription, and endonuclease domains. This modular structure allows combining of functional domains, e.g., dCas9 (DNA binding), MMLV reverse transcriptase (reverse transcription), FokI (endonuclease). In some embodiments, multiple functional domains may arise from a single protein, e.g., Cas9 or Cas9 nickase (DNA binding, endonuclease). In some embodiments, a gene modifying polypeptide includes one or more domains that, collectively, facilitate 1) binding the template nucleic acid, 2) binding the target DNA molecule, and 3) facilitate integration of the at least a portion of the template nucleic acid into the target DNA. In some embodiments, the gene modifying polypeptide is an engineered polypeptide that comprises one or more amino acid substitutions to a corresponding naturally occurring sequence. In some embodiments, the gene modifying polypeptide comprises two or more domains that are heterologous relative to each other, e.g., through a heterologous fusion (or other conjugate) of otherwise wild-type domains, or well as fusions of modified domains, e.g., by way of replacement or fusion of a heterologous sub-domain or other substituted domain. For instance, in some embodiments, one or more of: the RT domain is heterologous to the DBD; the DBD is heterologous to the endonuclease domain; or the RT domain is heterologous to the endonuclease domain.


In some embodiments, a template RNA molecule for use in the system comprises, from 5′ to 3′ (1) a gRNA spacer; (2) a gRNA scaffold; (3) heterologous object sequence (4) a primer binding site (PBS) sequence. In some embodiments:

    • (1) Is a gRNA spacer of ˜18-22 nt, e.g., is 20 nt
    • (2) Is a gRNA scaffold comprising one or more hairpin loops, e.g., 1, 2, of 3 loops for associating the template with a Cas domain, e.g., a nickase Cas9 domain. In some embodiments, the gRNA scaffold comprises the sequence, from 5′ to 3′,











(SEQ ID NO: 5008)



GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAG






TCCGTTATCAACTTGAAAAAGTGGGACCGAGTCGGTCC.








    • (3) In some embodiments, the heterologous object sequence is, e.g., 7-74, e.g., 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, or 70-80 nt or, 80-90 nt in length. In some embodiments, the first (most 5′) base of the sequence is not C.

    • (4) In some embodiments, the PBS sequence that binds the target priming sequence after nicking occurs is e.g., 3-20 nt, e.g., 7-15 nt, e.g., 12-14 nt. In some embodiments, the PBS sequence has 40-60% GC content.





In some embodiments, a second gRNA associated with the system may help drive complete integration. In some embodiments, the second gRNA may target a location that is 0-200 nt away from the first-strand nick, e.g., 0-50, 50-100, 100-200 nt away from the first-strand nick. In some embodiments, the second gRNA can only bind its target sequence after the edit is made, e.g., the gRNA binds a sequence present in the heterologous object sequence, but not in the initial target sequence.


In some embodiments, a gene modifying system described herein is used to make an edit in HEK293, K562, U205, or HeLa cells. In some embodiment, a gene modifying system is used to make an edit in primary cells, e.g., primary cortical neurons from E18.5 mice.


In some embodiments, a gene modifying polypeptide as described herein comprises a reverse transcriptase or RT domain (e.g., as described herein) that comprises a MoMLV RT sequence or variant thereof. In embodiments, the MoMLV RT sequence comprises one or more mutations selected from D200N, L603W, T330P, T306K, W313F, D524G, E562Q, D583N, P51L, S67R, E67K, T197A, H204R, E302K, F309N, L435G, N454K, H594Q, D653N, R110S, and K103L. In embodiments, the MoMLV RT sequence comprises a combination of mutations, such as D200N, L603W, and T330P, optionally further including T306K and/or W313F.


In some embodiments, an endonuclease domain (e.g., as described herein) nCas9, e.g., comprising an N863A mutation (e.g., in spCas9) or a H840A mutation.


In some embodiments, the heterologous object sequence (e.g., of a system as described herein) is about 1-50, 50-100, 100-200, 200-300, 300-400, 400-500, 500-600, 600-700, 700-800, 800-900, 900-1000, or more, nucleotides in length.


In some embodiments, the RT and endonuclease domains are joined by a flexible linker, e.g., comprising the amino acid sequence SGGSSGGSSGSETPGTSESATPESSGGSSGGSS (SEQ ID NO: 5006).


In some embodiments, the endonuclease domain is N-terminal relative to the RT domain. In some embodiments, the endonuclease domain is C-terminal relative to the RT domain.


In some embodiments, the system incorporates a heterologous object sequence into a target site by TPRT, e.g., as described herein.


In some embodiments, a gene modifying polypeptide comprises a DNA binding domain. In some embodiments, a gene modifying polypeptide comprises an RNA binding domain. In some embodiments, the RNA binding domain comprises an RNA binding domain of B-box protein, MS2 coat protein, dCas, or an element of a sequence of a table herein. In some embodiments, the RNA binding domain is capable of binding to a template RNA with greater affinity than a reference RNA binding domain.


In some embodiments, a gene modifying system is capable of producing an insertion into the target site of at least 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 nucleotides (and optionally no more than 500, 400, 300, 200, or 100 nucleotides). In some embodiments, a gene modifying system is capable of producing an insertion into the target site of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 nucleotides (and optionally no more than 500, 400, 300, 200, or 100 nucleotides). In some embodiments, a gene modifying system is capable of producing an insertion into the target site of at least 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5 or 10 kilobases (and optionally no more than 1, 5, 10, or 20 kilobases). In some embodiments, a gene modifying system is capable of producing a deletion of at least 81, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200 nucleotides (and optionally no more than 500, 400, 300, or 200 nucleotides). In some embodiments, a gene modifying system is capable of producing a deletion of at least 81, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200 nucleotides (and optionally no more than 500, 400, 300, or 200 nucleotides). In some embodiments, a gene modifying system is capable of producing a deletion of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200 nucleotides (and optionally no more than 500, 400, 300, or 200 nucleotides). In some embodiments, a gene modifying system is capable of producing a deletion of at least 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5 or 10 kilobases (and optionally no more than 1, 5, 10, or 20 kilobases). In some embodiments, a gene modifying system is capable of producing a substitution into the target site of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, or 100 or more nucleotides. In some embodiments, a gene modifying system is capable of producing a substitution in the target site of 1-2, 2-3, 3-4, 4-5, 5-10, 10-15, 15-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, 80-90, or 90-100 nucleotides.


In some embodiments, the substitution is a transition mutation. In some embodiments, the substitution is a transversion mutation. In some embodiments, the substitution converts an adenine to a thymine, an adenine to a guanine, an adenine to a cytosine, a guanine to a thymine, a guanine to a cytosine, a guanine to an adenine, a thymine to a cytosine, a thymine to an adenine, a thymine to a guanine, a cytosine to an adenine, a cytosine to a guanine, or a cytosine to a thymine.


In some embodiments, an insertion, deletion, substitution, or combination thereof, increases or decreases expression (e.g. transcription or translation) of a gene. In some embodiments, an insertion, deletion, substitution, or combination thereof, increases or decreases expression (e.g. transcription or translation) of a gene by altering, adding, or deleting sequences in a promoter or enhancer, e.g. sequences that bind transcription factors. In some embodiments, an insertion, deletion, substitution, or combination thereof alters translation of a gene (e.g. alters an amino acid sequence), inserts or deletes a start or stop codon, alters or fixes the translation frame of a gene. In some embodiments, an insertion, deletion, substitution, or combination thereof alters splicing of a gene, e.g. by inserting, deleting, or altering a splice acceptor or donor site. In some embodiments, an insertion, deletion, substitution, or combination thereof alters transcript or protein half-life. In some embodiments, an insertion, deletion, substitution, or combination thereof alters protein localization in the cell (e.g. from the cytoplasm to a mitochondria, from the cytoplasm into the extracellular space (e.g. adds a secretion tag)). In some embodiments, an insertion, deletion, substitution, or combination thereof alters (e.g. improves) protein folding (e.g. to prevent accumulation of misfolded proteins). In some embodiments, an insertion, deletion, substitution, or combination thereof, alters, increases, decreases the activity of a gene, e.g. a protein encoded by the gene.


Exemplary gene modifying polypeptides, and systems comprising them and methods of using them are described, e.g., in PCT/US2021/020948, which is incorporated herein by reference with respect to retroviral RT domains, including the amino acid and nucleic acid sequences therein.


Exemplary gene modifying polypeptides and retroviral RT domain sequences are also described, e.g., in International Application No. PCT/US21/20948 filed Mar. 4, 2021, e.g., at Table 30, Table 31, and Table 44 therein; the entire application is incorporated by reference herein with respect to retroviral RTs, e.g., in said sequences and tables. Accordingly, a gene modifying polypeptide described herein may comprise an amino acid sequence according to any of the Tables mentioned in this paragraph, or a domain thereof (e.g., a retroviral RT domain), or a functional fragment or variant of any of the foregoing, or an amino acid sequence having at least 70%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In some embodiments, a polypeptide for use in any of the systems described herein can be a molecular reconstruction or ancestral reconstruction based upon the aligned polypeptide sequence of multiple homologous proteins. In some embodiments, a reverse transcriptase domain for use in any of the systems described herein can be a molecular reconstruction or an ancestral reconstruction, or can be modified at particular residues, based upon alignments of reverse transcriptase domains from the same or different sources. A skilled artisan can, based on the Accession numbers provided herein, align polypeptides or nucleic acid sequences, e.g., by using routine sequence analysis tools as Basic Local Alignment Search Tool (BLAST) or CD-Search for conserved domain analysis. Molecular reconstructions can be created based upon sequence consensus, e.g. using approaches described in Ivics et al., Cell 1997, 501-510; Wagstaff et al., Molecular Biology and Evolution 2013, 88-99.


Polypeptide Components of Gene Modifying Systems

In some embodiments, the gene modifying polypeptide possesses the functions of DNA target site binding, template nucleic acid (e.g., RNA) binding, DNA target site cleavage, and template nucleic acid (e.g., RNA) writing, e.g., reverse transcription. In some embodiments, each functions is contained within a distinct domain. In some embodiments, a function may be attributed to two or more domains (e.g., two or more domains, together, exhibit the functionality). In some embodiments, two or more domains may have the same or similar function (e.g., two or more domains each independently have DNA-binding functionality, e.g., for two different DNA sequences). In other embodiments, one or more domains may be capable of enabling one or more functions, e.g., a Cas9 domain enabling both DNA binding and target site cleavage. In some embodiments, the domains are all located within a single polypeptide. In some embodiments, a first domain is in one polypeptide and a second domain is in a second polypeptide. For example, in some embodiments, the sequences may be split between a first polypeptide and a second polypeptide, e.g., wherein the first polypeptide comprises a reverse transcriptase (RT) domain and wherein the second polypeptide comprises a DNA-binding domain and an endonuclease domain, e.g., a nickase domain. As a further example, in some embodiments, the first polypeptide and the second polypeptide each comprise a DNA binding domain (e.g., a first DNA binding domain and a second DNA binding domain). In some embodiments, the first and second polypeptide may be brought together post-translationally via a split-intein to form a single gene modifying polypeptide.


In some aspects, a gene modifying polypeptide described herein comprises (e.g., a system described herein comprises a gene modifying polypeptide that comprises): 1) a Cas domain (e.g., a Cas nickase domain, e.g., a Cas9 nickase domain); 2) a reverse transcriptase (RT) domain of Table D, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity thereto, wherein the RT domain is C-terminal of the Cas domain; and a linker disposed between the RT domain and the Cas domain, wherein the linker has a sequence from the same row of Table D as the RT domain, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity thereto.


In some embodiments, the RT domain has a sequence with 100% identity to the RT domain of Table D and the linker has a sequence with 100% identity to the linker sequence from the same row of Table D as the RT domain. In some embodiments, the Cas domain comprises a sequence of Table 8, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto. In some embodiments, the gene modifying polypeptide comprises an amino acid sequence according to any of SEQ ID NOs: 1-3332 in the sequence listing, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity thereto.


In some embodiments, the gene modifying polypeptide comprises a GG amino acid sequence between the Cas domain and the linker, an AG amino acid sequence between the RT domain and the second NLS, and/or a GG amino acid sequence between the linker and the RT domain. In some embodiments, the gene modifying polypeptide comprises a sequence of SEQ ID NO: 4000 which comprises the first NLS and the Cas domain, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto. In some embodiments, the gene modifying polypeptide comprises a sequence of SEQ ID NO: 4001 which comprises the second NLS, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99% identity thereto.











Exemplary N-terminal NLS-Cas9 domain



(SEQ ID NO: 4000)



MPAAKRVKLDGGDKKYSIGLDIGTNSVGWAVITDEYKVPSK






KFKVLGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARR






RYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEED






KKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKAD






LRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQ






TYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLP






GEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDT






YDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNT






EITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEI






FFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELL






VKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFY






PFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSE






ETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPK






HSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVD






LLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNAS






LGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDRE






MIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGI






RDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKEDIQK






AQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKV






MGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKE






LGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDI






NRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNV






PSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSE






LDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI






REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN






AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIG






KATAKYFFYSNIMNFFKTEITLANGEIRKRPLIETNGETG






EIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESI






LPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVE






KGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEV






KKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPS






KYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII






EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENII






HLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQ






SITGLYETRIDLSQLGGDGG






Exemplary C-terminal sequence comprising



an NLS



(SEQ ID NO: 4001)



AGKRTADGSEFEKRTADGSEFESPKKKAKVE






Writing Domain (RT Domain)


In certain aspects of the present invention, the writing domain of the gene modifying system possesses reverse transcriptase activity and is also referred to as a reverse transcriptase domain (a RT domain). In some embodiments, the RT domain comprises an RT catalytic portion and RNA-binding region (e.g., a region that binds the template RNA).


In some embodiments, a nucleic acid encoding the reverse transcriptase is altered from its natural sequence to have altered codon usage, e.g. improved for human cells. In some embodiments the reverse transcriptase domain is a heterologous reverse transcriptase from a retrovirus. In some embodiments, the RT domain comprising a gene modifying polypeptide has been mutated from its original amino acid sequence, e.g., has at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 substitutions. In some embodiments, the RT domain is derived from the RT of a retrovirus, e.g., HIV-1 RT, Moloney Murine Leukemia Virus (MMLV) RT, avian myeloblastosis virus (AMV) RT, or Rous Sarcoma Virus (RSV) RT.


In some embodiments, the retroviral reverse transcriptase (RT) domain exhibits enhanced stringency of target-primed reverse transcription (TPRT) initiation, e.g., relative to an endogenous RT domain. In some embodiments, the RT domain initiates TPRT when the 3 nt in the target site immediately upstream of the first strand nick, e.g., the genomic DNA priming the RNA template, have at least 66% or 100% complementarity to the 3 nt of homology in the RNA template. In some embodiments, the RT domain initiates TPRT when there are less than 5 nt mismatched (e.g., less than 1, 2, 3, 4, or 5 nt mismatched) between the template RNA homology and the target DNA priming reverse transcription. In some embodiments, the RT domain is modified such that the stringency for mismatches in priming the TPRT reaction is increased, e.g., wherein the RT domain does not tolerate any mismatches or tolerates fewer mismatches in the priming region relative to a wild-type (e.g., unmodified) RT domain. In some embodiments, the RT domain comprises a HIV-1 RT domain. In embodiments, the HIV-1 RT domain initiates lower levels of synthesis even with three nucleotide mismatches relative to an alternative RT domain (e.g., as described by Jamburuthugoda and Eickbush J Mol Biol 407(5):661-672 (2011); incorporated herein by reference in its entirety). In some embodiments, the RT domain forms a dimer (e.g., a heterodimer or homodimer). In some embodiments, the RT domain is monomeric. In some embodiments, an RT domain, naturally functions as a monomer or as a dimer (e.g., heterodimer or homodimer). In some embodiments, an RT domain naturally functions as a monomer, e.g., is derived from a virus wherein it functions as a monomer. In embodiments, the RT domain is selected from an RT domain from murine leukemia virus (MLV; sometimes referred to as MoMLV) (e.g., P03355), porcine endogenous retrovirus (PERV) (e.g., UniProt Q4VFZ2), mouse mammary tumor virus (MMTV) (e.g., UniProt P03365), Avian reticuloendotheliosis virus (AVIRE) (e.g., UniProtKB accession: P03360); Feline leukemia virus (FLV or FeLV) (e.g., e.g., UniProtKB accession: P10273); Mason-Pfizer monkey virus (MPMV) (e.g., UniProt P07572), bovine leukemia virus (BLV) (e.g., UniProt P03361), human T-cell leukemia virus-1 (HTLV-1) (e.g., UniProt P03362), human foamy virus (HFV) (e.g., UniProt P14350), simian foamy virus (SFV) (e.g., SFV3L) (e.g., UniProt P23074 or P27401), or bovine foamy/syncytial virus (BFV/BSV) (e.g., UniProt O41894), or a functional fragment or variant thereof (e.g., an amino acid sequence having at least 70%, 80%, 90%, 95%, or 99% identity thereto). In some embodiments, an RT domain is dimeric in its natural functioning. In some embodiments, the RT domain is derived from a virus wherein it functions as a dimer. In embodiments, the RT domain is selected from an RT domain from avian sarcoma/leukemia virus (ASLV) (e.g., UniProt A0A142BKH1), Rous sarcoma virus (RSV) (e.g., UniProt P03354), avian myeloblastosis virus (AMV) (e.g., UniProt Q83133), human immunodeficiency virus type I (HIV-1) (e.g., UniProt P03369), human immunodeficiency virus type II (HIV-2) (e.g., UniProt P15833), simian immunodeficiency virus (SIV) (e.g., UniProt P05896), bovine immunodeficiency virus (BIV) (e.g., UniProt P19560), equine infectious anemia virus (EIAV) (e.g., UniProt P03371), or feline immunodeficiency virus (FIV) (e.g., UniProt P16088) (Herschhorn and Hizi Cell Mol Life Sci 67(16):2717-2747 (2010)), or a functional fragment or variant thereof (e.g., an amino acid sequence having at least 70%, 80%, 90%, 95%, or 99% identity thereto). Naturally heterodimeric RT domains may, in some embodiments, also be functional as homodimers. In some embodiments, dimeric RT domains are expressed as fusion proteins, e.g., as homodimeric fusion proteins or heterodimeric fusion proteins. In some embodiments, the RT function of the system is fulfilled by multiple RT domains (e.g., as described herein). In further embodiments, the multiple RT domains are fused or separate, e.g., may be on the same polypeptide or on different polypeptides.


In some embodiments, a gene modifying system described herein comprises an integrase domain, e.g., wherein the integrase domain may be part of the RT domain. In some embodiments, an RT domain (e.g., as described herein) comprises an integrase domain. In some embodiments, an RT domain (e.g., as described herein) lacks an integrase domain, or comprises an integrase domain that has been inactivated by mutation or deleted. In some embodiment, a gene modifying system described herein comprises an RNase H domain, e.g., wherein the RNase H domain may be part of the RT domain. In some embodiments, the RNase H domain is not part of the RT domain and is covalently linked via a flexible linker. In some embodiments, an RT domain (e.g., as described herein) comprises an RNase H domain, e.g., an endogenous RNAse H domain or a heterologous RNase H domain. In some embodiments, an RT domain (e.g., as described herein) lacks an RNase H domain. In some embodiments, an RT domain (e.g., as described herein) comprises an RNase H domain that has been added, deleted, mutated, or swapped for a heterologous RNase H domain. In some embodiments, the polypeptide comprises an inactivated endogenous RNase H domain. In some embodiments, an endogenous RNase H domain from one of the other domains of the polypeptide is genetically removed such that it is not included in the polypeptide, e.g., the endogenous RNase H domain is partially or completely truncated from the comprising domain. In some embodiments, mutation of an RNase H domain yields a polypeptide exhibiting lower RNase activity, e.g., as determined by the methods described in Kotewicz et al. Nucleic Acids Res 16(1):265-277 (1988) (incorporated herein by reference in its entirety), e.g., lower by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% compared to an otherwise similar domain without the mutation. In some embodiments, RNase H activity is abolished.


In some embodiments, an RT domain is mutated to increase fidelity compared to an otherwise similar domain without the mutation. For instance, in some embodiments, a YADD (SEQ ID NO: 25690) or YMDD (SEQ ID NO: 25691) motif in an RT domain (e.g., in a reverse transcriptase) is replaced with YVDD (SEQ ID NO: 25692). In embodiments, replacement of the YADD (SEQ ID NO: 25690) or YMDD (SEQ ID NO: 25691) or YVDD (SEQ ID NO: 25692) results in higher fidelity in retroviral reverse transcriptase activity (e.g., as described in Jamburuthugoda and Eickbush J Mol Biol 2011; incorporated herein by reference in its entirety).


In some embodiments, a gene modifying polypeptide described herein comprises an RT domain having an amino acid sequence according to Table 6, or a sequence having at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity thereto. In some embodiments, a nucleic acid described herein encodes an RT domain having an amino acid sequence according to Table 6, or a sequence having at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity thereto.









TABLE 6







Exemplary reverse transcriptase domains from retroviruses









RT
SEQ ID



Name
NO:
RT amino acid sequence





AVIRE_
8,001
TAPLEEEYRLFLEAPIQNVTLLEQWKREIPKVWAEINPPGLASTQAPIHV


P03360

QLLSTALPVRVRQYPITLEAKRSLRETIRKFRAAGILRPVHSPWNTPLLP




VRKSGTSEYRMVQDLREVNKRVETIHPTVPNPYTLLSLLPPDRIWYSVLD




LKDAFFCIPLAPESQLIFAFEWADAEEGESGQLTWTRLPQGFKNSPTLFD




EALNRDLQGFRLDHPSVSLLQYVDDLLIAADTQAACLSATRDLLMTLAEL




GYRVSGKKAQLCQEEVTYLGFKIHKGSRSLSNSRTQAILQIPVPKTKRQV




REFLGTIGYCRLWIPGFAELAQPLYAATRGGNDPLVWGEKEEEAFQSLKL




ALTQPPALALPSLDKPFQLFVEETSGAAKGVLTQALGPWKRPVAYLSKRL




DPVAAGWPRCLRAIAAAALLTREASKLTFGQDIEITSSHNLESLLRSPPD




KWLTNARITQYQVLLLDPPRVRFKQTAALNPATLLPETDDTLPIHHCLDT




LDSLTSTRPDLTDQPLAQAEATLFTDGSSYIRDGKRYAGAAVVTLDSVIW




AEPLPIGTSAQKAELIALTKALEWSKDKSVNIYTDSRYAFATLHVHGMIY




RERGLLTAGGKAIKNAPEILALLTAVWLPKRVAVMHCKGHQKDDAPTSTG




NRRADEVAREVAIRPLSTQATIS





AVIRE_
8,002
TAPLEEEYRLFLEAPIQNVTLLEQWKREIPKVWAEINPPGLASTQAPIHV


P03360_

QLLSTALPVRVRQYPITLEAKRSLRETIRKFRAAGILRPVHSPWNTPLLP


3mut

VRKSGTSEYRMVQDLREVNKRVETIHPTVPNPYTLLSLLPPDRIWYSVLD




LKDAFFCIPLAPESQLIFAFEWADAEEGESGQLTWTRLPQGFKNSPTLFN




EALNRDLQGFRLDHPSVSLLQYVDDLLIAADTQAACLSATRDLLMTLAEL




GYRVSGKKAQLCQEEVTYLGFKIHKGSRSLSNSRTQAILQIPVPKTKRQV




REFLGTIGYCRLWIPGFAELAQPLYAATRPGNDPLVWGEKEEEAFQSLKL




ALTQPPALALPSLDKPFQLFVEETSGAAKGVLTQALGPWKRPVAYLSKRL




DPVAAGWPRCLRAIAAAALLTREASKLTFGQDIEITSSHNLESLLRSPPD




KWLTNARITQYQVLLLDPPRVRFKQTAALNPATLLPETDDTLPIHHCLDT




LDSLTSTRPDLTDQPLAQAEATLFTDGSSYIRDGKRYAGAAVVTLDSVIW




AEPLPIGTSAQKAELIALTKALEWSKDKSVNIYTDSRYAFATLHVHGMIY




RERGWLTAGGKAIKNAPEILALLTAVWLPKRVAVMHCKGHQKDDAPTSTG




NRRADEVAREVAIRPLSTQATIS





AVIRE_
8,003
TAPLEEEYRLFLEAPIQNVTLLEQWKREIPKVWAEINPPGLASTQAPIHV


P03360_

QLLSTALPVRVRQYPITLEAKRSLRETIRKFRAAGILRPVHSPWNTPLLP


3mutA

VRKSGTSEYRMVQDLREVNKRVETIHPTVPNPYTLLSLLPPDRIWYSVLD




LKDAFFCIPLAPESQLIFAFEWADAEEGESGQLTWTRLPQGFKNSPTLFN




EALNRDLQGFRLDHPSVSLLQYVDDLLIAADTQAACLSATRDLLMTLAEL




GYRVSGKKAQLCQEEVTYLGFKIHKGSRSLSNSRTQAILQIPVPKTKRQV




REFLGKIGYCRLFIPGFAELAQPLYAATRPGNDPLVWGEKEEEAFQSLKL




ALTQPPALALPSLDKPFQLFVEETSGAAKGVLTQALGPWKRPVAYLSKRL




DPVAAGWPRCLRAIAAAALLTREASKLTFGQDIEITSSHNLESLLRSPPD




KWLTNARITQYQVLLLDPPRVRFKQTAALNPATLLPETDDTLPIHHQLDT




LDSLTSTRPDLTDQPLAQAEATLFTDGSSYIRDGKRYAGAAVVTLDSVIW




AEPLPIGTSAQKAELIALTKALEWSKDKSVNIYTDSRYAFATLHVHGMIY




RERGWLTAGGKAIKNAPEILALLTAVWLPKRVAVMHCKGHQKDDAPTSTG




NRRADEVAREVAIRPLSTQATIS





BAEVM_
8,004
TVSLQDEHRLFDIPVTTSLPDVWLQDFPQAWAETGGLGRAKCQAPIIIDL


P10272

KPTAVPVSIKQYPMSLEAHMGIRQHIIKFLELGVLRPCRSPWNTPLLPVK




KPGTQDYRPVQDLREINKRTVDIHPTVPNPYNLLSTLKPDYSWYTVLDLK




DAFFCLPLAPQSQELFAFEWKDPERGISGQLTWTRLPQGFKNSPTLFDEA




LHRDLTDFRTQHPEVTLLQYVDDLLLAAPTKKACTQGTRHLLQELGEKGY




RASAKKAQICQTKVTYLGYILSEGKRWLTPGRIETVARIPPPRNPREVRE




FLGTAGFCRLWIPGFAELAAPLYALTKESTPFTWQTEHQLAFEALKKALL




SAPALGLPDTSKPFTLFLDERQGIAKGVLTQKLGPWKRPVAYLSKKLDPV




AAGWPPCLRIMAATAMLVKDSAKLTLGQPLTVITPHTLEAIVRQPPDRWI




TNARLTHYQALLLDTDRVQFGPPVTLNPATLLPVPENQPSPHDCRQVLAE




THGTREDLKDQELPDADHTWYTDGSSYLDSGTRRAGAAVVDGHNTIWAQS




LPPGTSAQKAELIALTKALELSKGKKANIYTDSRYAFATAHTHGSIYERR




GLLTSEGKEIKNKAEIIALLKALFLPQEVAIIHCPGHQKGQDPVAVGNRQ




ADRVARQAAMAEVLTLATEPDNTSHIT





BAEVM_
8,005
TVSLQDEHRLFDIPVTTSLPDVWLQDFPQAWAETGGLGRAKCQAPIIIDL


P10272_

KPTAVPVSIKQYPMSLEAHMGIRQHIIKFLELGVLRPCRSPWNTPLLPVK


3mut

KPGTQDYRPVQDLREINKRTVDIHPTVPNPYNLLSTLKPDYSWYTVLDLK




DAFFCLPLAPQSQELFAFEWKDPERGISGQLTWTRLPQGFKNSPTLFNEA




LHRDLTDFRTQHPEVTLLQYVDDLLLAAPTKKACTQGTRHLLQELGEKGY




RASAKKAQICQTKVTYLGYILSEGKRWLTPGRIETVARIPPPRNPREVRE




FLGTAGFCRLWIPGFAELAAPLYALTKPSTPFTWQTEHQLAFEALKKALL




SAPALGLPDTSKPFTLFLDERQGIAKGVLTQKLGPWKRPVAYLSKKLDPV




AAGWPPCLRIMAATAMLVKDSAKLTLGQPLTVITPHTLEAIVRQPPDRWI




TNARLTHYQALLLDTDRVQFGPPVTLNPATLLPVPENQPSPHDCRQVLAE




THGTREDLKDQELPDADHTWYTDGSSYLDSGTRRAGAAVVDGHNTIWAQS




LPPGTSAQKAELIALTKALELSKGKKANIYTDSRYAFATAHTHGSIYERR




GWLTSEGKEIKNKAEIIALLKALFLPQEVAIIHCPGHQKGQDPVAVGNRQ




ADRVARQAAMAEVLTLATEPDNTSHIT





BAEVM_
8,006
TVSLQDEHRLFDIPVTTSLPDVWLQDFPQAWAETGGLGRAKCQAPIIIDL


P10272_

KPTAVPVSIKQYPMSLEAHMGIRQHIIKFLELGVLRPCRSPWNTPLLPVK


3mutA

KPGTQDYRPVQDLREINKRTVDIHPTVPNPYNLLSTLKPDYSWYTVLDLK




DAFFCLPLAPQSQELFAFEWKDPERGISGQLTWTRLPQGFKNSPTLFNEA




LHRDLTDFRTQHPEVTLLQYVDDLLLAAPTKKACTQGTRHLLQELGEKGY




RASAKKAQICQTKVTYLGYILSEGKRWLTPGRIETVARIPPPRNPREVRE




FLGKAGFCRLFIPGFAELAAPLYALTKPSTPFTWQTEHQLAFEALKKALL




SAPALGLPDTSKPFTLFLDERQGIAKGVLTQKLGPWKRPVAYLSKKLDPV




AAGWPPCLRIMAATAMLVKDSAKLTLGQPLTVITPHTLEAIVRQPPDRWI




TNARLTHYQALLLDTDRVQFGPPVTLNPATLLPVPENQPSPHDCRQVLAE




THGTREDLKDQELPDADHTWYTDGSSYLDSGTRRAGAAVVDGHNTIWAQS




LPPGTSAQKAELIALTKALELSKGKKANIYTDSRYAFATAHTHGSIYERR




GWLTSEGKEIKNKAEIIALLKALFLPQEVAIIHCPGHQKGQDPVAVGNRQ




ADRVARQAAMAEVLTLATEPDNTSHIT





BLVAU_
8,007
GVLDAPPSHIGLEHLPPPPEVPQFPLNLERLQALQDLVHRSLEAGYISPW


P25059

DGPGNNPVFPVRKPNGAWRFVHDLRVTNALTKPIPALSPGPPDLTAIPTH




LPHIICLDLKDAFFQIPVEDRFRSYFAFTLPTPGGLQPHRRFAWRVLPQG




FINSPALFERALQEPLRQVSAAFSQSLLVSYMDDILYVSPTEEQRLQCYQ




TMAAHLRDLGFQVASEKTRQTPSPVPFLGQMVHERMVTYQSLPTLQISSP




ISLHQLQTVLGDLQWVSRGTPTTRRPLQLLYSSLKGIDDPRAIIHLSPEQ




QQGIAELRQALSHNARSRYNEQEPLLAYVHLTRAGSTLVLFQKGAQFPLA




YFQTPLTDNQASPWGLLLLLGCQYLQAQALSSYAKTILKYYHNLPKTSLD




NWIQSSEDPRVQELLQLWPQISSQGIQPPGPWKTLVTRAEVFLTPQFSPE




PIPAALCLFSDGAARRGAYCLWKDHLLDFQAVPAPESAQKGELAGLLAGL




AAAPPEPLNIWVDSKYLYSLLRTLVLGAWLQPDPVPSYALLYKSLLRHPA




IFVGHVRSHSSASHPIASLNNYVDQL





BLVAU_
8,008
GVLDAPPSHIGLEHLPPPPEVPQFPLNLERLQALQDLVHRSLEAGYISPW


P25059_

DGPGNNPVFPVRKPNGAWRFVHDLRVTNALTKPIPALSPGPPDLTAIPTH


2mut

LPHIICLDLKDAFFQIPVEDRFRSYFAFTLPTPGGLQPHRRFAWRVLPQG




FINSPALFQRALQEPLRQVSAAFSQSLLVSYMDDILYVSPTEEQRLQCYQ




TMAAHLRDLGFQVASEKTRQTPSPVPFLGQMVHERMVTYQSLPTLQISSP




ISLHQLQTVLGDLQWVSRGTPTTRRPLQLLYSSLKPIDDPRAIIHLSPEQ




QQGIAELRQALSHNARSRYNEQEPLLAYVHLTRAGSTLVLFQKGAQFPLA




YFQTPLTDNQASPWGLLLLLGCQYLQAQALSSYAKTILKYYHNLPKTSLD




NWIQSSEDPRVQELLQLWPQISSQGIQPPGPWKTLVTRAEVFLTPQFSPE




PIPAALCLFSDGAARRGAYCLWKDHLLDFQAVPAPESAQKGELAGLLAGL




AAAPPEPLNIWVDSKYLYSLLRTLVLGAWLQPDPVPSYALLYKSLLRHPA




IFVGHVRSHSSASHPIASLNNYVDQL





BLVJ_
8,009
GVLDTPPSHIGLEHLPPPPEVPQFPLNLERLQALQDLVHRSLEAGYISPW


P03361

DGPGNNPVFPVRKPNGAWRFVHDLRATNALTKPIPALSPGPPDLTAIPTH




PPHIICLDLKDAFFQIPVEDRFRFYLSFTLPSPGGLQPHRRFAWRVLPQG




FINSPALFERALQEPLRQVSAAFSQSLLVSYMDDILYASPTEEQRSQCYQ




ALAARLRDLGFQVASEKTSQTPSPVPFLGQMVHEQIVTYQSLPTLQISSP




ISLHQLQAVLGDLQWVSRGTPTTRRPLQLLYSSLKRHHDPRAIIQLSPEQ




LQGIAELRQALSHNARSRYNEQEPLLAYVHLTRAGSTLVLFQKGAQFPLA




YFQTPLTDNQASPWGLLLLLGCQYLQTQALSSYAKPILKYYHNLPKTSLD




NWIQSSEDPRVQELLQLWPQISSQGIQPPGPWKTLITRAEVFLTPQFSPD




PIPAALCLFSDGATGRGAYCLWKDHLLDFQAVPAPESAQKGELAGLLAGL




AAAPPEPVNIWVDSKYLYSLLRTLVLGAWLQPDPVPSYALLYKSLLRHPA




IVVGHVRSHSSASHPIASLNNYVDQL





BLVJ_
8,010
GVLDTPPSHIGLEHLPPPPEVPQFPLNLERLQALQDLVHRSLEAGYISPW


P03361_

DGPGNNPVFPVRKPNGAWRFVHDLRATNALTKPIPALSPGPPDLTAIPTH


2mut

PPHIICLDLKDAFFQIPVEDRFRFYLSFTLPSPGGLQPHRRFAWRVLPQG




FINSPALFNRALQEPLRQVSAAFSQSLLVSYMDDILYASPTEEQRSQCYQ




ALAARLRDLGFQVASEKTSQTPSPVPFLGQMVHEQIVTYQSLPTLQISSP




ISLHQLQAVLGDLQWVSRGTPTTRRPLQLLYSSLKRHHDPRAIIQLSPEQ




LQGIAELRQALSHNARSRYNEQEPLLAYVHLTRAGSTLVLFQKGAQFPLA




YFQTPLTDNQASPWGLLLLLGCQYLQTQALSSYAKPILKYYHNLPKTSLD




NWIQSSEDPRVQELLQLWPQISSQGIQPPGPWKTLITRAEVFLTPQFSPD




PIPAALCLFSDGATGRGAYCLWKDHLLDFQAVPAPESAQKGELAGLLAGL




AAAPPEPVNIWVDSKYLYSLLRTWVLGAWLQPDPVPSYALLYKSLLRHPA




IVVGHVRSHSSASHPIASLNNYVDQL





BLVJ_
8,011
GVLDTPPSHIGLEHLPPPPEVPQFPLNLERLQALQDLVHRSLEAGYISPW


P03361_

DGPGNNPVFPVRKPNGAWRFVHDLRATNALTKPIPALSPGPPDLTAPPTH


2mutB

PPHIICLDLKDAFFQIPVEDRFRFYLSFTLPSPGGLQPHRRFAWRVLPQG




FINSPALFQRALQEPLRQVSAAFSQSLLVSYMDDILYASPTEEQRSQCYQ




ALAARLRDLGFQVASEKTSQTPSPVPFLGQMVHEQIVTYQSLPTLQISSP




ISLHQLQAVLGDLQWVSRGTPTTRRPLQLLYSSLKRHHDPRAIIQLSPEQ




LQGIAELRQALSHNARSRYNEQEPLLAYVHLTRAGSTLVLFQKGAQFPLA




YFQTPLTDNQASPWGLLLLLGCQYLQTQALSSYAKPILKYYHNLPKTSLD




NWIQSSEDPRVQELLQLWPQISSQGIQPPGPWKTLITRAEVFLTPQFSPD




PIPAALCLFSDGATGRGAYCLWKDHLLDFQAVPAPESAQKGELAGLLAGL




AAAPPEPVNIWVDSKYLYSLLRTWVLGAWLQPDPVPSYALLYKSLLRHPA




IVVGHVRSHSSASHPIASLNNYVDQL





FFV_
8,012
MDLLKPLTVERKGVKIKGYWNSQADITCVPKDLLQGEEPVRQQNVTTIHG


O93209

TQEGDVYYVNLKIDGRRINTEVIGTTLDYAIITPGDVPWILKKPLELTIK




LDLEEQQGTLLNNSILSKKGKEELKQLFEKYSALWQSWENQVGHRRIRPH




KIATGTVKPTPQKQYHINPKAKPDIQIVINDLLKQGVLIQKESTMNTPVY




PVPKPNGRWRMVLDYRAVNKVTPLIAVQNQHSYGILGSLFKGRYKTTIDL




SNGFWAHPIVPEDYWITAFTWQGKQYCWTVLPQGFLNSPGLFTGDVVDLL




QGIPNVEVYVDDVYISHDSEKEHLEYLDILFNRLKEAGYIISLKKSNIAN




SIVDFLGFQITNEGRGLTDTFKEKLENITAPTTLKQLQSILGLLNFARNF




IPDFTELIAPLYALIPKSTKNYVPWQIEHSTTLETLITKLNGAEYLQGRK




GDKTLIMKVNASYTTGYIRYYNEGEKKPISYVSIVFSKTELKFTELEKLL




TTVHKGLLKALDLSMGQNIHVYSPIVSMQNIQKTPQTAKKALASRWLSWL




SYLEDPRIRFFYDPQMPALKDLPAVDTGKDNKKHPSNFQHIFYTDGSAIT




SPTKEGHLNAGMGIVYFINKDGNLQKQQEWSISLGNHTAQFAEIAAFEFA




LKKCLPLGGNILVVTDSNYVAKAYNEELDVWASNGFVNNRKKPLKHISKW




KSVADLKRLRPDVVVTHEPGHQKLDSSPHAYGNNLADQLATQASFKVH





FFV_
8,013
MDLLKPLTVERKGVKIKGYWNSQADITCVPKDLLQGEEPVRQQNVTTIHG


O93209_

TQEGDVYYVNLKIDGRRINTEVIGTTLDYAIITPGDVPWILKKPLELTIK


2mut

LDLEEQQGTLLNNSILSKKGKEELKQLFEKYSALWQSWENQVGHRRIRPH




KIATGTVKPTPQKQYHINPKAKPDIQIVINDLLKQGVLIQKESTMNTPVY




PVPKPNGRWRMVLDYRAVNKVTPLIAVQNQHSYGILGSLFKGRYKTTIDL




SNGFWAHPIVPEDYWITAFTWQGKQYCWTVLPQGFLNSPGLFNGDVVDLL




QGIPNVEVYVDDVYISHDSEKEHLEYLDILFNRLKEAGYIISLKKSNIAN




SIVDFLGFQITNEGRGLTDTFKEKLENITAPTTLKQLQSILGLLNFARNF




IPDFTELIAPLYALIPKSPKNYVPWQIEHSTTLETLITKLNGAEYLQGRK




GDKTLIMKVNASYTTGYIRYYNEGEKKPISYVSIVFSKTELKFTELEKLL




TTVHKGLLKALDLSMGQNIHVYSPIVSMQNIQKTPQTAKKALASRWLSWL




SYLEDPRIRFFYDPQMPALKDLPAVDTGKDNKKHPSNFQHIFYTDGSAIT




SPTKEGHLNAGMGIVYFINKDGNLQKQQEWSISLGNHTAQFAEIAAFEFA




LKKCLPLGGNILVVTDSNYVAKAYNEELDVWASNGFVNNRKKPLKHISKW




KSVADLKRLRPDVVVTHEPGHQKLDSSPHAYGNNLADQLATQASFKVH





FFV_
8,014
MDLLKPLTVERKGVKIKGYWNSQADITCVPKDLLQGEEPVRQQNVTTIHG


O93209_

TQEGDVYYVNLKIDGRRINTEVIGTTLDYAIITPGDVPWILKKPLELTIK


2mutA

LDLEEQQGTLLNNSILSKKGKEELKQLFEKYSALWQSWENQVGHRRIRPH




KIATGTVKPTPQKQYHINPKAKPDIQIVINDLLKQGVLIQKESTMNTPVY




PVPKPNGRWRMVLDYRAVNKVTPLIAVQNQHSYGILGSLFKGRYKTTIDL




SNGFWAHPIVPEDYWITAFTWQGKQYCWTVLPQGFLNSPGLFNGDVVDLL




QGIPNVEVYVDDVYISHDSEKEHLEYLDILFNRLKEAGYIISLKKSNIAN




SIVDFLGFQITNEGRGLTDTFKEKLENITAPTTLKQLQSILGKLNFARNF




IPDFTELIAPLYALIPKSPKNYVPWQIEHSTTLETLITKLNGAEYLQGRK




GDKTLIMKVNASYTTGYIRYYNEGEKKPISYVSIVFSKTELKFTELEKLL




TTVHKGLLKALDLSMGQNIHVYSPIVSMQNIQKTPQTAKKALASRWLSWL




SYLEDPRIRFFYDPQMPALKDLPAVDTGKDNKKHPSNFQHIFYTDGSAIT




SPTKEGHLNAGMGIVYFINKDGNLQKQQEWSISLGNHTAQFAEIAAFEFA




LKKCLPLGGNILVVTDSNYVAKAYNEELDVWASNGFVNNRKKPLKHISKW




KSVADLKRLRPDVVVTHEPGHQKLDSSPHAYGNNLADQLATQASFKVH





FFV_
8,015
VPWILKKPLELTIKLDLEEQQGTLLNNSILSKKGKEELKQLFEKYSALWQ


O93209-

SWENQVGHRRIRPHKIATGTVKPTPQKQYHINPKAKPDIQIVINDLLKQG


Pro

VLIQKESTMNTPVYPVPKPNGRWRMVLDYRAVNKVTPLIAVQNQHSYGIL




GSLFKGRYKTTIDLSNGFWAHPIVPEDYWITAFTWQGKQYCWTVLPQGFL




NSPGLFTGDVVDLLQGIPNVEVYVDDVYISHDSEKEHLEYLDILFNRLKE




AGYIISLKKSNIANSIVDFLGFQITNEGRGLTDTFKEKLENITAPTTLKQ




LQSILGLLNFARNFIPDFTELIAPLYALIPKSTKNYVPWQIEHSTTLETL




ITKLNGAEYLQGRKGDKTLIMKVNASYTTGYIRYYNEGEKKPISYVSIVF




SKTELKFTELEKLLTTVHKGLLKALDLSMGQNIHVYSPIVSMQNIQKTPQ




TAKKALASRWLSWLSYLEDPRIRFFYDPQMPALKDLPAVDTGKDNKKHPS




NFQHIFYTDGSAITSPTKEGHLNAGMGIVYFINKDGNLQKQQEWSISLGN




HTAQFAEIAAFEFALKKCLPLGGNILVVTDSNYVAKAYNEELDVWASNGF




VNNRKKPLKHISKWKSVADLKRLRPDVVVTHEPGHQKLDSSPHAYGNNLA




DQLATQASFKVH





FFV_
8,016
VPWILKKPLELTIKLDLEEQQGTLLNNSILSKKGKEELKQLFEKYSALWQ


O93209-

SWENQVGHRRIRPHKIATGTVKPTPQKQYHINPKAKPDIQIVINDLLKQG


Pro_

VLIQKESTMNTPVYPVPKPNGRWRMVLDYRAVNKVTPLIAVQNQHSYGIL


2mut

GSLFKGRYKTTIDLSNGFWAHPIVPEDYWITAFTWQGKQYCWTVLPQGFL




NSPGLFNGDWVDLLQGIPNVEVYVDDVYISHDSEKEHLEYLDILFNRLKE




AGYIISLKKSNIANSIVDFLGFQITNEGRGLTDTFKEKLENITAPTTLKQ




LQSILGLLNFARNFIPDFTELIAPLYALIPKSPKNYVPWQIEHSTTLETL




ITKLNGAEYLQGRKGDKTLIMKVNASYTTGYIRYYNEGEKKPISYVSIVF




SKTELKFTELEKLLTTVHKGLLKALDLSMGQNIHVYSPIVSMQNIQKTPQ




TAKKALASRWLSWLSYLEDPRIRFFYDPQMPALKDLPAVDTGKDNKKHPS




NFQHIFYTDGSAITSPTKEGHLNAGMGIVYFINKDGNLQKQQEWSISLGN




HTAQFAEIAAFEFALKKCLPLGGNILVVTDSNYVAKAYNEELDVWASNGF




VNNRKKPLKHISKWKSVADLKRLRPDVVVTHEPGHQKLDSSPHAYGNNLA




DQLATQASFKVH





FFV_
8,017
VPWILKKPLELTIKLDLEEQQGTLLNNSILSKKGKEELKQLFEKYSALWQ


O93209-

SWENQVGHRRIRPHKIATGTVKPTPQKQYHINPKAKPDIQIVINDLLKQG


Pro_

VLIQKESTMNTPVYPVPKPNGRWRMVLDYRAVNKVTPLIAVQNQHSYGIL


2mutA

GSLFKGRYKTTIDLSNGFWAHPIVPEDYWITAFTWQGKQYCWTVLPQGFL




NSPGLFNGDVVDLLQGIPNVEVYVDDVYISHDSEKEHLEYLDILFNRLKE




AGYIISLKKSNIANSIVDFLGFQITNEGRGLTDTFKEKLENITAPTTLKQ




LQSILGKLNFARNFIPDFTELIAPLYALIPKSPKNYVPWQIEHSTTLETL




ITKLNGAEYLQGRKGDKTLIMKVNASYTTGYIRYYNEGEKKPISYVSIVF




SKTELKFTELEKLLTTVHKGLLKALDLSMGQNIHVYSPIVSMQNIQKTPQ




TAKKALASRWLSWLSYLEDPRIRFFYDPQMPALKDLPAVDTGKDNKKHPS




NFQHIFYTDGSAITSPTKEGHLNAGMGIVYFINKDGNLQKQQEWSISLGN




HTAQFAEIAAFEFALKKCLPLGGNILVVTDSNYVAKAYNEELDVWASNGF




VNNRKKPLKHISKWKSVADLKRLRPDVVVTHEPGHQKLDSSPHAYGNNLA




DQLATQASFKVH





FLV_
8,018
TLQLEEEYRLFEPESTQKQEMDIWLKNFPQAWAETGGMGTAHCQAPVLIQ


P10273

LKATATPISIRQYPMPHEAYQGIKPHIRRMLDQGILKPCQSPWNTPLLPV




KKPGTEDYRPVQDLREVNKRVEDIHPTVPNPYNLLSTLPPSHPWYTVLDL




KDAFFCLRLHSESQLLFAFEWRDPEIGLSGQLTWTRLPQGFKNSPTLFDE




ALHSDLADFRVRYPALVLLQYVDDLLLAAATRTECLEGTKALLETLGNKG




YRASAKKAQICLQEVTYLGYSLKDGQRWLTKARKEAILSIPVPKNSRQVR




EFLGTAGYCRLWIPGFAELAAPLYPLTRPGTLFQWGTEQQLAFEDIKKAL




LSSPALGLPDITKPFELFIDENSGFAKGVLVQKLGPWKRPVAYLSKKLDT




VASGWPPCLRMVAAIAILVKDAGKLTLGQPLTILTSHPVEALVRQPPNKW




LSNARMTHYQAMLLDAERVHFGPTVSLNPATLLPLPSGGNHHDCLQILAE




THGTRPDLTDQPLPDADLTWYTDGSSFIRNGEREAGAAVTTESEVIWAAP




LPPGTSAQRAELIALTQALKMAEGKKLTVYTDSRYAFATTHVHGEIYRRR




GLLTSEGKEIKNKNEILALLEALFLPKRLSIIHCPGHQKGDSPQAKGNRL




ADDTAKKAATETHSSLTVLP





FLV_
8,019
TLQLEEEYRLFEPESTQKQEMDIWLKNFPQAWAETGGMGTAHCQAPVLIQ


P10273_

LKATATPISIRQYPMPHEAYQGIKPHIRRMLDQGILKPCQSPWNTPLLPV


3mut

KKPGTEDYRPVQDLREVNKRVEDIHPTVPNPYNLLSTLPPSHPWYTVLDL




KDAFFCLRLHSESQLLFAFEWRDPEIGLSGQLTWTRLPQGFKNSPTLFNE




ALHSDLADFRVRYPALVLLQYVDDLLLAAATRTECLEGTKALLETLGNKG




YRASAKKAQICLQEVTYLGYSLKDGQRWLTKARKEAILSIPVPKNSRQVR




EFLGTAGYCRLWIPGFAELAAPLYPLTRPGTLFQWGTEQQLAFEDIKKAL




LSSPALGLPDITKPFELFIDENSGFAKGVLVQKLGPWKRPVAYLSKKLDT




VASGWPPCLRMVAAIAILVKDAGKLTLGQPLTILTSHPVEALVRQPPNKW




LSNARMTHYQAMLLDAERVHFGPTVSLNPATLLPLPSGGNHHDCLQILAE




THGTRPDLTDQPLPDADLTWYTDGSSFIRNGEREAGAAVTTESEVIWAAP




LPPGTSAQRAELIALTQALKMAEGKKLTVYTDSRYAFATTHVHGEIYRRR




GWLTSEGKEIKNKNEILALLEALFLPKRLSIIHCPGHQKGDSPQAKGNRL




ADDTAKKAATETHSSLTVLP





FLV_
8,020
TLQLEEEYRLFEPESTQKQEMDIWLKNFPQAWAETGGMGTAHCQAPVLIQ


P10273_

LKATATPISIRQYPMPHEAYQGIKPHIRRMLDQGILKPCQSPWNTPLLPV


3mutA

KKPGTEDYRPVQDLREVNKRVEDIHPTVPNPYNLLSTLPPSHPWYTVLDL




KDAFFCLRLHSESQLLFAFEWRDPEIGLSGQLTWTRLPQGFKNSPTLFNE




ALHSDLADFRVRYPALVLLQYVDDLLLAAATRTECLEGTKALLETLGNKG




YRASAKKAQICLQEVTYLGYSLKDGQRWLTKARKEAILSIPVPKNSRQVR




EFLGKAGYCRLFIPGFAELAAPLYPLTRPGTLFQWGTEQQLAFEDIKKAL




LSSPALGLPDITKPFELFIDENSGFAKGVLVQKLGPWKRPVAYLSKKLDT




VASGWPPCLRMVAAIAILVKDAGKLTLGQPLTILTSHPVEALVRQPPNKW




LSNARMTHYQAMLLDAERVHFGPTVSLNPATLLPLPSGGNHHDCLQILAE




THGTRPDLTDQPLPDADLTWYTDGSSFIRNGEREAGAAVTTESEVIWAAP




LPPGTSAQRAELIALTQALKMAEGKKLTVYTDSRYAFATTHVHGEIYRRR




GWLTSEGKEIKNKNEILALLEALFLPKRLSIIHCPGHQKGDSPQAKGNRL




ADDTAKKAATETHSSLTVLP





FOAMV_
8,021
MNPLQLLQPLPAEIKGTKLLAHWNSGATITCIPESFLEDEQPIKKTLIKT


P14350

IHGEKQQNVYYVTFKVKGRKVEAEVIASPYEYILLSPTDVPWLTQQPLQL




TILVPLQEYQEKILSKTALPEDQKQQLKTLFVKYDNLWQHWENQVGHRKI




RPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGVLTPQNSTMNT




PVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILATIVRQKYKTT




LDLANGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFLNSPALFTADVV




DLLKEIPNVQVYVDDIYLSHDDPKEHVQQLEKVFQILLQAGYVVSLKKSE




IGQKTVEFLGFNITKEGRGLTDTFKTKLLNITPPKDLKQLQSILGLLNFA




RNFIPNFAELVQPLYNLIASAKGKYIEWSEENTKQLNMVIEALNTASNLE




ERLPEQRLVIKVNTSPSAGYVRYYNETGKKPIMYLNYVFSKAELKFSMLE




KLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSQSPVKHPSQYEGVFYTDG




SAIKSPDPTKSNNAGMGIVHATYKPEYQVLNQWSIPLGNHTAQMAEIAAV




EFACKKALKIPGPVLVITDSFYVAESANKELPYWKSNGFVNNKKKPLKHI




SKWKSIAECLSMKPDITIQHEKGISLQIPVFILKGNALADKLATQGSYVV




N





FOAMV_
8,022
MNPLQLLQPLPAEIKGTKLLAHWNSGATITCIPESFLEDEQPIKKTLIKT


P14350_

IHGEKQQNVYYVTFKVKGRKVEAEVIASPYEYILLSPTDVPWLTQQPLQL


2mut

TILVPLQEYQEKILSKTALPEDQKQQLKTLFVKYDNLWQHWENQVGHRKI




RPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGVLTPQNSTMNT




PVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILATIVRQKYKTT




LDLANGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFLNSPALFNADVV




DLLKEIPNVQVYVDDIYLSHDDPKEHVQQLEKVFQILLQAGYVVSLKKSE




IGQKTVEFLGFNITKEGRGLTDTFKTKLLNITPPKDLKQLQSILGLLNFA




RNFIPNFAELVQPLYNLIAPAKGKYIEWSEENTKQLNMVIEALNTASNLE




ERLPEQRLVIKVNTSPSAGYVRYYNETGKKPIMYLNYVFSKAELKFSMLE




KLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSQSPVKHPSQYEGVFYTDG




SAIKSPDPTKSNNAGMGIVHATYKPEYQVLNQWSIPLGNHTAQMAEIAAV




EFACKKALKIPGPVLVITDSFYVAESANKELPYWKSNGFVNNKKKPLKHI




SKWKSIAECLSMKPDITIQHEKGISLQIPVFILKGNALADKLATQGSYVV




N





FOAMV_
8,023
MNPLQLLQPLPAEIKGTKLLAHWNSGATITCIPESFLEDEQPIKKTLIKT


P14350_

IHGEKQQNVYYVTFKVKGRKVEAEVIASPYEYILLSPTDVPWLTQQPLQL


2mutA

TILVPLQEYQEKILSKTALPEDQKQQLKTLFVKYDNLWQHWENQVGHRKI




RPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGVLTPQNSTMNT




PVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILATIVRQKYKTT




LDLANGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFLNSPALFNADVV




DLLKEIPNVQVYVDDIYLSHDDPKEHVQQLEKVFQILLQAGYVVSLKKSE




IGQKTVEFLGFNITKEGRGLTDTFKTKLLNITPPKDLKQLQSILGKLNFA




RNFIPNFAELVQPLYNLIAPAKGKYIEWSEENTKQLNMVIEALNTASNLE




ERLPEQRLVIKVNTSPSAGYVRYYNETGKKPIMYLNYVFSKAELKFSMLE




KLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSQSPVKHPSQYEGVFYTDG




SAIKSPDPTKSNNAGMGIVHATYKPEYQVLNQWSIPLGNHTAQMAEIAAV




EFACKKALKIPGPVLVITDSFYVAESANKELPYWKSNGFVNNKKKPLKHI




SKWKSIAECLSMKPDITIQHEKGISLQIPVFILKGNALADKLATQGSYVV




N





FOAMV_
8,024
VPWLTQQPLQLTILVPLQEYQEKILSKTALPEDQKQQLKTLFVKYDNLWQ


P14350-

HWENQVGHRKIRPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro

VLTPQNSTMNTPVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGIL




ATIVRQKYKTTLDLANGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFL




NSPALFTADVVDLLKEIPNVQVYVDDIYLSHDDPKEHVQQLEKVFQILLQ




AGYVVSLKKSEIGQKTVEFLGFNITKEGRGLTDTFKTKLLNITPPKDLKQ




LQSILGLLNFARNFIPNFAELVQPLYNLIASAKGKYIEWSEENTKQLNMV




IEALNTASNLEERLPEQRLVIKVNTSPSAGYVRYYNETGKKPIMYLNYVF




SKAELKFSMLEKLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSQSPVKHP




SQYEGVFYTDGSAIKSPDPTKSNNAGMGIVHATYKPEYQVLNQWSIPLGN




HTAQMAEIAAVEFACKKALKIPGPVLVITDSFYVAESANKELPYWKSNGF




VNNKKKPLKHISKWKSIAECLSMKPDITIQHEKGISLQIPVFILKGNALA




DKLATQGSYVVN





FOAMV_
8,025
VPWLTQQPLQLTILVPLQEYQEKILSKTALPEDQKQQLKTLFVKYDNLWQ


P14350-

HWENQVGHRKIRPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro_

VLTPQNSTMNTPVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGIL


2mut

ATIVRQKYKTTLDLANGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFL




NSPALFNADVVDLLKEIPNVQVYVDDIYLSHDDPKEHVQQLEKVFQILLQ




AGYVVSLKKSEIGQKTVEFLGFNITKEGRGLTDTFKTKLLNITPPKDLKQ




LQSILGLLNFARNFIPNFAELVQPLYNLIAPAKGKYIEWSEENTKQLNMV




IEALNTASNLEERLPEQRLVIKVNTSPSAGYVRYYNETGKKPIMYLNYVF




SKAELKFSMLEKLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSQSPVKHP




SQYEGVFYTDGSAIKSPDPTKSNNAGMGIVHATYKPEYQVLNQWSIPLGN




HTAQMAEIAAVEFACKKALKIPGPVLVITDSFYVAESANKELPYWKSNGF




VNNKKKPLKHISKWKSIAECLSMKPDITIQHEKGISLQIPVFILKGNALA




DKLATQGSYVVN





FOAMV_
8,026
VPWLTQQPLQLTILVPLQEYQEKILSKTALPEDQKQQLKTLFVKYDNLWQ


P14350-

HWENQVGHRKIRPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro_

VLTPQNSTMNTPVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGIL


2mutA

ATIVRQKYKTTLDLANGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFL




NSPALFNADVVDLLKEIPNVQVYVDDIYLSHDDPKEHVQQLEKVFQILLQ




AGYVVSLKKSEIGQKTVEFLGFNITKEGRGLTDTFKTKLLNITPPKDLKQ




LQSILGKLNFARNFIPNFAELVQPLYNLIAPAKGKYIEWSEENTKQLNMV




IEALNTASNLEERLPEQRLVIKVNTSPSAGYVRYYNETGKKPIMYLNYVF




SKAELKFSMLEKLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSQSPVKHP




SQYEGVFYTDGSAIKSPDPTKSNNAGMGIVHATYKPEYQVLNQWSIPLGN




HTAQMAEIAAVEFACKKALKIPGPVLVITDSFYVAESANKELPYWKSNGF




VNNKKKPLKHISKWKSIAECLSMKPDITIQHEKGISLQIPVFILKGNALA




DKLATQGSYVVN





GALV_
8,027
VLNLEEEYRLHEKPVPSSIDPSWLQLFPTVWAERAGMGLANQVPPVVVEL


P21414

RSGASPVAVRQYPMSKEAREGIRPHIQKFLDLGVLVPCRSPWNTPLLPVK




KPGTNDYRPVQDLREINKRVQDIHPTVPNPYNLLSSLPPSYTWYSVLDLK




DAFFCLRLHPNSQPLFAFEWKDPEKGNTGQLTWTRLPQGFKNSPTLFDEA




LHRDLAPFRALNPQVVLLQYVDDLLVAAPTYEDCKKGTQKLLQELSKLGY




RVSAKKAQLCQREVTYLGYLLKEGKRWLTPARKATVMKIPVPTTPRQVRE




FLGTAGFCRLWIPGFASLAAPLYPLTKESIPFIWTEEHQQAFDHIKKALL




SAPALALPDLTKPFTLYIDERAGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPTCLKAVAAVALLLKDADKLTLGQNVTVIASHSLESIVRQPPDRWM




TNARMTHYQSLLLNERVSFAPPAVLNPATLLPVESEATPVHRCSEILAEE




TGTRRDLEDQPLPGVPTWYTDGSSFITEGKRRAGAPIVDGKRTVWASSLP




EGTSAQKAELVALTQALRLAEGKNINIYTDSRYAFATAHIHGAIYKQRGL




LTSAGKDIKNKEEILALLEAIHLPRRVAIIHCPGHQRGSNPVATGNRRAD




EAAKQAALSTRVLAGTTKP





GALV_
8,028
VLNLEEEYRLHEKPVPSSIDPSWLQLFPTVWAERAGMGLANQVPPVVVEL


P21414_

RSGASPVAVRQYPMSKEAREGIRPHIQKFLDLGVLVPCRSPWNTPLLPVK


3mut

KPGTNDYRPVQDLREINKRVQDIHPTVPNPYNLLSSLPPSYTWYSVLDLK




DAFFCLRLHPNSQPLFAFEWKDPEKGNTGQLTWTRLPQGFKNSPTLFNEA




LHRDLAPFRALNPQVVLLQYVDDLLVAAPTYEDCKKGTQKLLQELSKLGY




RVSAKKAQLCQREVTYLGYLLKEGKRWLTPARKATVMKIPVPTTPRQVRE




FLGTAGFCRLWIPGFASLAAPLYPLTKPSIPFIWTEEHQQAFDHIKKALL




SAPALALPDLTKPFTLYIDERAGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPTCLKAVAAVALLLKDADKLTLGQNVTVIASHSLESIVRQPPDRWM




TNARMTHYQSLLLNERVSFAPPAVLNPATLLPVESEATPVHRCSEILAEE




TGTRRDLEDQPLPGVPTWYTDGSSFITEGKRRAGAPIVDGKRTVWASSLP




EGTSAQKAELVALTQALRLAEGKNINIYTDSRYAFATAHIHGAIYKQRGW




LTSAGKDIKNKEEILALLEAIHLPRRVAIIHCPGHQRGSNPVATGNRRAD




EAAKQAALSTRVLAGTTKP





GALV_
8,029
VLNLEEEYRLHEKPVPSSIDPSWLQLFPTVWAERAGMGLANQVPPVVVEL


P21414_

RSGASPVAVRQYPMSKEAREGIRPHIQKFLDLGVLVPCRSPWNTPLLPVK


3mutA

KPGTNDYRPVQDLREINKRVQDIHPTVPNPYNLLSSLPPSYTWYSVLDLK




DAFFCLRLHPNSQPLFAFEWKDPEKGNTGQLTWTRLPQGFKNSPTLFNEA




LHRDLAPFRALNPQVVLLQYVDDLLVAAPTYEDCKKGTQKLLQELSKLGY




RVSAKKAQLCQREVTYLGYLLKEGKRWLTPARKATVMKIPVPTTPRQVRE




FLGKAGFCRLFIPGFASLAAPLYPLTKPSIPFIWTEEHQQAFDHIKKALL




SAPALALPDLTKPFTLYIDERAGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPTCLKAVAAVALLLKDADKLTLGQNVTVIASHSLESIVRQPPDRWM




TNARMTHYQSLLLNERVSFAPPAVLNPATLLPVESEATPVHRCSEILAEE




TGTRRDLEDQPLPGVPTWYTDGSSFITEGKRRAGAPIVDGKRTVWASSLP




EGTSAQKAELVALTQALRLAEGKNINIYTDSRYAFATAHIHGAIYKQRGW




LTSAGKDIKNKEEILALLEAIHLPRRVAIIHCPGHQRGSNPVATGNRRAD




EAAKQAALSTRVLAGTTKP





HTL1A_
8,030
AVLGLEHLPRPPQISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNP


P03362

VFPVKKANGTWRFIHDLRATNSLTIDLSSSSPGPPDLSSLPTTLAHLQTI




DLRDAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWKVLPQGFKNSPTL




FEMQLAHILQPIRQAFPQCTILQYMDDILLASPSHEDLLLLSEATMASLI




SHGLPVSENKTQQTPGTIKFLGQIISPNHLTYDAVPTVPIRSRWALPELQ




ALLGEIQWVSKGTPTLRQPLHSLYCALQRHTDPRDQIYLNPSQVQSLVQL




RQALSQNCRSRLVQTLPLLGAIMLTLTGTTTVVFQSKEQWPLVWLHAPLP




HTSQCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISTQTFNQFIQTSD




HPSVPILLHHSHRFKNLGAQTGELWNTFLKTAAPLAPVKALMPVFTLSPV




IINTAPCLFSDGSTSRAAYILWDKQILSQRSFPLPPPHKSAQRAELLGLL




HGLSSARSWRCLNIFLDSKYLYHYLRTLALGTFQGRSSQAPFQALLPRLL




SRKVVYLHHVRSHTNLPDPISRLNALTDALLITPVLQL





HTL1A_
8,031
AVLGLEHLPRPPQISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNP


P03362_

VFPVKKANGTWRFIHDLRATNSLTIDLSSSSPGPPDLSSLPTTLAHLQTI


2mut

DLRDAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWKVLPQGFKNSPTL




FQMQLAHILQPIRQAFPQCTILQYMDDILLASPSHEDLLLLSEATMASLI




SHGLPVSENKTQQTPGTIKFLGQIISPNHLTYDAVPTVPIRSRWALPELQ




ALLGEIQWVSKGTPTLRQPLHSLYCALQPHTDPRDQIYLNPSQVQSLVQL




RQALSQNCRSRLVQTLPLLGAIMLTLTGTTTVVFQSKEQWPLVWLHAPLP




HTSQCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISTQTFNQFIQTSD




HPSVPILLHHSHRFKNLGAQTGELWNTFLKTAAPLAPVKALMPVFTLSPV




IINTAPCLFSDGSTSRAAYILWDKQILSQRSFPLPPPHKSAQRAELLGLL




HGLSSARSWRCLNIFLDSKYLYHYLRTLALGTFQGRSSQAPFQALLPRLL




SRKVVYLHHVRSHTNLPDPISRLNALTDALLITPVLQL





HTL1A_
8,032
AVLGLEHLPRPPQISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNP


P03362_

VFPVKKANGTWRFIHDLRATNSLTIDLSSSSPGPPDLSSPPTTLAHLQTI


2mutB

DLRDAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWKVLPQGFKNSPTL




FQMQLAHILQPIRQAFPQCTILQYMDDILLASPSHEDLLLLSEATMASLI




SHGLPVSENKTQQTPGTIKFLGQIISPNHLTYDAVPTVPIRSRWALPELQ




ALLGEIQWVSKGTPTLRQPLHSLYCALQPHTDPRDQIYLNPSQVQSLVQL




RQALSQNCRSRLVQTLPLLGAIMLTLTGTTTVVFQSKEQWPLVWLHAPLP




HTSQCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISTQTFNQFIQTSD




HPSVPILLHHSHRFKNLGAQTGELWNTFLKTAAPLAPVKALMPVFTLSPV




IINTAPCLFSDGSTSRAAYILWDKQILSQRSFPLPPPHKSAQRAELLGLL




HGLSSARSWRCLNIFLDSKYLYHYLRTLALGTFQGRSSQAPFQALLPRLL




SRKVVYLHHVRSHTNLPDPISRLNALTDALLITPVLQL





HTL1C_
8,033
AVLGLEHLPRPPEISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNP


P14078

VFPVKKANGTWRFIHDLRATNSLTIDLSSSSPGPPDLSSLPTTLAHLQTI




DLKDAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWRVLPQGFKNSPTL




FEMQLAHILQPIRQAFPQCTILQYMDDILLASPSHADLQLLSEATMASLI




SHGLPVSENKTQQTPGTIKFLGQIISPNHLTYDAVPKVPIRSRWALPELQ




ALLGEIQWVSKGTPTLRQPLHSLYCALQRHTDPRDQIYLNPSQVQSLVQL




RQALSQNCRSRLVQTLPLLGAIMLTLTGTTTVVFQSKQQWPLVWLHAPLP




HTSQCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISTQTFNQFIQTSD




HPSVPILLHHSHRFKNLGAQTGELWNTFLKTTAPLAPVKALMPVFTLSPV




IINTAPCLFSDGSTSQAAYILWDKHILSQRSFPLPPPHKSAQRAELLGLL




HGLSSARSWRCLNIFLDSKYLYHYLRTLALGTFQGRSSQAPFQALLPRLL




SRKVVYLHHVRSHTNLPDPISRLNALTDALLITPVLQL





HTL1C_
8,034
AVLGLEHLPRPPEISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNP


P14078_

VFPVKKANGTWRFIHDLRATNSLTIDLSSSSPGPPDLSSLPTTLAHLQTI


2mut

DLKDAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWRVLPQGFKNSPTL




FQMQLAHILQPIRQAFPQCTILQYMDDILLASPSHADLQLLSEATMASLI




SHGLPVSENKTQQTPGTIKFLGQIISPNHLTYDAVPKVPIRSRWALPELQ




ALLGEIQWVSKGTPTLRQPLHSLYCALQPHTDPRDQIYLNPSQVQSLVQL




RQALSQNCRSRLVQTLPLLGAIMLTLTGTTTVVFQSKQQWPLVWLHAPLP




HTSQCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISTQTFNQFIQTSD




HPSVPILLHHSHRFKNLGAQTGELWNTFLKTTAPLAPVKALMPVFTLSPV




IINTAPCLFSDGSTSQAAYILWDKHILSQRSFPLPPPHKSAQRAELLGLL




HGLSSARSWRCLNIFLDSKYLYHYLRTLALGTFQGRSSQAPFQALLPRLL




SRKVVYLHHVRSHTNLPDPISRLNALTDALLITPVLQL





HTL1L_
8,035
GLEHLPRPPEISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNPVFP


P0C211

VKKANGTWRFIHDLRATNSLTVDLSSSSPGPPDLSSLPTTLAHLQTIDLK




DAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWKVLPQGFKNSPTLFEM




QLASILQPIRQAFPQCVILQYMDDILLASPSPEDLQQLSEATMASLISHG




LPVSQDKTQQTPGTIKFLGQIISPNHITYDAVPTVPIRSRWALPELQALL




GEIQWVSKGTPTLRQPLHSLYCALQGHTDPRDQIYLNPSQVQSLMQLQQA




LSQNCRSRLAQTLPLLGAIMLTLTGTTTVVFQSKQQWPLVWLHAPLPHTS




QCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISIQTFNQFIQTSDHPS




VPILLHHSHRFKNLGAQTGELWNTFLKTAAPLAPVKALTPVFTLSPIIIN




TAPCLFSDGSTSQAAYILWDKHILSQRSFPLPPPHKSAQQAELLGLLHGL




SSARSWHCLNIFLDSKYLYHYLRTLALGTFQGKSSQAPFQALLPRLLAHK




VIYLHHVRSHTNLPDPISKLNALTDALLITPIL





HTL1L_
8,036
GLEHLPRPPEISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNPVFP


P0C211_

VKKANGTWRFIHDLRATNSLTVDLSSSSPGPPDLSSLPTTLAHLQTIDLK


2mut

DAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWKVLPQGFKNSPTLFQM




QLASILQPIRQAFPQCVILQYMDDILLASPSPEDLQQLSEATMASLISHG




LPVSQDKTQQTPGTIKFLGQIISPNHITYDAVPTVPIRSRWALPELQALL




GEIQWVSKGTPTLRQPLHSLYCALQGHTDPRDQIYLNPSQVQSLMQLQQA




LSQNCRSRLAQTLPLLGAIMLTLTGTTTVVFQSKQQWPLVWLHAPLPHTS




QCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISIQTFNQFIQTSDHPS




VPILLHHSHRFKNLGAQTGELWNTFLKTAAPLAPVKALTPVFTLSPIIIN




TAPCLFSDGSTSQAAYILWDKHILSQRSFPLPPPHKSAQQAELLGLLHGL




SSARSWHCLNIFLDSKYLYHYLRTLAWGTFQGKSSQAPFQALLPRLLAHK




VIYLHHVRSHTNLPDPISKLNALTDALLITPIL





HTL1L_
8,037
GLEHLPRPPEISQFPLNPERLQALQHLVRKALEAGHIEPYTGPGNNPVFP


P0C211_

VKKANGTWRFIHDLRATNSLTVDLSSSSPGPPDLSSPPTTLAHLQTIDLK


2mutB

DAFFQIPLPKQFQPYFAFTVPQQCNYGPGTRYAWKVLPQGFKNSPTLFQM




QLASILQPIRQAFPQCVILQYMDDILLASPSPEDLQQLSEATMASLISHG




LPVSQDKTQQTPGTIKFLGQIISPNHITYDAVPTVPIRSRWALPELQALL




GEIQWVSKGTPTLRQPLHSLYCALQGHTDPRDQIYLNPSQVQSLMQLQQA




LSQNCRSRLAQTLPLLGAIMLTLTGTTTVVFQSKQQWPLVWLHAPLPHTS




QCPWGQLLASAVLLLDKYTLQSYGLLCQTIHHNISIQTFNQFIQTSDHPS




VPILLHHSHRFKNLGAQTGELWNTFLKTAAPLAPVKALTPVFTLSPIIIN




TAPCLFSDGSTSQAAYILWDKHILSQRSFPLPPPHKSAQQAELLGLLHGL




SSARSWHCLNIFLDSKYLYHYLRTLAWGTFQGKSSQAPFQALLPRLLAHK




VIYLHHVRSHTNLPDPISKLNALTDALLITPIL





HTL32_
8,038
GLEHLPPPPEVSQFPLNPERLQALTDLVSRALEAKHIEPYQGPGNNPIFP


Q0R5R2

VKKPNGKWRFIHDLRATNSVTRDLASPSPGPPDLTSLPQGLPHLRTIDLT




DAFFQIPLPTIFQPYFAFTLPQPNNYGPGTRYSWRVLPQGFKNSPTLFEQ




QLSHILTPVRKTFPNSLIIQYMDDILLASPAPGELAALTDKVTNALTKEG




LPLSPEKTQATPGPIHFLGQVISQDCITYETLPSINVKSTWSLAELQSML




GELQWVSKGTPVLRSSLHQLYLALRGHRDPRDTIKLTSIQVQALRTIQKA




LTLNCRSRLVNQLPILALIMLRPTGTTAVLFQTKQKWPLVWLHTPHPATS




LRPWGQLLANAVIILDKYSLQHYGQVCKSFHHNISNQALTYYLHTSDQSS




VAILLQHSHRFHNLGAQPSGPWRSLLQMPQIFQNIDVLRPPFTISPVVIN




HAPCLFSDGSASKAAFIIWDRQVIHQQVLSLPSTCSAQAGELFGLLAGLQ




KSQPWVALNIFLDSKFLIGHLRRMALGAFPGPSTQCELHTQLLPLLQGKT




VYVHHVRSHTLLQDPISRLNEATDALMLAPLLPL





HTL32_
8,039
GLEHLPPPPEVSQFPLNPERLQALTDLVSRALEAKHIEPYQGPGNNPIFP


Q0R5R2_

VKKPNGKWRFIHDLRATNSVTRDLASPSPGPPDLTSLPQGLPHLRTIDLT


2mut

DAFFQIPLPTIFQPYFAFTLPQPNNYGPGTRYSWRVLPQGFKNSPTLFQQ




QLSHILTPVRKTFPNSLIIQYMDDILLASPAPGELAALTDKVTNALTKEG




LPLSPEKTQATPGPIHFLGQVISQDCITYETLPSINVKSTWSLAELQSML




GELQWVSKGTPVLRSSLHQLYLALRGHRDPRDTIKLTSIQVQALRTIQKA




LTLNCRSRLVNQLPILALIMLRPTGTTAVLFQTKQKWPLVWLHTPHPATS




LRPWGQLLANAVIILDKYSLQHYGQVCKSFHHNISNQALTYYLHTSDQSS




VAILLQHSHRFHNLGAQPSGPWRSLLQMPQIFQNIDVLRPPFTISPWVIN




HAPCLFSDGSASKAAFIIWDRQVIHQQVLSLPSTCSAQAGELFGLLAGLQ




KSQPWVALNIFLDSKFLIGHLRRMAWGAFPGPSTQCELHTQLLPLLQGKT




VYVHHVRSHTLLQDPISRLNEATDALMLAPLLPL





HTL32_
8,040
GLEHLPPPPEVSQFPLNPERLQALTDLVSRALEAKHIEPYQGPGNNPIFP


Q0R5R2_

VKKPNGKWRFIHDLRATNSVTRDLASPSPGPPDLTSPPQGLPHLRTIDLT


2mutB

DAFFQIPLPTIFQPYFAFTLPQPNNYGPGTRYSWRVLPQGFKNSPTLFQQ




QLSHILTPVRKTFPNSLIIQYMDDILLASPAPGELAALTDKVTNALTKEG




LPLSPEKTQATPGPIHFLGQVISQDCITYETLPSINVKSTWSLAELQSML




GELQWVSKGTPVLRSSLHQLYLALRGHRDPRDTIKLTSIQVQALRTIQKA




LTLNCRSRLVNQLPILALIMLRPTGTTAVLFQTKQKWPLVWLHTPHPATS




LRPWGQLLANAVIILDKYSLQHYGQVCKSFHHNISNQALTYYLHTSDQSS




VAILLQHSHRFHNLGAQPSGPWRSLLQMPQIFQNIDVLRPPFTISPVVIN




HAPCLFSDGSASKAAFIIWDRQVIHQQVLSLPSTCSAQAGELFGLLAGLQ




KSQPWVALNIFLDSKFLIGHLRRMAWGAFPGPSTQCELHTQLLPLLQGKT




VYVHHVRSHTLLQDPISRLNEATDALMLAPLLPL





HTL3P_
8,041
GLEHLPPPPEVSQFPLNPERLQALTDLVSRALEAKHIEPYQGPGNNPIFP


Q4U0X6

VKKPNGKWRFIHDLRATNSLTRDLASPSPGPPDLTSLPQDLPHLRTIDLT




DAFFQIPLPAVFQPYFAFTLPQPNNHGPGTRYSWRVLPQGFKNSPTLFEQ




QLSHILAPVRKAFPNSLIIQYMDDILLASPALRELTALTDKVTNALTKEG




LPMSLEKTQATPGSIHFLGQVISPDCITYETLPSIHVKSIWSLAELQSML




GELQWVSKGTPVLRSSLHQLYLALRGHRDPRDTIELTSTQVQALKTIQKA




LALNCRSRLVSQLPILALIILRPTGTTAVLFQTKQKWPLVWLHTPHPATS




LRPWGQLLANAIITLDKYSLQHYGQICKSFHHNISNQALTYYLHTSDQSS




VAILLQHSHRFHNLGAQPSGPWRSLLQVPQIFQNIDVLRPPFIISPVVID




HAPCLFSDGATSKAAFILWDKQVIHQQVLPLPSTCSAQAGELFGLLAGLQ




KSKPWPALNIFLDSKFLIGHLRRMALGAFLGPSTQCDLHARLFPLLQGKT




VYVHHVRSHTLLQDPISRLNEATDALMLAPLLPL





HTL3P_
8,042
GLEHLPPPPEVSQFPLNPERLQALTDLVSRALEAKHIEPYQGPGNNPIFP


Q4U0X6_

VKKPNGKWRFIHDLRATNSLTRDLASPSPGPPDLTSLPQDLPHLRTIDLT


2mut

DAFFQIPLPAVFQPYFAFTLPQPNNHGPGTRYSWRVLPQGFKNSPTLFQQ




QLSHILAPVRKAFPNSLIIQYMDDILLASPALRELTALTDKVTNALTKEG




LPMSLEKTQATPGSIHFLGQVISPDCITYETLPSIHVKSIWSLAELQSML




GELQWVSKGTPVLRSSLHQLYLALRGHRDPRDTIELTSTQVQALKTIQKA




LALNCRSRLVSQLPILALIILRPTGTTAVLFQTKQKWPLVWLHTPHPATS




LRPWGQLLANAIITLDKYSLQHYGQICKSFHHNISNQALTYYLHTSDQSS




VAILLQHSHRFHNLGAQPSGPWRSLLQVPQIFQNIDVLRPPFIISPVVID




HAPCLFSDGATSKAAFILWDKQVIHQQVLPLPSTCSAQAGELFGLLAGLQ




KSKPWPALNIFLDSKFLIGHLRRMAWGAFLGPSTQCDLHARLFPLLQGKT




VYVHHVRSHTLLQDPISRLNEATDALMLAPLLPL





HTL3P_
8,043
GLEHLPPPPEVSQFPLNPERLQALTDLVSRALEAKHIEPYQGPGNNPIFP


Q4U0X6_

VKKPNGKWRFIHDLRATNSLTRDLASPSPGPPDLTSPPQDLPHLRTIDLT


2mutB

DAFFQIPLPAVFQPYFAFTLPQPNNHGPGTRYSWRVLPQGFKNSPTLFQQ




QLSHILAPVRKAFPNSLIIQYMDDILLASPALRELTALTDKVTNALTKEG




LPMSLEKTQATPGSIHFLGQVISPDCITYETLPSIHVKSIWSLAELQSML




GELQWVSKGTPVLRSSLHQLYLALRGHRDPRDTIELTSTQVQALKTIQKA




LALNCRSRLVSQLPILALIILRPTGTTAVLFQTKQKWPLVWLHTPHPATS




LRPWGQLLANAIITLDKYSLQHYGQICKSFHHNISNQALTYYLHTSDQSS




VAILLQHSHRFHNLGAQPSGPWRSLLQVPQIFQNIDVLRPPFIISPVVID




HAPCLFSDGATSKAAFILWDKQVIHQQVLPLPSTCSAQAGELFGLLAGLQ




KSKPWPALNIFLDSKFLIGHLRRMAWGAFLGPSTQCDLHARLFPLLQGKT




VYVHHVRSHTLLQDPISRLNEATDALMLAPLLPL





HTLV2_
8,044
HLPPPPQVDQFPLNLPERLQALNDLVSKALEAGHIEPYSGPGNNPVFPVK


P03363_

KPNGKWRFIHDLRATNAITTTLTSPSPGPPDLTSLPTALPHLQTIDLTDA


2mut

FFQIPLPKQYQPYFAFTIPQPCNYGPGTRYAWTVLPQGFKNSPTLFQQQL




AAVLNPMRKMFPTSTIVQYMDDILLASPTNEELQQLSQLTLQALTTHGLP




ISQEKTQQTPGQIRFLGQVISPNHITYESTPTIPIKSQWTLTELQVILGE




IQWVSKGTPILRKHLQSLYSALHPYRDPRACITLTPQQLHALHAIQQALQ




HNCRGRLNPALPLLGLISLSTSGTTSVIFQPKQNWPLAWLHTPHPPTSLC




PWGHLLACTILTLDKYTLQHYGQLCQSFHHNMSKQALCDFLRNSPHPSVG




ILIHHMGRFHNLGSQPSGPWKTLLHLPTLLQEPRLLRPIFTLSPVVLDTA




PCLFSDGSPQKAAYVLWDQTILQQDITPLPSHETHSAQKGELLALICGLR




AAKPWPSLNIFLDSKYLIKYLHSLAIGAFLGTSAHQTLQAALPPLLQGKT




IYLHHVRSHTNLPDPISTFNEYTDSLILAPLVPL





JSRV_
8,045
PLGTSDSPVTHADPIDWKSEEPVWVDQWPLTQEKLSAAQQLVQEQLRLGH


P31623

IEPSTSAWNSPIFVIKKKSGKWRLLQDLRKVNETMMHMGALQPGLPTPSA




IPDKSYIIVIDLKDCFYTIPLAPQDCKRFAFSLPSVNFKEPMQRYQWRVL




PQGMTNSPTLCQKFVATAIAPVRQRFPQLYLVHYMDDILLAHTDEHLLYQ




AFSILKQHLSLNGLVIADEKIQTHFPYNYLGFSLYPRVYNTQLVKLQTDH




LKTLNDFQKLLGDINWIRPYLKLPTYTLQPLFDILKGDSDPASPRTLSLE




GRTALQSIEEAIRQQQITYCDYQRSWGLYILPTPRAPTGVLYQDKPLRWI




YLSATPTKHLLPYYELVAKIIAKGRHEAIQYFGMEPPFICVPYALEQQDW




LFQFSDNWSIAFANYPGQITHHYPSDKLLQFASSHAFIFPKIVRRQPIPE




ATLIFTDGSSNGTAALIINHQTYYAQTSFSSAQVVELFAVHQALLTVPTS




FNLFTDSSYVVGALQMIETVPIIGTTSPEVLNLFTLIQQVLHCRQHPCFF




GHIRAHSTLPGALVQGNHTADVLTKQVFFQS





JSRV_
8,046
PLGTSDSPVTHADPIDWKSEEPVWVDQWPLTQEKLSAAQQLVQEQLRLGH


P31623_

IEPSTSAWNSPIFVIKKKSGKWRLLQDLRKVNETMMHMGALQPGLPTPSP


2mutB

IPDKSYIIVIDLKDCFYTIPLAPQDCKRFAFSLPSVNFKEPMQRYQWRVL




PQGMTNSPTLCQKFVATAIAPVRQRFPQLYLVHYMDDILLAHTDEHLLYQ




AFSILKQHLSLNGLVIADEKIQTHFPYNYLGFSLYPRVYNTQLVKLQTDH




LKTLNDFQKLLGDINWIRPYLKLPTYTLQPLFDILKGDSDPASPRTLSLE




GRTALQSIEEAIRQQQITYCDYQRSWGLYILPTPRAPTGVLYQDKPLRWI




YLSATPTKHLLPYYELVAKIIAKGRHEAIQYFGMEPPFICVPYALEQQDW




LFQFSDNWSIAFANYPGQITHHYPSDKLLQFASSHAFIFPKIVRRQPIPE




ATLIFTDGSSNGTAALIINHQTYYAQTSFSSAQVVELFAVHQALLTVPTS




FNLFTDSSYVVGALQMIETVPIIGTTSPEVLNLFTLIQQVLHCRQHPCFF




GHIRAHSTLPGALVQGNHTADVLTKQVFFQS





KORV_
8,047
TLGDQGSRGSDPLPEPRVTLTVEGIPTEFLVNTGAEHSVLTKPMGKMGSK


Q9TTC1

RTVVAGATGSKVYPWTTKRLLKIGQKQVTHSFLVIPECPAPLLGRDLLTK




LKAQIQFSTEGPQVTWEDRPAMCLVLNLEEEYRLHEKPVPPSIDPSWLQL




FPMVWAEKAGMGLANQVPPVVVELKSDASPVAVRQYPMSKEAREGIRPHI




QRFLDLGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNKRVQDIHPT




VPNPYNLLSSLPPSHTWYSVLDLKDAFFCLKLHPNSQPLFAFEWRDPEKG




NTGQLTWTRLPQGFKNSPTLFDEALHRDLASFRALNPQVVMLQYVDDLLV




AAPTYRDCKEGTRRLLQELSKLGYRVSAKKAQLCREEVTYLGYLLKGGKR




WLTPARKATVMKIPTPTTPRQVREFLGTAGFCRLWIPGFASLAAPLYPLT




REKVPFTWTEAHQEAFGRIKEALLSAPALALPDLTKPFALYVDEKEGVAR




GVLTQTLGPWRRPVAYLSKKLDPVASGWPTCLKAIAAVALLLKDADKLTL




GQNVLVIAPHNLESIVRQPPDRWMTNARMTHYQSLLLNERVSFAPPAILN




PATLLPVESDDTPIHICSEILAEETGTRPDLRDQPLPGVPAWYTDGSSFI




MDGRRQAGAAIVDNKRTVWASNLPEGTSAQKAELIALTQALRLAEGKSIN




IYTDSRYAFATAHVHGAIYKQRGLLTSAGKDIKNKEEILALLEAIHLPKR




VAIIHCPGHQRGTDPVATGNRKADEAAKQAAQSTRILTETTKN





KORV_
8,048
TLGDQGSRGSDPLPEPRVTLTVEGIPTEFLVNTGAEHSVLTKPMGKMGSK


Q9TTC1_

RTVVAGATGSKVYPWTTKRLLKIGQKQVTHSFLVIPECPAPLLGRDLLTK


3mut

LKAQIQFSTEGPQVTWEDRPAMCLVLNLEEEYRLHEKPVPPSIDPSWLQL




FPMVWAEKAGMGLANQVPPVVVELKSDASPVAVRQYPMSKEAREGIRPHI




QRFLDLGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNKRVQDIHPT




VPNPYNLLSSLPPSHTWYSVLDLKDAFFCLKLHPNSQPLFAFEWRDPEKG




NTGQLTWTRLPQGFKNSPTLFNEALHRDLASFRALNPQVVMLQYVDDLLV




AAPTYRDCKEGTRRLLQELSKLGYRVSAKKAQLCREEVTYLGYLLKGGKR




WLTPARKATVMKIPTPTTPRQVREFLGTAGFCRLWIPGFASLAAPLYPLT




RPKVPFTWTEAHQEAFGRIKEALLSAPALALPDLTKPFALYVDEKEGVAR




GVLTQTLGPWRRPVAYLSKKLDPVASGWPTCLKAIAAVALLLKDADKLTL




GQNVLVIAPHNLESIVRQPPDRWMTNARMTHYQSLLLNERVSFAPPAILN




PATLLPVESDDTPIHICSEILAEETGTRPDLRDQPLPGVPAWYTDGSSFI




MDGRRQAGAAIVDNKRTVWASNLPEGTSAQKAELIALTQALRLAEGKSIN




IYTDSRYAFATAHVHGAIYKQRGWLTSAGKDIKNKEEILALLEAIHLPKR




VAIIHCPGHQRGTDPVATGNRKADEAAKQAAQSTRILTETTKN





KORV_
8,049
TLGDQGSRGSDPLPEPRVTLTVEGIPTEFLVNTGAEHSVLTKPMGKMGSK


Q9TTC1_

RTVVAGATGSKVYPWTTKRLLKIGQKQVTHSFLVIPECPAPLLGRDLLTK


3mutA

LKAQIQFSTEGPQVTWEDRPAMCLVLNLEEEYRLHEKPVPPSIDPSWLQL




FPMVWAEKAGMGLANQVPPVVVELKSDASPVAVRQYPMSKEAREGIRPHI




QRFLDLGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNKRVQDIHPT




VPNPYNLLSSLPPSHTWYSVLDLKDAFFCLKLHPNSQPLFAFEWRDPEKG




NTGQLTWTRLPQGFKNSPTLFNEALHRDLASFRALNPQVVMLQYVDDLLV




AAPTYRDCKEGTRRLLQELSKLGYRVSAKKAQLCREEVTYLGYLLKGGKR




WLTPARKATVMKIPTPTTPRQVREFLGKAGFCRLFIPGFASLAAPLYPLT




RPKVPFTWTEAHQEAFGRIKEALLSAPALALPDLTKPFALYVDEKEGVAR




GVLTQTLGPWRRPVAYLSKKLDPVASGWPTCLKAIAAVALLLKDADKLTL




GQNVLVIAPHNLESIVRQPPDRWMTNARMTHYQSLLLNERVSFAPPAILN




PATLLPVESDDTPIHICSEILAEETGTRPDLRDQPLPGVPAWYTDGSSFI




MDGRRQAGAAIVDNKRTVWASNLPEGTSAQKAELIALTQALRLAEGKSIN




IYTDSRYAFATAHVHGAIYKQRGWLTSAGKDIKNKEEILALLEAIHLPKR




VAIIHCPGHQRGTDPVATGNRKADEAAKQAAQSTRILTETTKN





KORV_
8,050
LLGRDLLTKLKAQIQFSTEGPQVTWEDRPAMCLVLNLEEEYRLHEKPVPP


Q9TTC1-

SIDPSWLQLFPMVWAEKAGMGLANQVPPVVVELKSDASPVAVRQYPMSKE


Pro

AREGIRPHIQRFLDLGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVN




KRVQDIHPTVPNPYNLLSSLPPSHTWYSVLDLKDAFFCLKLHPNSQPLFA




FEWRDPEKGNTGQLTWTRLPQGFKNSPTLFDEALHRDLASFRALNPQVVM




LQYVDDLLVAAPTYRDCKEGTRRLLQELSKLGYRVSAKKAQLCREEVTYL




GYLLKGGKRWLTPARKATVMKIPTPTTPRQVREFLGTAGFCRLWIPGFAS




LAAPLYPLTREKVPFTWTEAHQEAFGRIKEALLSAPALALPDLTKPFALY




VDEKEGVARGVLTQTLGPWRRPVAYLSKKLDPVASGWPTCLKAIAAVALL




LKDADKLTLGQNVLVIAPHNLESIVRQPPDRWMTNARMTHYQSLLLNERV




SFAPPAILNPATLLPVESDDTPIHICSEILAEETGTRPDLRDQPLPGVPA




WYTDGSSFIMDGRRQAGAAIVDNKRTVWASNLPEGTSAQKAELIALTQAL




RLAEGKSINIYTDSRYAFATAHVHGAIYKQRGLLTSAGKDIKNKEEILAL




LEAIHLPKRVAIIHCPGHQRGTDPVATGNRKADEAAKQAAQSTRILTETT




KN





KORV_
8,051
LLGRDLLTKLKAQIQFSTEGPQVTWEDRPAMCLVLNLEEEYRLHEKPVPP


Q9TTC1-

SIDPSWLQLFPMVWAEKAGMGLANQVPPVVVELKSDASPVAVRQYPMSKE


Pro_

AREGIRPHIQRFLDLGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVN


3mut

KRVQDIHPTVPNPYNLLSSLPPSHTWYSVLDLKDAFFCLKLHPNSQPLFA




FEWRDPEKGNTGQLTWTRLPQGFKNSPTLFNEALHRDLASFRALNPQVVM




LQYVDDLLVAAPTYRDCKEGTRRLLQELSKLGYRVSAKKAQLCREEVTYL




GYLLKGGKRWLTPARKATVMKIPTPTTPRQVREFLGTAGFCRLWIPGFAS




LAAPLYPLTRPKVPFTWTEAHQEAFGRIKEALLSAPALALPDLTKPFALY




VDEKEGVARGVLTQTLGPWRRPVAYLSKKLDPVASGWPTCLKAIAAVALL




LKDADKLTLGQNVLVIAPHNLESIVRQPPDRWMTNARMTHYQSLLLNERV




SFAPPAILNPATLLPVESDDTPIHICSEILAEETGTRPDLRDQPLPGVPA




WYTDGSSFIMDGRRQAGAAIVDNKRTVWASNLPEGTSAQKAELIALTQAL




RLAEGKSINIYTDSRYAFATAHVHGAIYKQRGWLTSAGKDIKNKEEILAL




LEAIHLPKRVAIIHCPGHQRGTDPVATGNRKADEAAKQAAQSTRILTETT




KN





KORV_
8,052
LLGRDLLTKLKAQIQFSTEGPQVTWEDRPAMCLVLNLEEEYRLHEKPVPP


Q9TTC1-

SIDPSWLQLFPMVWAEKAGMGLANQVPPVVVELKSDASPVAVRQYPMSKE


Pro_

AREGIRPHIQRFLDLGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVN


3mutA

KRVQDIHPTVPNPYNLLSSLPPSHTWYSVLDLKDAFFCLKLHPNSQPLFA




FEWRDPEKGNTGQLTWTRLPQGFKNSPTLFNEALHRDLASFRALNPQVVM




LQYVDDLLVAAPTYRDCKEGTRRLLQELSKLGYRVSAKKAQLCREEVTYL




GYLLKGGKRWLTPARKATVMKIPTPTTPRQVREFLGKAGFCRLFIPGFAS




LAAPLYPLTRPKVPFTWTEAHQEAFGRIKEALLSAPALALPDLTKPFALY




VDEKEGVARGVLTQTLGPWRRPVAYLSKKLDPVASGWPTCLKAIAAVALL




LKDADKLTLGQNVLVIAPHNLESIVRQPPDRWMTNARMTHYQSLLLNERV




SFAPPAILNPATLLPVESDDTPIHICSEILAEETGTRPDLRDQPLPGVPA




WYTDGSSFIMDGRRQAGAAIVDNKRTVWASNLPEGTSAQKAELIALTQAL




RLAEGKSINIYTDSRYAFATAHVHGAIYKQRGWLTSAGKDIKNKEEILAL




LEAIHLPKRVAIIHCPGHQRGTDPVATGNRKADEAAKQAAQSTRILTETT




KN





MLVAV_
8,053
TLNLEDEYRLYETSAEPEVSPGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03356

PLKATSTPVSIKQYPMSQEAKLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHRWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLLTLGNL




GYRASAKKAQLCQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLRKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVAV_
8,054
TLNLEDEYRLYETSAEPEVSPGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03356_

PLKATSTPVSIKQYPMSQEAKLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHRWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLLTLGNL




GYRASAKKAQLCQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLRKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPWVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVAV_
8,055
TLNLEDEYRLYETSAEPEVSPGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03356_

PLKATSTPVSIKQYPMSQEAKLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHRWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLLTLGNL




GYRASAKKAQLCQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLRKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVBM_
8,056
TLGIEDEYRLHETSTEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


Q7SVK7

PLKATSTPVSIQQYPMSHEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPVPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFSWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAG




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVBM_
8,057
TLGIEDEYRLHETSTEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


Q7SVK7

PLKATSTPVSIQQYPMSHEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPVPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFSWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAG




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVBM_
8,058
TLGIEDEYRLHETSTEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


Q7SVK7_

PLKATSTPVSIQQYPMSHEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPVPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFSWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPWVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAG




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVBM_
8,059
TLGIEDEYRLHETSTEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


Q7SVK7_

PLKATSTPVSIQQYPMSHEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPVPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFSWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILA




ETHGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAG




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVBM_
8,060
LGIEDEYRLHETSTEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLIIP


Q7SVK7_

LKATSTPVSIQQYPMSHEARLGIKPHIQRLLDQGILVPCQSPWNTPLLPV


3mutAWS

KKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLDL




KDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFNE




ALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLQTLGDLG




YRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPVPKTPRQLR




EFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFSWGPDQQKAYQEIKQAL




LTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLDP




VAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDRW




LSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILAE




THGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAGA




LPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRRR




GWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNRL




ADQAAREAAIKTPPDTSTLLI





MLVBM_
8,061
LGIEDEYRLHETSTEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLIIP


Q7SVK7_

LKATSTPVSIQQYPMSHEARLGIKPHIQRLLDQGILVPCQSPWNTPLLPV


3mutAWS

KKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLDL




KDAFFCLRLHPTSQPLFAFEWRDPGMGISGQLTWTRLPQGFKNSPTLFNE




ALHRDLADFRIQHPDLILLQYVDDILLAATSELDCQQGTRALLQTLGDLG




YRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPVPKTPRQLR




EFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFSWGPDQQKAYQEIKQAL




LTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLDP




VAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDRW




LSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEEGAPHDCLEILAE




THGTRPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAGA




LPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYRRR




GWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNRL




ADQAAREAAIKTPPDTSTLLI





MLVCB_
8,062
TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P08361

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLAGFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPIPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFNWGPDQQKAFQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEGLQHDCLDILA




EAHGTRSDLMDQPLPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNSAEARGNR




MADQAAREVATRETPETSTLL





MLVCB_
8,063
TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P08361_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLAGFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPIPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAFQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHDCLDILA




EAHGTRSDLMDQPLPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNSAEARGNR




MADQAAREVATRETPETSTLL





MLVCB_
8,064
TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P08361_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLAGFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPIPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAFQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHDCLDILA




EAHGTRSDLMDQPLPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNSAEARGNR




MADQAAREVATRETPETSTLL





MLVF5_
8,065
TLNIEDEYRLHETSKGPDVPLGSTWLSDFPQAWAETGGMGLAFRQAPLII


P26810

SLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQSLFAFEWKDPEMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGLCRLWIPGFAEMAAPLYPLTKTGTLFKWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDVGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPIVALNPATLLPLPEEGLQHDCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSFLQEGQRRAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAAGKKLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNHAEARGNR




MADQAAREVATRETPETSTLL





MLVF5_
8,066
TLNIEDEYRLHETSKGPDVPLGSTWLSDFPQAWAETGGMGLAFRQAPLII


P26810_

SLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQSLFAFEWKDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGLCRLWIPGFAEMAAPLYPLTKPGTLFKWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDVGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPIVALNPATLLPLPEEGLQHDCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSFLQEGQRRAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAAGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNHAEARGNR




MADQAAREVATRETPETSTLL





MLVF5_
8,067
TLNIEDEYRLHETSKGPDVPLGSTWLSDFPQAWAETGGMGLAFRQAPLII


P26810_

SLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQSLFAFEWKDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGLCRLFIPGFAEMAAPLYPLTKPGTLFKWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDVGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPIVALNPATLLPLPEEGLQHDCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSFLQEGQRRAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAAGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNHAEARGNR




MADQAAREVATRETPETSTLL





MLVFF_
8,068
TLNIEDEYRLHETSKGPDVPLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P26809_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQSLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFEWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPIVALNPATLLPLPEEGLQHDCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVVWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNRAEARGNR




MADQAAREVATRETPETSTLL





MLVFF_
8,069
TLNIEDEYRLHETSKGPDVPLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P26809_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQSLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGDL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFEWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPIVALNPATLLPLPEEGLQHDCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVVWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGNRAEARGNR




MADQAAREVATRETPETSTLL





MLVMS_
8,070
TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLL





MLVMS_
8,137
TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


reference

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLLIENSSP





MLVMS_
8,071
TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLL





MLVMS_
8,072
TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLL





MLVMS_
8,073
TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLL





MLVMS_
8,074
TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA_

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD


WS

LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAY




LSKKLDPVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALV




KQPPDRWLSNARMTHYQALLLDTDRVQFGPWVALNPATLLPLPEEGLQHN




CLDILAEAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTET




EVIWAKALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIH




GEIYRRRGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSA




EARGNRMADQAARKAAITETPDTSTLL





MLVMS_
8,075
TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA_

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD


WS

LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLL





MLVMS_
8,076
TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


PLV919

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLLIENSSPSGGSKRTADGSEFE





MLVMS_
8,077
TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLII


P03355_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


PLV919

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEGLQHNCLDILA




EAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAK




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR




RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNR




MADQAARKAAITETPDTSTLLIENSSPSGGSKRTADGSEFE





MLVRD_
8,078
TLNIEDEYRLHEISTEPDVSPGSTWLSDFPQAWAETGGMGLAVRQAPLII


P11227

PLKATSTPVSIKQYPMSQEAKLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQGLREVNKRVEDIHPTVPNPYNLLSGLPTSHRWYTVLD




LKDAFFCLRLHPTSQPLFASEWRDPGMGISGQLTWTRLPQGFKNSPTLFD




EALHRGLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLKTLGNL




GYRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPRFAEMAAPLYPLTKTGTLFNWGPDQQKAYHEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPWVALNPATLLPLPEEGAPHDCLEILA




ETHGTEPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYKR




RGLLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MLVRD_
8,079
TLNIEDEYRLHEISTEPDVSPGSTWLSDFPQAWAETGGMGLAVRQAPLII


P11227_

PLKATSTPVSIKQYPMSQEAKLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQGLREVNKRVEDIHPTVPNPYNLLSGLPTSHRWYTVLD




LKDAFFCLRLHPTSQPLFASEWRDPGMGISGQLTWTRLPQGFKNSPTLFN




EALHRGLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLKTLGNL




GYRASAKKAQICQKQVKYLGYLLREGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPRFAEMAAPLYPLTKPGTLFNWGPDQQKAYHEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPWVALNPATLLPLPEEGAPHDCLEILA




ETHGTEPDLTDQPIPDADHTWYTDGSSFLQEGQRKAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKRLNVYTDSRYAFATAHIHGEIYKR




RGWLTSEGREIKNKSEILALLKALFLPKRLSIIHCLGHQKGDSAEARGNR




LADQAAREAAIKTPPDTSTLL





MMTVB_
8,080
WVQEISDSRPMLHIYLNGRRFLGLLNTGADKTCIAGRDWPANWPIHQTES


P03365

SLQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDI




KVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQ




ALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMH




DMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSP




NFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMD




DILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQG




DSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILN




GDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYT




PTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDP




DYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTA




IIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQA




EIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHL




QRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,081
WVQEISDSRPMLHIYLNGRRFLGLLNTGADKTCIAGRDWPANWPIHQTES


P03365

SLQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDI




KVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQ




ALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMH




DMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSP




NFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMD




DILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQG




DSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILN




GDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYT




PTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDP




DYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTA




IIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQA




EIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHL




QRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,082
WVQEISDSRPMLHIYLNGRRFLGLLNTGADKTCIAGRDWPANWPIHQTES


P03365_

SLQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDI


2mut

KVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQ




ALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMH




DMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSP




NFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMD




DILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQG




DSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILN




PDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYT




PTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDP




DYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTA




IIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQA




EIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHL




QRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,083
VQEISDSRPMLHIYLNGRRFLGLLDTGADKTCIAGRDWPANWPIHQTESS


P03365_

LQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDIK


2mut_

VRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQA


WS

LQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMHD




MGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSPN




FKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMDD




ILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQGD




SVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILNP




DSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYTP




TACLWQDGWVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDPD




YIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTAI




IFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQAE




IVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHLQ




RLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILTA





MMTVB_
8,084
VQEISDSRPMLHIYLNGRRFLGLLDTGADKTCIAGRDWPANWPIHQTESS


P03365_

LQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDIK


2mut_

VRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQA


WS

LQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMHD




MGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSPN




FKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMDD




ILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQGD




SVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILNP




DSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYTP




TACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDPD




YIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTAI




IFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQAE




IVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHLQ




RLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILTA





MMTVB_
8,085
WVQEISDSRPMLHIYLNGRRFLGLLNTGADKTCIAGRDWPANWPIHQTES


P03365_

SLQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDI


2mutB

KVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQ




ALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMH




DMGALQPGLPSPVAPPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSP




NFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMD




DILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQG




DSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILN




PDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYT




PTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDP




DYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTA




IIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQA




EIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHL




QRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,086
WVQEISDSRPMLHIYLNGRRFLGLLNTGADKTCIAGRDWPANWPIHQTES


P03365_

SLQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDI


2mutB

KVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQ




ALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMH




DMGALQPGLPSPVAPPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSP




NFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMD




DILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQG




DSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILN




PDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYT




PTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDP




DYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTA




IIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQA




EIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHL




QRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,087
VQEISDSRPMLHIYLNGRRFLGLLDTGADKTCIAGRDWPANWPIHQTESS


P03365_

LQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDIK


2mutB_

VRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQA


WS

LQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMHD




MGALQPGLPSPPAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSPN




FKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMDD




ILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQGD




SVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILNP




DSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYTP




TACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDPD




YIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTAI




IFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQAE




IVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHLQ




RLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILTA





MMTVB_
8,088
VQEISDSRPMLHIYLNGRRFLGLLDTGADKTCIAGRDWPANWPIHQTESS


P03365_

LQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDIK


2mutB_

VRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQA


WS

LQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMHD




MGALQPGLPSPPAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSPN




FKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMDD




ILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQGD




SVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILNP




DSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYTP




TACLWQDGWVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDPD




YIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTAI




IFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQAE




IVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHLQ




RLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILTA





MMTVB_
8,089
VQEISDSRPMLHIYLNGRRFLGLLDTGADKTCIAGRDWPANWPIHQTESS


P03365_

LQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDIK


WS

VRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQA




LQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMHD




MGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSPN




FKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMDD




ILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQGD




SVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILNG




DSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYTP




TACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDPD




YIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTAI




IFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQAE




IVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHLQ




RLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILTA





MMTVB_
8,090
VQEISDSRPMLHIYLNGRRFLGLLDTGADKTCIAGRDWPANWPIHQTESS


P03365_

LQGLGMACGVARSSQPLRWQHEDKSGIIHPFVIPTLPFTLWGRDIMKDIK


WS

VRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQWPLKQEKLQA




LQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDLRAVNATMHD




MGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKRFAFSVPSPN




FKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQDSYIVHYMDD




ILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLKYLGTHIQGD




SVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGELKPLFEILNG




DSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSLCILKTEYTP




TACLWQDGWVEWIHLPHISPKVITPYDIFCTQLIIKGRHRSKELFSKDPD




YIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPLLTFTLQTAI




IFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKENTQNTAQQAE




IVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTKIYTELKHLQ




RLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILTA





MMTVB_
8,091
GRDIMKDIKVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQW


P03365-

PLKQEKLQALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDL


Pro

RAVNATMHDMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKR




FAFSVPSPNFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQD




SYIVHYMDDILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLK




YLGTHIQGDSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGEL




KPLFEILNGDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSL




CILKTEYTPTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRS




KELFSKDPDYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPL




LTFTLQTAIIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKEN




TQNTAQQAEIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTK




IYTELKHLQRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,092
GRDIMKDIKVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQW


P03365-

PLKQEKLQALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDL


Pro

RAVNATMHDMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKR




FAFSVPSPNFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQD




SYIVHYMDDILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLK




YLGTHIQGDSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGEL




KPLFEILNGDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSL




CILKTEYTPTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRS




KELFSKDPDYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPL




LTFTLQTAIIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKEN




TQNTAQQAEIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTK




IYTELKHLQRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,093
GRDIMKDIKVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQW


P03365-

PLKQEKLQALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDL




RAVNATMHDMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKR




FAFSVPSPNFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQD




SYIVHYMDDILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLK




YLGTHIQGDSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGEL




KPLFEILNPDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSL




CILKTEYTPTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRS




KELFSKDPDYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPL




LTFTLQTAIIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKEN




TQNTAQQAEIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTK




IYTELKHLQRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,094
GRDIMKDIKVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQW


P03365-

PLKQEKLQALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDL


Pro_

RAVNATMHDMGALQPGLPSPVAVPKGWEIIIIDLQDCFFNIKLHPEDCKR


2mut

FAFSVPSPNFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQD




SYIVHYMDDILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLK




YLGTHIQGDSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGEL




KPLFEILNPDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSL




CILKTEYTPTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRS




KELFSKDPDYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPL




LTFTLQTAIIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKEN




TQNTAQQAEIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTK




IYTELKHLQRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,095
GRDIMKDIKVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQW


P03365-

PLKQEKLQALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDL


Pro_

RAVNATMHDMGALQPGLPSPVAPPKGWEIIIIDLQDCFFNIKLHPEDCKR


2mutB

FAFSVPSPNFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQD




SYIVHYMDDILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLK




YLGTHIQGDSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGEL




KPLFEILNPDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSL




CILKTEYTPTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRS




KELFSKDPDYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPL




LTFTLQTAIIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKEN




TQNTAQQAEIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTK




IYTELKHLQRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MMTVB_
8,096
GRDIMKDIKVRLMTDSPDDSQDLMIGAIESNLFADQISWKSDQPVWLNQW


P03365-

PLKQEKLQALQQLVTEQLQLGHLEESNSPWNTPVFVIKKKSGKWRLLQDL


Pro_

RAVNATMHDMGALQPGLPSPVAPPKGWEIIIIDLQDCFFNIKLHPEDCKR


2mutB

FAFSVPSPNFKRPYQRFQWKVLPQGMKNSPTLCQKFVDKAILTVRDKYQD




SYIVHYMDDILLAHPSRSIVDEILTSMIQALNKHGLVVSTEKIQKYDNLK




YLGTHIQGDSVSYQKLQIRTDKLRTLNDFQKLLGNINWIRPFLKLTTGEL




KPLFEILNPDSNPISTRKLTPEACKALQLMNERLSTARVKRLDLSQPWSL




CILKTEYTPTACLWQDGVVEWIHLPHISPKVITPYDIFCTQLIIKGRHRS




KELFSKDPDYIVVPYTKVQFDLLLQEKEDWPISLLGFLGEVHFHLPKDPL




LTFTLQTAIIFPHMTSTTPLEKGIVIFTDGSANGRSVTYIQGREPIIKEN




TQNTAQQAEIVAVITAFEEVSQPFNLYTDSKYVTGLFPEIETATLSPRTK




IYTELKHLQRLIHKRQEKFYIGHIRGHTGLPGPLAQGNAYADSLTRILT





MPMV_
8,097
LTAAIDILAPQQCAEPITWKSDEPVWVDQWPLTNDKLAAAQQLVQEQLEA


P07572

GHITESSSPWNTPIFVIKKKSGKWRLLQDLRAVNATMVLMGALQPGLPSP




VAIPQGYLKIIIDLKDCFFSIPLHPSDQKRFAFSLPSTNFKEPMQRFQWK




VLPQGMANSPTLCQKYVATAIHKVRHAWKQMYIIHYMDDILIAGKDGQQV




LQCFDQLKQELTAAGLHIAPEKVQLQDPYTYLGFELNGPKITNQKAVIRK




DKLQTLNDFQKLLGDINWLRPYLKLTTGDLKPLFDTLKGDSDPNSHRSLS




KEALASLEKVETAIAEQFVTHINYSLPLIFLIFNTALTPTGLFWQDNPIM




WIHLPASPKKVLLPYYDAIADLIILGRDHSKKYFGIEPSTIIQPYSKSQI




DWLMQNTEMWPIACASFVGILDNHYPPNKLIQFCKLHTFVFPQIISKTPL




NNALLVFTDGSSTGMAAYTLTDTTIKFQTNLNSAQLVELQALIAVLSAFP




NQPLNIYTDSAYLAHSIPLLETVAQIKHISETAKLFLQCQQLIYNRSIPF




YIGHVRAHSGLPGPIAQGNQRADLATKIVASNINT





MPMV_
8,098
LTAAIDILAPQQCAEPITWKSDEPVWVDQWPLTNDKLAAAQQLVQEQLEA


P07572_

GHITESSSPWNTPIFVIKKKSGKWRLLQDLRAVNATMVLMGALQPGLPSP


2mutB

VAPPQGYLKIIIDLKDCFFSIPLHPSDQKRFAFSLPSTNFKEPMQRFQWK




VLPQGMANSPTLCQKYVATAIHKVRHAWKQMYIIHYMDDILIAGKDGQQV




LQCFDQLKQELTAAGLHIAPEKVQLQDPYTYLGFELNGPKITNQKAVIRK




DKLQTLNDFQKLLGDINWLRPYLKLTTGDLKPLFDTLKPDSDPNSHRSLS




KEALASLEKVETAIAEQFVTHINYSLPLIFLIFNTALTPTGLFWQDNPIM




WIHLPASPKKVLLPYYDAIADLIILGRDHSKKYFGIEPSTIIQPYSKSQI




DWLMQNTEMWPIACASFVGILDNHYPPNKLIQFCKLHTFVFPQIISKTPL




NNALLVFTDGSSTGMAAYTLTDTTIKFQTNLNSAQLVELQALIAVLSAFP




NQPLNIYTDSAYLAHSIPLLETVAQIKHISETAKLFLQCQQLIYNRSIPF




YIGHVRAHSGLPGPIAQGNQRADLATKIVASNINT





PERV_
8,099
TLQLDDEYRLYSPLVKPDQNIQFWLEQFPQAWAETAGMGLAKQVPPQVIQ


Q4VFZ2

LKASATPVSVRQYPLSKEAQEGIRPHVQRLIQQGILVPVQSPWNTPLLPV




RKPGTNDYRPVQDLREVNKRVQDIHPTVPNPYNLLCALPPQRSWYTVLDL




KDAFFCLRLHPTSQPLFAFEWRDPGTGRTGQLTWTRLPQGFKNSPTIFDE




ALHRDLANFRIQHPQVTLLQYVDDLLLAGATKQDCLEGTKALLLELSDLG




YRASAKKAQICRREVTYLGYSLRDGQRWLTEARKKTVVQIPAPTTAKQVR




EFLGTAGFCRLWIPGFATLAAPLYPLTKEKGEFSWAPEHQKAFDAIKKAL




LSAPALALPDVTKPFTLYVDERKGVARGVLTQTLGPWRRPVAYLSKKLDP




VASGWPVCLKAIAAVAILVKDADKLTLGQNITVIAPHALENIVRQPPDRW




MTNARMTHYQSLLLTERVTFAPPAALNPATLLPEETDEPVTHDCHQLLIE




ETGVRKDLTDIPLTGEVLTWFTDGSSYVVEGKRMAGAAVVDGTRTIWASS




LPEGTSAQKAELMALTQALRLAEGKSINIYTDSRYAFATAHVHGAIYKQR




GLLTSAGREIKNKEEILSLLEALHLPKRLAIIHCPGHQKAKDPISRGNQM




ADRVAKQAAQGVNLL





PERV_
8,100
TLQLDDEYRLYSPLVKPDQNIQFWLEQFPQAWAETAGMGLAKQVPPQVIQ


Q4VFZ2

LKASATPVSVRQYPLSKEAQEGIRPHVQRLIQQGILVPVQSPWNTPLLPV




RKPGTNDYRPVQDLREVNKRVQDIHPTVPNPYNLLCALPPQRSWYTVLDL




KDAFFCLRLHPTSQPLFAFEWRDPGTGRTGQLTWTRLPQGFKNSPTIFDE




ALHRDLANFRIQHPQVTLLQYVDDLLLAGATKQDCLEGTKALLLELSDLG




YRASAKKAQICRREVTYLGYSLRDGQRWLTEARKKTVQIPAPTTAKQVRE




FLGTAGFCRLWIPGFATLAAPLYPLTKEKGEFSWAPEHQKAFDAIKKALL




SAPALALPDVTKPFTLYVDERKGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPVCLKAIAAVAILVKDADKLTLGQNITVIAPHALENIVRQPPDRWM




TNARMTHYQSLLLTERVTFAPPAALNPATLLPEETDEPVTHDCHQLLIEE




TGVRKDLTDIPLTGEVLTWFTDGSSYVVEGKRMAGAAVVDGTRTIWASSL




PEGTSAQKAELMALTQALRLAEGKSINIYTDSRYAFATAHVHGAIYKQRG




LLTSAGREIKNKEEILSLLEALHLPKRLAIIHCPGHQKAKDPISRGNQMA




DRVAKQAAQGVNLL





PERV_
8,101
TLQLDDEYRLYSPLVKPDQNIQFWLEQFPQAWAETAGMGLAKQVPPQVIQ


Q4VFZ2_

LKASATPVSVRQYPLSKEAQEGIRPHVQRLIQQGILVPVQSPWNTPLLPV


3mut

RKPGTNDYRPVQDLREVNKRVQDIHPTVPNPYNLLCALPPQRSWYTVLDL




KDAFFCLRLHPTSQPLFAFEWRDPGTGRTGQLTWTRLPQGFKNSPTIFNE




ALHRDLANFRIQHPQVTLLQYVDDLLLAGATKQDCLEGTKALLLELSDLG




YRASAKKAQICRREVTYLGYSLRDGQRWLTEARKKTVQIPAPTTAKQVRE




FLGTAGFCRLWIPGFATLAAPLYPLTKPKGEFSWAPEHQKAFDAIKKALL




SAPALALPDVTKPFTLYVDERKGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPVCLKAIAAVAILVKDADKLTLGQNITVIAPHALENIVRQPPDRWM




TNARMTHYQSLLLTERVTFAPPAALNPATLLPEETDEPVTHDCHQLLIEE




TGVRKDLTDIPLTGEVLTWFTDGSSYVVEGKRMAGAAVVDGTRTIWASSL




PEGTSAQKAELMALTQALRLAEGKSINIYTDSRYAFATAHVHGAIYKQRG




WLTSAGREIKNKEEILSLLEALHLPKRLAIIHCPGHQKAKDPISRGNQMA




DRVAKQAAQGVNLL





PERV_
8,102
TLQLDDEYRLYSPLVKPDQNIQFWLEQFPQAWAETAGMGLAKQVPPQVIQ


Q4VFZ2_

LKASATPVSVRQYPLSKEAQEGIRPHVQRLIQQGILVPVQSPWNTPLLPV


3mut

RKPGTNDYRPVQDLREVNKRVQDIHPTVPNPYNLLCALPPQRSWYTVLDL




KDAFFCLRLHPTSQPLFAFEWRDPGTGRTGQLTWTRLPQGFKNSPTIFNE




ALHRDLANFRIQHPQVTLLQYVDDLLLAGATKQDCLEGTKALLLELSDLG




YRASAKKAQICRREVTYLGYSLRDGQRWLTEARKKTVVQIPAPTTAKQVR




EFLGTAGFCRLWIPGFATLAAPLYPLTKPKGEFSWAPEHQKAFDAIKKAL




LSAPALALPDVTKPFTLYVDERKGVARGVLTQTLGPWRRPVAYLSKKLDP




VASGWPVCLKAIAAVAILVKDADKLTLGQNITVIAPHALENIVRQPPDRW




MTNARMTHYQSLLLTERVTFAPPAALNPATLLPEETDEPVTHDCHQLLIE




ETGVRKDLTDIPLTGEVLTWFTDGSSYVVEGKRMAGAAVVDGTRTIWASS




LPEGTSAQKAELMALTQALRLAEGKSINIYTDSRYAFATAHVHGAIYKQR




GWLTSAGREIKNKEEILSLLEALHLPKRLAIIHCPGHQKAKDPISRGNQM




ADRVAKQAAQGVNLL





PERV_
8,103
LDDEYRLYSPLVKPDQNIQFWLEQFPQAWAETAGMGLAKQVPPQVIQLKA


Q4VFZ2_

SATPVSVRQYPLSKEAQEGIRPHVQRLIQQGILVPVQSPWNTPLLPVRKP


3mutA_

GTNDYRPVQDLREVNKRVQDIHPTVPNPYNLLCALPPQRSWYTVLDLKDA


WS

FFCLRLHPTSQPLFAFEWRDPGTGRTGQLTWTRLPQGFKNSPTIFNEALH




RDLANFRIQHPQVTLLQYVDDLLLAGATKQDCLEGTKALLLELSDLGYRA




SAKKAQICRREVTYLGYSLRDGQRWLTEARKKTVVQIPAPTTAKQVREFL




GKAGFCRLFIPGFATLAAPLYPLTKPKGEFSWAPEHQKAFDAIKKALLSA




PALALPDVTKPFTLYVDERKGVARGVLTQTLGPWRRPVAYLSKKLDPVAS




GWPVCLKAIAAVAILVKDADKLTLGQNITVIAPHALENIVRQPPDRWMTN




ARMTHYQSLLLTERVTFAPPAALNPATLLPEETDEPVTHDCHQLLIEETG




VRKDLTDIPLTGEVLTWFTDGSSYVVEGKRMAGAAVVDGTRTIWASSLPE




GTSAQKAELMALTQALRLAEGKSINIYTDSRYAFATAHVHGAIYKQRGWL




TSAGREIKNKEEILSLLEALHLPKRLAIIHCPGHQKAKDPISRGNQMADR




VAKQAAQGVNLLP





PERV_
8,104
LDDEYRLYSPLVKPDQNIQFWLEQFPQAWAETAGMGLAKQVPPQVIQLKA


Q4VFZ2_

SATPVSVRQYPLSKEAQEGIRPHVQRLIQQGILVPVQSPWNTPLLPVRKP


3mutA_

GTNDYRPVQDLREVNKRVQDIHPTVPNPYNLLCALPPQRSWYTVLDLKDA


WS

FFCLRLHPTSQPLFAFEWRDPGTGRTGQLTWTRLPQGFKNSPTIFNEALH




RDLANFRIQHPQVTLLQYVDDLLLAGATKQDCLEGTKALLLELSDLGYRA




SAKKAQICRREVTYLGYSLRDGQRWLTEARKKTVVQIPAPTTAKQVREFL




GKAGFCRLFIPGFATLAAPLYPLTKPKGEFSWAPEHQKAFDAIKKALLSA




PALALPDVTKPFTLYVDERKGVARGVLTQTLGPWRRPVAYLSKKLDPVAS




GWPVCLKAIAAVAILVKDADKLTLGQNITVIAPHALENIVRQPPDRWMTN




ARMTHYQSLLLTERVTFAPPAALNPATLLPEETDEPVTHDCHQLLIEETG




VRKDLTDIPLTGEVLTWFTDGSSYVVEGKRMAGAAVVDGTRTIWASSLPE




GTSAQKAELMALTQALRLAEGKSINIYTDSRYAFATAHVHGAIYKQRGWL




TSAGREIKNKEEILSLLEALHLPKRLAIIHCPGHQKAKDPISRGNQMADR




VAKQAAQGVNLLP





SFV1_
8,105
MDPLQLLQPLEAEIKGTKLKAHWNSGATITCVPEAFLEDERPIQTMLIKT


P23074

IHGEKQQDVYYLTFKVQGRKVEAEVLASPYDYILLNPSDVPWLMKKPLQL




TVLVPLHEYQERLLQQTALPKEQKELLQKLFLKYDALWQHWENQVGHRRI




KPHNIATGTLAPRPQKQYPINPKAKPSIQIVIDDLLKQGVLIQQNSTMNT




PVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGILSSIYRGKYKTT




LDLTNGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFLNSPALFTADWV




DLLKEIPNVQAYVDDIYISHDDPQEHLEQLEKIFSILLNAGYVVSLKKSE




IAQREVEFLGFNITKEGRGLTDTFKQKLLNITPPKDLKQLQSILGLLNFA




RNFIPNYSELVKPLYTIVANANGKFISWTEDNSNQLQHIISVLNQADNLE




ERNPETRLIIKVNSSPSAGYIRYYNEGSKRPIMYVNYIFSKAEAKFTQTE




KLLTTMHKGLIKAMDLAMGQEILVYSPIVSMTKIQRTPLPERKALPVRWI




TWMTYLEDPRIQFHYDKSLPELQQIPNVTEDVIAKTKHPSEFAMVFYTDG




SAIKHPDVNKSHSAGMGIAQVQFIPEYKIVHQWSIPLGDHTAQLAEIAAV




EFACKKALKISGPVLIVTDSFYVAESANKELPYWKSNGFLNNKKKPLRHV




SKWKSIAECLQLKPDIIIMHEKGHQQPMTTLHTEGNNLADKLATQGSYVV




H





SFV1_
8,106
MDPLQLLQPLEAEIKGTKLKAHWNSGATITCVPEAFLEDERPIQTMLIKT


P23074_

IHGEKQQDVYYLTFKVQGRKVEAEVLASPYDYILLNPSDVPWLMKKPLQL


2mut

TVLVPLHEYQERLLQQTALPKEQKELLQKLFLKYDALWQHWENQVGHRRI




KPHNIATGTLAPRPQKQYPINPKAKPSIQIVIDDLLKQGVLIQQNSTMNT




PVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGILSSIYRGKYKTT




LDLTNGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFLNSPALFNADWD




LLKEIPNVQAYVDDIYISHDDPQEHLEQLEKIFSILLNAGYVVSLKKSEI




AQREVEFLGFNITKEGRGLTDTFKQKLLNITPPKDLKQLQSILGLLNFAR




NFIPNYSELVKPLYTIVAPANGKFISWTEDNSNQLQHIISVLNQADNLEE




RNPETRLIIKVNSSPSAGYIRYYNEGSKRPIMYVNYIFSKAEAKFTQTEK




LLTTMHKGLIKAMDLAMGQEILVYSPIVSMTKIQRTPLPERKALPVRWIT




WMTYLEDPRIQFHYDKSLPELQQIPNVTEDVIAKTKHPSEFAMVFYTDGS




AIKHPDVNKSHSAGMGIAQVQFIPEYKIVHQWSIPLGDHTAQLAEIAAVE




FACKKALKISGPVLIVTDSFYVAESANKELPYWKSNGFLNNKKKPLRHVS




KWKSIAECLQLKPDIIIMHEKGHQQPMTTLHTEGNNLADKLATQGSYVVH





SFV1_
8,107
MDPLQLLQPLEAEIKGTKLKAHWNSGATITCVPEAFLEDERPIQTMLIKT


P23074_

IHGEKQQDVYYLTFKVQGRKVEAEVLASPYDYILLNPSDVPWLMKKPLQL


2mutA

TVLVPLHEYQERLLQQTALPKEQKELLQKLFLKYDALWQHWENQVGHRRI




KPHNIATGTLAPRPQKQYPINPKAKPSIQIVIDDLLKQGVLIQQNSTMNT




PVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGILSSIYRGKYKTT




LDLTNGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFLNSPALFNADWV




DLLKEIPNVQAYVDDIYISHDDPQEHLEQLEKIFSILLNAGYVVSLKKSE




IAQREVEFLGFNITKEGRGLTDTFKQKLLNITPPKDLKQLQSILGKLNFA




RNFIPNYSELVKPLYTIVAPANGKFISWTEDNSNQLQHIISVLNQADNLE




ERNPETRLIIKVNSSPSAGYIRYYNEGSKRPIMYVNYIFSKAEAKFTQTE




KLLTTMHKGLIKAMDLAMGQEILVYSPIVSMTKIQRTPLPERKALPVRWI




TWMTYLEDPRIQFHYDKSLPELQQIPNVTEDVIAKTKHPSEFAMVFYTDG




SAIKHPDVNKSHSAGMGIAQVQFIPEYKIVHQWSIPLGDHTAQLAEIAAV




EFACKKALKISGPVLIVTDSFYVAESANKELPYWKSNGFLNNKKKPLRHV




SKWKSIAECLQLKPDIIIMHEKGHQQPMTTLHTEGNNLADKLATQGSYVV




H





SFV1_
8,108
VPWLMKKPLQLTVLVPLHEYQERLLQQTALPKEQKELLQKLFLKYDALWQ


P23074-

HWENQVGHRRIKPHNIATGTLAPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro

VLIQQNSTMNTPVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGIL




SSIYRGKYKTTLDLTNGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFL




NSPALFTADVVDLLKEIPNVQAYVDDIYISHDDPQEHLEQLEKIFSILLN




AGYVVSLKKSEIAQREVEFLGFNITKEGRGLTDTFKQKLLNITPPKDLKQ




LQSILGLLNFARNFIPNYSELVKPLYTIVANANGKFISWTEDNSNQLQHI




ISVLNQADNLEERNPETRLIIKVNSSPSAGYIRYYNEGSKRPIMYVNYIF




SKAEAKFTQTEKLLTTMHKGLIKAMDLAMGQEILVYSPIVSMTKIQRTPL




PERKALPVRWITWMTYLEDPRIQFHYDKSLPELQQIPNVTEDVIAKTKHP




SEFAMVFYTDGSAIKHPDVNKSHSAGMGIAQVQFIPEYKIVHQWSIPLGD




HTAQLAEIAAVEFACKKALKISGPVLIVTDSFYVAESANKELPYWKSNGF




LNNKKKPLRHVSKWKSIAECLQLKPDIIIMHEKGHQQPMTTLHTEGNNLA




DKLATQGSYVVH





SFV1_
8,109
VPWLMKKPLQLTVLVPLHEYQERLLQQTALPKEQKELLQKLFLKYDALWQ


P23074_

HWENQVGHRRIKPHNIATGTLAPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro_

VLIQQNSTMNTPVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGIL


2mut

SSIYRGKYKTTLDLTNGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFL




NSPALFNADVVDLLKEIPNVQAYVDDIYISHDDPQEHLEQLEKIFSILLN




AGYVVSLKKSEIAQREVEFLGFNITKEGRGLTDTFKQKLLNITPPKDLKQ




LQSILGLLNFARNFIPNYSELVKPLYTIVAPANGKFISWTEDNSNQLQHI




ISVLNQADNLEERNPETRLIIKVNSSPSAGYIRYYNEGSKRPIMYVNYIF




SKAEAKFTQTEKLLTTMHKGLIKAMDLAMGQEILVYSPIVSMTKIQRTPL




PERKALPVRWITWMTYLEDPRIQFHYDKSLPELQQIPNVTEDVIAKTKHP




SEFAMVFYTDGSAIKHPDVNKSHSAGMGIAQVQFIPEYKIVHQWSIPLGD




HTAQLAEIAAVEFACKKALKISGPVLIVTDSFYVAESANKELPYWKSNGF




LNNKKKPLRHVSKWKSIAECLQLKPDIIIMHEKGHQQPMTTLHTEGNNLA




DKLATQGSYVVH





SFV1_
8,110
VPWLMKKPLQLTVLVPLHEYQERLLQQTALPKEQKELLQKLFLKYDALWQ


P23074_

HWENQVGHRRIKPHNIATGTLAPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro_

VLIQQNSTMNTPVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGIL


2mutA

SSIYRGKYKTTLDLTNGFWAHPITPESYWLTAFTWQGKQYCWTRLPQGFL




NSPALFNADVVDLLKEIPNVQAYVDDIYISHDDPQEHLEQLEKIFSILLN




AGYVVSLKKSEIAQREVEFLGFNITKEGRGLTDTFKQKLLNITPPKDLKQ




LQSILGKLNFARNFIPNYSELVKPLYTIVAPANGKFISWTEDNSNQLQHI




ISVLNQADNLEERNPETRLIIKVNSSPSAGYIRYYNEGSKRPIMYVNYIF




SKAEAKFTQTEKLLTTMHKGLIKAMDLAMGQEILVYSPIVSMTKIQRTPL




PERKALPVRWITWMTYLEDPRIQFHYDKSLPELQQIPNVTEDVIAKTKHP




SEFAMVFYTDGSAIKHPDVNKSHSAGMGIAQVQFIPEYKIVHQWSIPLGD




HTAQLAEIAAVEFACKKALKISGPVLIVTDSFYVAESANKELPYWKSNGF




LNNKKKPLRHVSKWKSIAECLQLKPDIIIMHEKGHQQPMTTLHTEGNNLA




DKLATQGSYVVH





SFV3L_
8,111
MDPLQLLQPLEAEIKGTKLKAHWNSGATITCVPQAFLEEEVPIKNIWIKT


P27401

IHGEKEQPVYYLTFKIQGRKVEAEVISSPYDYILVSPSDIPWLMKKPLQL




TTLVPLQEYEERLLKQTMLTGSYKEKLQSLFLKYDALWQHWENQVGHRRI




KPHHIATGTVNPRPQKQYPINPKAKASIQTVINDLLKQGVLIQQNSIMNT




PVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGILSSIFRGKYKTT




LDLSNGFWAHSITPESYWLTAFTWLGQQYCWTRLPQGFLNSPALFTADVV




DLLKEVPNVQVYVDDIYISHDDPREHLEQLEKVFSLLLNAGYVVSLKKSE




IAQHEVEFLGFNITKEGRGLTETFKQKLLNITPPRDLKQLQSILGLLNFA




RNFIPNFSELVKPLYNIIATANGKYITWTTDNSQQLQNIISMLNSAENLE




ERNPEVRLIMKVNTSPSAGYIRFYNEFAKRPIMYLNYVYTKAEVKFTNTE




KLLTTIHKGLIKALDLGMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMSYLEDPRIQFHYDKTLPELQQVPTVTDDIIAKIKHPSEFSMVFYTDG




SAIKHPNVNKSHNAGMGIAQVQFKPEFTVINTWSIPLGDHTAQLAEVAAV




EFACKKALKIDGPVLIVTDSFYVAESVNKELPYWQSNGFFNNKKKPLKHV




SKWKSIADCIQLKPDIIIIHEKGHQPTASTFHTEGNNLADKLATQGSYVV




N





SFV3L_
8,112
MDPLQLLQPLEAEIKGTKLKAHWNSGATITCVPQAFLEEEVPIKNIWIKT


P27401_

IHGEKEQPVYYLTFKIQGRKVEAEVISSPYDYILVSPSDIPWLMKKPLQL


2mut

TTLVPLQEYEERLLKQTMLTGSYKEKLQSLFLKYDALWQHWENQVGHRRI




KPHHIATGTVNPRPQKQYPINPKAKASIQTVINDLLKQGVLIQQNSIMNT




PVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGILSSIFRGKYKTT




LDLSNGFWAHSITPESYWLTAFTWLGQQYCWTRLPQGFLNSPALFNADVV




DLLKEVPNVQVYVDDIYISHDDPREHLEQLEKVFSLLLNAGYVVSLKKSE




IAQHEVEFLGFNITKEGRGLTETFKQKLLNITPPRDLKQLQSILGLLNFA




RNFIPNFSELVKPLYNIIATAPGKYITWTTDNSQQLQNIISMLNSAENLE




ERNPEVRLIMKVNTSPSAGYIRFYNEFAKRPIMYLNYVYTKAEVKFTNTE




KLLTTIHKGLIKALDLGMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMSYLEDPRIQFHYDKTLPELQQVPTVTDDIIAKIKHPSEFSMVFYTDG




SAIKHPNVNKSHNAGMGIAQVQFKPEFTVINTWSIPLGDHTAQLAEVAAV




EFACKKALKIDGPVLIVTDSFYVAESVNKELPYWQSNGFFNNKKKPLKHV




SKWKSIADCIQLKPDIIIIHEKGHQPTASTFHTEGNNLADKLATQGSYVV




N





SFV3L_
8,113
MDPLQLLQPLEAEIKGTKLKAHWNSGATITCVPQAFLEEEVPIKNIWIKT


P27401_

IHGEKEQPVYYLTFKIQGRKVEAEVISSPYDYILVSPSDIPWLMKKPLQL


2mutA

TTLVPLQEYEERLLKQTMLTGSYKEKLQSLFLKYDALWQHWENQVGHRRI




KPHHIATGTVNPRPQKQYPINPKAKASIQTVINDLLKQGVLIQQNSIMNT




PVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGILSSIFRGKYKTT




LDLSNGFWAHSITPESYWLTAFTWLGQQYCWTRLPQGFLNSPALFNADVV




DLLKEVPNVQVYVDDIYISHDDPREHLEQLEKVFSLLLNAGYVVSLKKSE




IAQHEVEFLGFNITKEGRGLTETFKQKLLNITPPRDLKQLQSILGKLNFA




RNFIPNFSELVKPLYNIIATAPGKYITWTTDNSQQLQNIISMLNSAENLE




ERNPEVRLIMKVNTSPSAGYIRFYNEFAKRPIMYLNYVYTKAEVKFTNTE




KLLTTIHKGLIKALDLGMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMSYLEDPRIQFHYDKTLPELQQVPTVTDDIIAKIKHPSEFSMVFYTDG




SAIKHPNVNKSHNAGMGIAQVQFKPEFTVINTWSIPLGDHTAQLAEVAAV




EFACKKALKIDGPVLIVTDSFYVAESVNKELPYWQSNGFFNNKKKPLKHV




SKWKSIADCIQLKPDIIIIHEKGHQPTASTFHTEGNNLADKLATQGSYVV




N





SFV3L_
8,114
IPWLMKKPLQLTTLVPLQEYEERLLKQTMLTGSYKEKLQSLFLKYDALWQ


P27401-

HWENQVGHRRIKPHHIATGTVNPRPQKQYPINPKAKASIQTVINDLLKQG


Pro

VLIQQNSIMNTPVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGIL




SSIFRGKYKTTLDLSNGFWAHSITPESYWLTAFTWLGQQYCWTRLPQGFL




NSPALFTADVVDLLKEVPNVQVYVDDIYISHDDPREHLEQLEKVFSLLLN




AGYVVSLKKSEIAQHEVEFLGFNITKEGRGLTETFKQKLLNITPPRDLKQ




LQSILGLLNFARNFIPNFSELVKPLYNIIATANGKYITWTTDNSQQLQNI




ISMLNSAENLEERNPEVRLIMKVNTSPSAGYIRFYNEFAKRPIMYLNYVY




TKAEVKFTNTEKLLTTIHKGLIKALDLGMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMSYLEDPRIQFHYDKTLPELQQVPTVTDDIIAKIKHP




SEFSMVFYTDGSAIKHPNVNKSHNAGMGIAQVQFKPEFTVINTWSIPLGD




HTAQLAEVAAVEFACKKALKIDGPVLIVTDSFYVAESVNKELPYWQSNGF




FNNKKKPLKHVSKWKSIADCIQLKPDIIIIHEKGHQPTASTFHTEGNNLA




DKLATQGSYVVN





SFV3L_
8,115
IPWLMKKPLQLTTLVPLQEYEERLLKQTMLTGSYKEKLQSLFLKYDALWQ


P27401-

HWENQVGHRRIKPHHIATGTVNPRPQKQYPINPKAKASIQTVINDLLKQG


Pro_

VLIQQNSIMNTPVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGIL


2mut

SSIFRGKYKTTLDLSNGFWAHSITPESYWLTAFTWLGQQYCWTRLPQGFL




NSPALFNADVVDLLKEVPNVQVYVDDIYISHDDPREHLEQLEKVFSLLLN




AGYVVSLKKSEIAQHEVEFLGFNITKEGRGLTETFKQKLLNITPPRDLKQ




LQSILGLLNFARNFIPNFSELVKPLYNIIATAPGKYITWTTDNSQQLQNI




ISMLNSAENLEERNPEVRLIMKVNTSPSAGYIRFYNEFAKRPIMYLNYVY




TKAEVKFTNTEKLLTTIHKGLIKALDLGMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMSYLEDPRIQFHYDKTLPELQQVPTVTDDIIAKIKHP




SEFSMVFYTDGSAIKHPNVNKSHNAGMGIAQVQFKPEFTVINTWSIPLGD




HTAQLAEVAAVEFACKKALKIDGPVLIVTDSFYVAESVNKELPYWQSNGF




FNNKKKPLKHVSKWKSIADCIQLKPDIIIIHEKGHQPTASTFHTEGNNLA




DKLATQGSYVVN





SFV3L_
8,116
IPWLMKKPLQLTTLVPLQEYEERLLKQTMLTGSYKEKLQSLFLKYDALWQ


P27401-

HWENQVGHRRIKPHHIATGTVNPRPQKQYPINPKAKASIQTVINDLLKQG


Pro_

VLIQQNSIMNTPVYPVPKPDGKWRMVLDYREVNKTIPLIAAQNQHSAGIL


2mutA

SSIFRGKYKTTLDLSNGFWAHSITPESYWLTAFTWLGQQYCWTRLPQGFL




NSPALFNADVVDLLKEVPNVQVYVDDIYISHDDPREHLEQLEKVFSLLLN




AGYVVSLKKSEIAQHEVEFLGFNITKEGRGLTETFKQKLLNITPPRDLKQ




LQSILGKLNFARNFIPNFSELVKPLYNIIATAPGKYITWTTDNSQQLQNI




ISMLNSAENLEERNPEVRLIMKVNTSPSAGYIRFYNEFAKRPIMYLNYVY




TKAEVKFTNTEKLLTTIHKGLIKALDLGMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMSYLEDPRIQFHYDKTLPELQQVPTVTDDIIAKIKHP




SEFSMVFYTDGSAIKHPNVNKSHNAGMGIAQVQFKPEFTVINTWSIPLGD




HTAQLAEVAAVEFACKKALKIDGPVLIVTDSFYVAESVNKELPYWQSNGF




FNNKKKPLKHVSKWKSIADCIQLKPDIIIIHEKGHQPTASTFHTEGNNLA




DKLATQGSYVVN





SFVCP_
8,117
MNPLQLLQPLPAEVKGTKLLAHWNSGATITCIPESFLEDEQPIKQTLIKT


Q87040

IHGEKQQNVYYLTFKVKGRKVEAEVIASPYEYILLSPTDVPWLTQQPLQL




TILVPLQEYQDRINKTALPEEQKQQLKALFTKYDNLWQHWENQVGHRKIR




PHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGVLTPQNSTMNTP




VYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILATIVRQKYKTTL




DLANGFWAHPITPDSYWLTAFTWQGKQYCWTRLPQGFLNSPALFTADAVD




LLKEVPNVQVYVDDIYLSHDNPHEHIQQLEKVFQILLQAGYVVSLKKSEI




GQRTVEFLGFNITKEGRGLTDTFKTKLLNVTPPKDLKQLQSILGLLNFAR




NFIPNFAELVQTLYNLIASSKGKYIEWTEDNTKQLNKVIEALNTASNLEE




RLPDQRLVIKVNTSPSAGYVRYYNESGKKPIMYLNYVFSKAELKFSMLEK




LLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLPERKALPIRWIT




WMTYLEDPRIQFHYDKTLPELKHIPDVYTSSIPPLKHPSQYEGVFCTDGS




AIKSPDPTKSNNAGMGIVHAIYNPEYKILNQWSIPLGHHTAQMAEIAAVE




FACKKALKVPGPVLVITDSFYVAESANKELPYWKSNGFVNNKKEPLKHIS




KWKSIAECLSIKPDITIQHEKGHQPINTSIHTEGNALADKLATQGSYVVN





SFVCP_
8,118
MNPLQLLQPLPAEVKGTKLLAHWNSGATITCIPESFLEDEQPIKQTLIKT


Q87040_

IHGEKQQNVYYLTFKVKGRKVEAEVIASPYEYILLSPTDVPWLTQQPLQL


2mut

TILVPLQEYQDRILNKTALPEEQKQQLKALFTKYDNLWQHWENQVGHRKI




RPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGVLTPQNSTMNT




PVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILATIVRQKYKTT




LDLANGFWAHPITPDSYWLTAFTWQGKQYCWTRLPQGFLNSPALFNADAV




DLLKEVPNVQVYVDDIYLSHDNPHEHIQQLEKVFQILLQAGYVVSLKKSE




IGQRTVEFLGFNITKEGRGLTDTFKTKLLNVTPPKDLKQLQSILGLLNFA




RNFIPNFAELVQTLYNLIASSPGKYIEWTEDNTKQLNKVIEALNTASNLE




ERLPDQRLVIKVNTSPSAGYVRYYNESGKKPIMYLNYVFSKAELKFSMLE




KLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLPERKALPIRWI




TWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSIPPLKHPSQYEGVFCTDG




SAIKSPDPTKSNNAGMGIVHAIYNPEYKILNQWSIPLGHHTAQMAEIAAV




EFACKKALKVPGPVLVITDSFYVAESANKELPYWKSNGFVNNKKEPLKHI




SKWKSIAECLSIKPDITIQHEKGHQPINTSIHTEGNALADKLATQGSYVV




N





SFVCP_
8,119
MNPLQLLQPLPAEVKGTKLLAHWNSGATITCIPESFLEDEQPIKQTLIKT


Q87040_

IHGEKQQNVYYLTFKVKGRKVEAEVIASPYEYILLSPTDVPWLTQQPLQL


2mutA

TILVPLQEYQDRINKTALPEEQKQQLKALFTKYDNLWQHWENQVGHRKIR




PHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGVLTPQNSTMNTP




VYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILATIVRQKYKTTL




DLANGFWAHPITPDSYWLTAFTWQGKQYCWTRLPQGFLNSPALFNADAVD




LLKEVPNVQVYVDDIYLSHDNPHEHIQQLEKVFQILLQAGYVVSLKKSEI




GQRTVEFLGFNITKEGRGLTDTFKTKLLNVTPPKDLKQLQSILGKLNFAR




NFIPNFAELVQTLYNLIASSPGKYIEWTEDNTKQLNKVIEALNTASNLEE




RLPDQRLVIKVNTSPSAGYVRYYNESGKKPIMYLNYVFSKAELKFSMLEK




LLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLPERKALPIRWIT




WMTYLEDPRIQFHYDKTLPELKHIPDVYTSSIPPLKHPSQYEGVFCTDGS




AIKSPDPTKSNNAGMGIVHAIYNPEYKILNQWSIPLGHHTAQMAEIAAVE




FACKKALKVPGPVLVITDSFYVAESANKELPYWKSNGFVNNKKEPLKHIS




KWKSIAECLSIKPDITIQHEKGHQPINTSIHTEGNALADKLATQGSYVVN





SFVCP_
8,120
VPWLTQQPLQLTILVPLQEYQDRILNKTALPEEQKQQLKALFTKYDNLWQ


Q87040-

HWENQVGHRKIRPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro

VLTPQNSTMNTPVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGIL




ATIVRQKYKTTLDLANGFWAHPITPDSYWLTAFTWQGKQYCWTRLPQGFL




NSPALFTADAVDLLKEVPNVQVYVDDIYLSHDNPHEHIQQLEKVFQILLQ




AGYVVSLKKSEIGQRTVEFLGFNITKEGRGLTDTFKTKLLNVTPPKDLKQ




LQSILGLLNFARNFIPNFAELVQTLYNLIASSKGKYIEWTEDNTKQLNKV




IEALNTASNLEERLPDQRLVIKVNTSPSAGYVRYYNESGKKPIMYLNYVF




SKAELKFSMLEKLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSIPPLKHP




SQYEGVFCTDGSAIKSPDPTKSNNAGMGIVHAIYNPEYKILNQWSIPLGH




HTAQMAEIAAVEFACKKALKVPGPVLVITDSFYVAESANKELPYWKSNGF




VNNKKEPLKHISKWKSIAECLSIKPDITIQHEKGHQPINTSIHTEGNALA




DKLATQGSYVVN





SFVCP_
8,121
VPWLTQQPLQLTILVPLQEYQDRILNKTALPEEQKQQLKALFTKYDNLWQ


Q87040-

HWENQVGHRKIRPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQG


Pro_

VLTPQNSTMNTPVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGIL


2mut

ATIVRQKYKTTLDLANGFWAHPITPDSYWLTAFTWQGKQYCWTRLPQGFL




NSPALFNADAVDLLKEVPNVQVYVDDIYLSHDNPHEHIQQLEKVFQILLQ




AGYVVSLKKSEIGQRTVEFLGFNITKEGRGLTDTFKTKLLNVTPPKDLKQ




LQSILGLLNFARNFIPNFAELVQTLYNLIASSPGKYIEWTEDNTKQLNKV




IEALNTASNLEERLPDQRLVIKVNTSPSAGYVRYYNESGKKPIMYLNYVF




SKAELKFSMLEKLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPL




PERKALPIRWITWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSIPPLKHP




SQYEGVFCTDGSAIKSPDPTKSNNAGMGIVHAIYNPEYKILNQWSIPLGH




HTAQMAEIAAVEFACKKALKVPGPVLVITDSFYVAESANKELPYWKSNGF




VNNKKEPLKHISKWKSIAECLSIKPDITIQHEKGHQPINTSIHTEGNALA




DKLATQGSYVVN





SFVCP_
8,122
VPWLTQQPLQLTILVPLQEYQDRINKTALPEEQKQQLKALFTKYDNLWQH


Q87040-

WENQVGHRKIRPHNIATGDYPPRPQKQYPINPKAKPSIQIVIDDLLKQGV


Pro_

LTPQNSTMNTPVYPVPKPDGRWRMVLDYREVNKTIPLTAAQNQHSAGILA


2mutA

TIVRQKYKTTLDLANGFWAHPITPDSYWLTAFTWQGKQYCWTRLPQGFLN




SPALFNADAVDLLKEVPNVQVYVDDIYLSHDNPHEHIQQLEKVFQILLQA




GYVVSLKKSEIGQRTVEFLGFNITKEGRGLTDTFKTKLLNVTPPKDLKQL




QSILGKLNFARNFIPNFAELVQTLYNLIASSPGKYIEWTEDNTKQLNKVI




EALNTASNLEERLPDQRLVIKVNTSPSAGYVRYYNESGKKPIMYLNYVFS




KAELKFSMLEKLLTTMHKALIKAMDLAMGQEILVYSPIVSMTKIQKTPLP




ERKALPIRWITWMTYLEDPRIQFHYDKTLPELKHIPDVYTSSIPPLKHPS




QYEGVFCTDGSAIKSPDPTKSNNAGMGIVHAIYNPEYKILNQWSIPLGHH




TAQMAEIAAVEFACKKALKVPGPVLVITDSFYVAESANKELPYWKSNGFV




NNKKEPLKHISKWKSIAECLSIKPDITIQHEKGHQPINTSIHTEGNALAD




KLATQGSYVVN





SMRVH_
8,123
PRSRAIDIPVPHADKISWKITDPVWVDQWPLTYEKTLAAIALVQEQLAAG


P03364

HIEPTNSPWNTPIFIIKKKSGSWRLLQDLRAVNKVMVPMGALQPGLPSPV




AIPLNYHKIVIDLKDCFFTIPLHPEDRPYFAFSVPQINFQSPMPRYQWKV




LPQGMANSPTLCQKFVAAAIAPVRSQWPEAYILHYMDDILLACDSAEAAK




ACYAHIISCLTSYGLKIAPDKVQVSEPFSYLGFELHHQQVFTPRVCLKTD




HLKTLNDFQKLLGDIQWLRPYLKLPTSALVPLNNILKGDPNPLSVRALTP




EAKQSLALINKAIQNQSVQQISYNLPLVLLLLPTPHTPTAVFWQPNGTDP




TKNGSPLLWLHLPASPSKVLLTYPSLLAMLIIKGRYTGRQLFGRDPHSII




IPYTQDQLTWLLQTSDEWAIALSSFTGDIDNHYPSDPVIQFAKLHQFIFP




KITKCAPIPQATLVFTDGSSNGIAAYVIDNQPISIKSPYLSAQLVELYAI




LQVFTVLAHQPFNLYTDSAYIAQSVPLLETVPFIKSSTNATPLFSKLQQL




ILNRQHPFFIGHLRAHLNLPGPLAEGNALADAATQIFPIISD





SMRVH_
8,124
PRSRAIDIPVPHADKISWKITDPVWVDQWPLTYEKTLAAIALVQEQLAAG


P03364_

HIEPTNSPWNTPIFIIKKKSGSWRLLQDLRAVNKVMVPMGALQPGLPSPV


2mut

AIPLNYHKIVIDLKDCFFTIPLHPEDRPYFAFSVPQINFQSPMPRYQWKV




LPQGMANSPTLCQKFVAAAIAPVRSQWPEAYILHYMDDILLACDSAEAAK




ACYAHIISCLTSYGLKIAPDKVQVSEPFSYLGFELHHQQVFTPRVCLKTD




HLKTLNDFQKLLGDIQWLRPYLKLPTSALVPLNNILKPDPNPLSVRALTP




EAKQSLALINKAIQNQSVQQISYNLPLVLLLLPTPHTPTAVFWQPNGTDP




TKNGSPLLWLHLPASPSKVLLTYPSLLAMLIIKGRYTGRQLFGRDPHSII




IPYTQDQLTWLLQTSDEWAIALSSFTGDIDNHYPSDPVIQFAKLHQFIFP




KITKCAPIPQATLVFTDGSSNGIAAYVIDNQPISIKSPYLSAQLVELYAI




LQVFTVLAHQPFNLYTDSAYIAQSVPLLETVPFIKSSTNATPLFSKLQQL




ILNRQHPFFIGHLRAHLNLPGPLAEGNALADAATQIFPIISD





SMRVH_
8,125
PRSRAIDIPVPHADKISWKITDPVWVDQWPLTYEKTLAAIALVQEQLAAG


P03364_

HIEPTNSPWNTPIFIIKKKSGSWRLLQDLRAVNKVMVPMGALQPGLPSPV


2mutB

APPLNYHKIVIDLKDCFFTIPLHPEDRPYFAFSVPQINFQSPMPRYQWKV




LPQGMANSPTLCQKFVAAAIAPVRSQWPEAYILHYMDDILLACDSAEAAK




ACYAHIISCLTSYGLKIAPDKVQVSEPFSYLGFELHHQQVFTPRVCLKTD




HLKTLNDFQKLLGDIQWLRPYLKLPTSALVPLNNILKPDPNPLSVRALTP




EAKQSLALINKAIQNQSVQQISYNLPLVLLLLPTPHTPTAVFWQPNGTDP




TKNGSPLLWLHLPASPSKVLLTYPSLLAMLIIKGRYTGRQLFGRDPHSII




IPYTQDQLTWLLQTSDEWAIALSSFTGDIDNHYPSDPVIQFAKLHQFIFP




KITKCAPIPQATLVFTDGSSNGIAAYVIDNQPISIKSPYLSAQLVELYAI




LQVFTVLAHQPFNLYTDSAYIAQSVPLLETVPFIKSSTNATPLFSKLQQL




ILNRQHPFFIGHLRAHLNLPGPLAEGNALADAATQIFPIISD





SRV2_
8,126
LATAVDILAPQRYADPITWKSDEPVWVDQWPLTQEKLAAAQQLVQEQLQA


P51517

GHIIESNSPWNTPIFVIKKKSGKWRLLQDLRAVNATMVLMGALQPGLPSP




VAIPQGYFKIVIDLKDCFFTIPLQPVDQKRFAFSLPSTNFKQPMKRYQWK




VLPQGMANSPTLCQKYVAAAIEPVRKSWAQMYIIHYMDDILIAGKLGEQV




LQCFAQLKQALTTTGLQIAPEKVQLQDPYTYLGFQINGPKITNQKAVIRR




DKLQTLNDFQKLLGDINWLRPYLHLTTGDLKPLFDILKGDSNPNSPRSLS




EAALASLQKVETAIAEQFVTQIDYTQPLTFLIFNTTLTPTGLFWQNNPVM




WVHLPASPKKVLLPYYDAIADLIILGRDNSKKYFGLEPSTIIQPYSKSQI




HWLMQNTETWPIACASYAGNIDNHYPPNKLIQFCKLHAVVFPRIISKTPL




DNALLVFTDGSSTGIAAYTFEKTTVRFKTSHTSAQLVELQALIAVLSAFP




HRALNVYTDSAYLAHSIPLLETVSHIKHISDTAKFFLQCQQLIYNRSIPF




YLGHIRAHSGLPGPLSQGNHITDLATKVVATTLTT





SRV2_
8,127
LATAVDILAPQRYADPITWKSDEPVWVDQWPLTQEKLAAAQQLVQEQLQA


P51517_

GHIIESNSPWNTPIFVIKKKSGKWRLLQDLRAVNATMVLMGALQPGLPSP


2mutB

VAPPQGYFKIVIDLKDCFFTIPLQPVDQKRFAFSLPSTNFKQPMKRYQWK




VLPQGMANSPTLCQKYVAAAIEPVRKSWAQMYIIHYMDDILIAGKLGEQV




LQCFAQLKQALTTTGLQIAPEKVQLQDPYTYLGFQINGPKITNQKAVIRR




DKLQTLNDFQKLLGDINWLRPYLHLTTGDLKPLFDILKGDSNPNSPRSLS




EAALASLQKVETAIAEQFVTQIDYTQPLTFLIFNTTLTPTGLFWQNNPVM




WVHLPASPKKVLLPYYDAIADLIILGRDNSKKYFGLEPSTIIQPYSKSQI




HWLMQNTETWPIACASYAGNIDNHYPPNKLIQFCKLHAVVFPRIISKTPL




DNALLVFTDGSSTGIAAYTFEKTTVRFKTSHTSAQLVELQALIAVLSAFP




HRALNVYTDSAYLAHSIPLLETVSHIKHISDTAKFFLQCQQLIYNRSIPF




YLGHIRAHSGLPGPLSQGNHITDLATKVVATTLTT





WDSV_
8,128
SCQTKNTLNIDEYLLQFPDQLWASLPTDIGRMLVPPITIKIKDNASLPSI


O92815

RQYPLPKDKTEGLRPLISSLENQGILIKCHSPCNTPIFPIKKAGRDEYRM




IHDLRAINNIVAPLTAVVASPTTVLSNLAPSLHWFTVIDLSNAFFSVPIH




KDSQYLFAFTFEGHQYTWTVLPQGFIHSPTLFSQALYQSLHKIKFKISSE




ICIYMDDVLIASKDRDTNLKDTAVMLQHLASEGHKVSKKKLQLCQQEVVY




LGQLLTPEGRKILPDRKVTVSQFQQPTTIRQIRAFLGLVGYCRHWIPEFS




IHSKFLEKQLKKDTAEPFQLDDQQVEAFNKLKHAITTAPVLVVPDPAKPF




QLYTSHSEHASIAVLTQKHAGRTRPIAFLSSKFDAIESGLPPCLKACASI




HRSLTQADSFILGAPLIIYTTHAICTLLQRDRSQLVTASRFSKWEADLLR




PELTFVACSAVSPAHLYMQSCENNIPPHDCVLLTHTISRPRPDLSDLPIP




DPDMTLFSDGSYTTGRGGAAVVMHRPVTDDFIIIHQQPGGASAQTAELLA




LAAACHLATDKTVNIYTDSRYAYGWHDFGHLWMHRGFVTSAGTPIKNHKE




IEYLLKQIMKPKQVSVIKIEAHTKGVSMEVRGNAAADEAAKNAVFLVQR





WDSV_
8,129
SCQTKNTLNIDEYLLQFPDQLWASLPTDIGRMLVPPITIKIKDNASLPSI


O92815_

RQYPLPKDKTEGLRPLISSLENQGILIKCHSPCNTPIFPIKKAGRDEYRM


2mut

IHDLRAINNIVAPLTAVVASPTTVLSNLAPSLHWFTVIDLSNAFFSVPIH




KDSQYLFAFTFEGHQYTWTVLPQGFIHSPTLFNQALYQSLHKIKFKISSE




ICIYMDDVLIASKDRDTNLKDTAVMLQHLASEGHKVSKKKLQLCQQEVVY




LGQLLTPEGRKILPDRKVTVSQFQQPTTIRQIRAFLGLVGYCRHWIPEFS




IHSKFLEKQLKPDTAEPFQLDDQQVEAFNKLKHAITTAPVLWPDPAKPFQ




LYTSHSEHASIAVLTQKHAGRTRPIAFLSSKFDAIESGLPPCLKACASIH




RSLTQADSFILGAPLIIYTTHAICTLLQRDRSQLVTASRFSKWEADLLRP




ELTFVACSAVSPAHLYMQSCENNIPPHDCVLLTHTISRPRPDLSDLPIPD




PDMTLFSDGSYTTGRGGAAVVMHRPVTDDFIIIHQQPGGASAQTAELLAL




AAACHLATDKTVNIYTDSRYAYGVVHDFGHLWMHRGFVTSAGTPIKNHKE




IEYLLKQIMKPKQVSVIKIEAHTKGVSMEVRGNAAADEAAKNAVFLVQR





WDSV_
8,130
SCQTKNTLNIDEYLLQFPDQLWASLPTDIGRMLVPPITIKIKDNASLPSI


O92815_

RQYPLPKDKTEGLRPLISSLENQGILIKCHSPCNTPIFPIKKAGRDEYRM


2mutA

IHDLRAINNIVAPLTAVVASPTTVLSNLAPSLHWFTVIDLSNAFFSVPIH




KDSQYLFAFTFEGHQYTWTVLPQGFIHSPTLFNQALYQSLHKIKFKISSE




ICIYMDDVLIASKDRDTNLKDTAVMLQHLASEGHKVSKKKLQLCQQEVVY




LGQLLTPEGRKILPDRKVTVSQFQQPTTIRQIRAFLGKVGYCRHFIPEFS




IHSKFLEKQLKPDTAEPFQLDDQQVEAFNKLKHAITTAPVLVVPDPAKPF




QLYTSHSEHASIAVLTQKHAGRTRPIAFLSSKFDAIESGLPPCLKACASI




HRSLTQADSFILGAPLIIYTTHAICTLLQRDRSQLVTASRFSKWEADLLR




PELTFVACSAVSPAHLYMQSCENNIPPHDCVLLTHTISRPRPDLSDLPIP




DPDMTLFSDGSYTTGRGGAAVVMHRPVTDDFIIIHQQPGGASAQTAELLA




LAAACHLATDKTVNIYTDSRYAYGVHDFGHLWMHRGFVTSAGTPIKNHKE




IEYLLKQIMKPKQVSVIKIEAHTKGVSMEVRGNAAADEAAKNAVFLVQR





WMSV_
8,131
VLNLEEEYRLHEKPVPSSIDPSWLQLFPTVWAERAGMGLANQVPPVVVEL


P03359

RSGASPVAVRQYPMSKEAREGIRPHIQRFLDLGVLVPCQSPWNTPLLPVK




KPGTNDYRPVQDLREINKRVQDIHPTVPNPYNLLSSLPPSHTWYSVLDLK




DAFFCLKLHPNSQPLFAFEWRDPEKGNTGQLTWTRLPQGFKNSPTLFDEA




LHRDLAPFRALNPQVVLLQYVDDLLVAAPTYRDCKEGTQKLLQELSKLGY




RVSAKKAQLCQKEVTYLGYLLKEGKRWLTPARKATVMKIPPPTTPRQVRE




FLGTAGFCRLWIPGFASLAAPLYPLTKESIPFIWTEEHQKAFDRIKEALL




SAPALALPDLTKPFTLYVDERAGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPTCLKAVAAVALLLKDADKLTLGQNVTVIASHSLESIVRQPPDRWM




TNARMTHYQSLLLNERVSFAPPAVLNPATLLPVESEATPVHRCSEILAEE




TGTRRDLKDQPLPGVPAWYTDGSSFIAEGKRRAGAAIVDGKRTVWASSLP




EGTSAQKAELVALTQALRLAEGKDINIYTDSRYAFATAHIHGAIYKQRGL




LTSAGKDIKNKEEILALLEAIHLPKRVAIIHCPGHQKGNDPVATGNRRAD




EAAKQAALSTRVLAETTKP





WMSV_
8,132
VLNLEEEYRLHEKPVPSSIDPSWLQLFPTVWAERAGMGLANQVPPVVVEL


P03359_

RSGASPVAVRQYPMSKEAREGIRPHIQRFLDLGVLVPCQSPWNTPLLPVK


3mut

KPGTNDYRPVQDLREINKRVQDIHPTVPNPYNLLSSLPPSHTWYSVLDLK




DAFFCLKLHPNSQPLFAFEWRDPEKGNTGQLTWTRLPQGFKNSPTLFNEA




LHRDLAPFRALNPQWVLLQYVDDLLVAAPTYRDCKEGTQKLLQELSKLGY




RVSAKKAQLCQKEVTYLGYLLKEGKRWLTPARKATVMKIPPPTTPRQVRE




FLGTAGFCRLWIPGFASLAAPLYPLTKPSIPFIWTEEHQKAFDRIKEALL




SAPALALPDLTKPFTLYVDERAGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPTCLKAVAAVALLLKDADKLTLGQNVTVIASHSLESIVRQPPDRWM




TNARMTHYQSLLLNERVSFAPPAVLNPATLLPVESEATPVHRCSEILAEE




TGTRRDLKDQPLPGVPAWYTDGSSFIAEGKRRAGAAIVDGKRTVWASSLP




EGTSAQKAELVALTQALRLAEGKDINIYTDSRYAFATAHIHGAIYKQRGW




LTSAGKDIKNKEEILALLEAIHLPKRVAIIHCPGHQKGNDPVATGNRRAD




EAAKQAALSTRVLAETTKP





WMSV_
8,133
VLNLEEEYRLHEKPVPSSIDPSWLQLFPTVWAERAGMGLANQVPPVVVEL


P03359_

RSGASPVAVRQYPMSKEAREGIRPHIQRFLDLGVLVPCQSPWNTPLLPVK


3mutA

KPGTNDYRPVQDLREINKRVQDIHPTVPNPYNLLSSLPPSHTWYSVLDLK




DAFFCLKLHPNSQPLFAFEWRDPEKGNTGQLTWTRLPQGFKNSPTLFNEA




LHRDLAPFRALNPQVVLLQYVDDLLVAAPTYRDCKEGTQKLLQELSKLGY




RVSAKKAQLCQKEVTYLGYLLKEGKRWLTPARKATVMKIPPPTTPRQVRE




FLGKAGFCRLFIPGFASLAAPLYPLTKPSIPFIWTEEHQKAFDRIKEALL




SAPALALPDLTKPFTLYVDERAGVARGVLTQTLGPWRRPVAYLSKKLDPV




ASGWPTCLKAVAAVALLLKDADKLTLGQNVTVIASHSLESIVRQPPDRWM




TNARMTHYQSLLLNERVSFAPPAVLNPATLLPVESEATPVHRCSEILAEE




TGTRRDLKDQPLPGVPAWYTDGSSFIAEGKRRAGAAIVDGKRTVWASSLP




EGTSAQKAELVALTQALRLAEGKDINIYTDSRYAFATAHIHGAIYKQRGW




LTSAGKDIKNKEEILALLEAIHLPKRVAIIHCPGHQKGNDPVATGNRRAD




EAAKQAALSTRVLAETTKP





XMRV6_
8,134
TLNIEDEYRLHETSKEPDVPLGSTWLSDFPQAWAETGGMGLAVRQAPLII


A1Z651

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP




VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD




EALHRDLADFRIQHPDLILLQYVDDLLLAATSEQDCQRGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKTGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEKEAPHDCLEILA




ETHGTRPDLTDQPIPDADYTWYTDGSSFLQEGQRRAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHVHGEIYRR




RGLLTSEGREIKNKNEILALLKALFLPKRLSIIHCPGHQKGNSAEARGNR




MADQAAREAAMKAVLETSTLL





XMRV6_
8,135
TLNIEDEYRLHETSKEPDVPLGSTWLSDFPQAWAETGGMGLAVRQAPLII


A1Z651_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mut

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSEQDCQRGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGTAGFCRLWIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEKEAPHDCLEILA




ETHGTRPDLTDQPIPDADYTWYTDGSSFLQEGQRRAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHVHGEIYRR




RGWLTSEGREIKNKNEILALLKALFLPKRLSIIHCPGHQKGNSAEARGNR




MADQAAREAAMKAVLETSTLL





XMRV6_
8,136
TLNIEDEYRLHETSKEPDVPLGSTWLSDFPQAWAETGGMGLAVRQAPLII


A1Z651_

PLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPLLP


3mutA

VKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLD




LKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN




EALHRDLADFRIQHPDLILLQYVDDLLLAATSEQDCQRGTRALLQTLGNL




GYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQL




REFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQA




LLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD




PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPPDR




WLSNARMTHYQAMLLDTDRVQFGPVVALNPATLLPLPEKEAPHDCLEILA




ETHGTRPDLTDQPIPDADYTWYTDGSSFLQEGQRRAGAAVTTETEVIWAR




ALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHVHGEIYRR




RGWLTSEGREIKNKNEILALLKALFLPKRLSIIHCPGHQKGNSAEARGNR




MADQAAREAAMKAVLETSTLL









In some embodiments, reverse transcriptase domains are modified, for example by site-specific mutation. In some embodiments, reverse transcriptase domains are engineered to have improved properties, e.g. SuperScript IV (SSIV) reverse transcriptase derived from the MMLV RT. In some embodiments, the reverse transcriptase domain may be engineered to have lower error rates, e.g., as described in WO2001068895, incorporated herein by reference. In some embodiments, the reverse transcriptase domain may be engineered to be more thermostable. In some embodiments, the reverse transcriptase domain may be engineered to be more processive. In some embodiments, the reverse transcriptase domain may be engineered to have tolerance to inhibitors. In some embodiments, the reverse transcriptase domain may be engineered to be faster. In some embodiments, the reverse transcriptase domain may be engineered to better tolerate modified nucleotides in the RNA template. In some embodiments, the reverse transcriptase domain may be engineered to insert modified DNA nucleotides. In some embodiments, the reverse transcriptase domain is engineered to bind a template RNA. In some embodiments, one or more mutations are chosen from D200N, L603W, T330P, D524G, E562Q, D583N, P51L, S67R, E67K, T197A, H204R, E302K, F309N, W313F, L435G, N454K, H594Q, L671P, E69K, H8Y, T306K, or D653N in the RT domain of murine leukemia virus reverse transcriptase or a corresponding mutation at a corresponding position of another RT domain.


In some embodiments, a gene modifying polypeptide comprises the RT domain from a retroviral reverse transcriptase, e.g., a wild-type M-MLV RT, e.g., comprising the following sequence:











M-MLV (WT):



(SEQ ID NO: 5002)



TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMG






LAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQR






LLDQGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNK






RVEDIHPTVPNPYNLLSGLPPSHQWYTVLDLKDAFFCLRL






HPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD






EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGT






RALLQTLGNLGYRASAKKAQICQKQVKYLGYLLKEGQRWL






TEARKETVMGQPTPKTPRQLREFLGTAGFCRLWIPGFAEM






AAPLYPLTKTGTLFNWGPDQQKAYQEIKQALLTAPALGLP






DLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD






PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAV






EALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNP






ATLLPLPEEGLQHNCLDILAEAHGTRPDLTDQPLPDADHT






WYTDGSSLLQEGQRKAGAAVTTETEVIWAKALPAGTSAQR






AELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR






RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQK






GHSAEARGNRMADQAARKAAITETPDTSTLLI






In some embodiments, a gene modifying polypeptide comprises the RT domain from a retroviral reverse transcriptase, e.g., an M-MLV RT, e.g., comprising the following sequence:











(SEQ ID NO: 5003)



TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMG






LAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQR






LLDQGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNK






RVEDIHPTVPNPYNLLSGLPPSHQWYTVLDLKDAFFCLRL






HPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD






EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGT






RALLQTLGNLGYRASAKKAQICQKQVKYLGYLLKEGQRWL






TEARKETVMGQPTPKTPRQLREFLGTAGFCRLWIPGFAEM






AAPLYPLTKTGTLFNWGPDQQKAYQEIKQALLTAPALGLP






DLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD






PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAV






EALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNP






ATLLPLPEEGLQHNCLDILAEAHGTRPDLTDQPLPDADHT






WYTDGSSLLQEGQRKAGAAVTTETEVIWAKALPAGTSAQR






AELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR






RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQK






GHSAEARGNRMADQAARKAAITETPDTSTLL






In some embodiments, a gene modifying polypeptide comprises the RT domain from a retroviral reverse transcriptase comprising the sequence of amino acids 659-1329 of NP 057933. In embodiments, the gene modifying polypeptide further comprises one additional amino acid at the N-terminus of the sequence of amino acids 659-1329 of NP 057933, e.g., as shown below:











(SEQ ID NO: 5004)



TLNIEDEHRLHETSKEPDVSLGSTWLSDFPQAWAETGGMG






LAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQR






LLDQGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNK







RVEDIHPTVPNPYNLLSGLPPSHQWYTVLDLKDAFFCLRL








HPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFD








EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGT








RALLQTLGNLGYRASAKKAQICQKQVKYLGYLLKEGQRWL






TEARKETVMGQPTPKTPRQLREFLGTAGFCRLWIPGFAEM






AAPLYPLTKTGTLFNWGPDQQKAYQEIKQALLTAPALGLP






DLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD






PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAV






EALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNP






ATLLPLPEEGLQHNCLDILAEAHGTRPDLTDQPLPDADHT







WYTDGSSLLQEGQRKAGAAVTTETEVIWAKALPAGTSAQR








AELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR








RGLLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQK








GHSAEARGNRMADQAARKAA








Core RT (bold), annotated per above


RNAseH (underlined), annotated per above


In embodiments, the gene modifying polypeptide further comprises one additional amino acid at the C-terminus of the sequence of amino acids 659-1329 of NP 057933. In embodiments, the gene modifying polypeptide comprises an RNaseH1 domain (e.g., amino acids 1178-1318 of NP_057933).


In some embodiments, a retroviral reverse transcriptase domain, e.g., M-MLV RT, may comprise one or more mutations from a wild-type sequence that may improve features of the RT, e.g., thermostability, processivity, and/or template binding. In some embodiments, an M-MLV RT domain comprises, relative to the M-MLV (WT) sequence above, one or more mutations, e.g., selected from D200N, L603W, T330P, T306K, W313F, D524G, E562Q, D583N, P51L, S67R, E67K, T197A, H204R, E302K, F309N, L435G, N454K, H594Q, D653N, R110S, K103L, e.g., a combination of mutations, such as D200N, L603W, and T330P, optionally further including T306K and W313F. In some embodiments, an M-MLV RT used herein comprises the mutations D200N, L603W, T330P, T306K and W313F. In embodiments, the mutant M-MLV RT comprises the following amino acid sequence:











M-MLV (PE2):



(SEQ ID NO: 5005)



TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMG






LAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQR






LLDQGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNK






RVEDIHPTVPNPYNLLSGLPPSHQWYTVLDLKDAFFCLRL






HPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFN






EALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGT






RALLQTLGNLGYRASAKKAQICQKQVKYLGYLLKEGQRWL






TEARKETVMGQPTPKTPRQLREFLGKAGFCRLFIPGFAEM






AAPLYPLTKPGTLFNWGPDQQKAYQEIKQALLTAPALGLP






DLTKPFELFVDEKQGYAKGVLTQKLGPWRRPVAYLSKKLD






PVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAV






EALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNP






ATLLPLPEEGLQHNCLDILAEAHGTRPDLTDQPLPDADHT






WYTDGSSLLQEGQRKAGAAVTTETEVIWAKALPAGTSAQR






AELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRR






RGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPGHQK






GHSAEARGNRMADQAARKAAITETPDTSTLLI






In some embodiments, a writing domain (e.g., RT domain) comprises an RNA-binding domain, e.g., that specifically binds to an RNA sequence. In some embodiments, a template RNA comprises an RNA sequence that is specifically bound by the RNA-binding domain of the writing domain.


In some embodiments, the reverse transcription domain only recognizes and reverse transcribes a specific template, e.g., a template RNA of the system. In some embodiments, the template comprises a sequence or structure that enables recognition and reverse transcription by a reverse transcription domain. In some embodiments, the template comprises a sequence or structure that enables association with an RNA-binding domain of a polypeptide component of a genome engineering system described herein. In some embodiments, the genome engineering system reverse preferably transcribes a template comprising an association sequence over a template lacking an association sequence.


The writing domain may also comprise DNA-dependent DNA polymerase activity, e.g., comprise enzymatic activity capable of writing DNA into the genome from a template DNA sequence. In some embodiments, DNA-dependent DNA polymerization is employed to complete second-strand synthesis of a target site edit. In some embodiments, the DNA-dependent DNA polymerase activity is provided by a DNA polymerase domain in the polypeptide. In some embodiments, the DNA-dependent DNA polymerase activity is provided by a reverse transcriptase domain that is also capable of DNA-dependent DNA polymerization, e.g., second-strand synthesis. In some embodiments, the DNA-dependent DNA polymerase activity is provided by a second polypeptide of the system. In some embodiments, the DNA-dependent DNA polymerase activity is provided by an endogenous host cell polymerase that is optionally recruited to the target site by a component of the genome engineering system.


In some embodiments, the reverse transcriptase domain has a lower probability of premature termination rate (Par) in vitro relative to a reference reverse transcriptase domain. In some embodiments, the reference reverse transcriptase domain is a viral reverse transcriptase domain, e.g., the RT domain from M-MLV.


In some embodiments, the reverse transcriptase domain has a lower probability of premature termination rate (Par) in vitro of less than about 5×10−3/nt, 5×10−4/nt, or 5×10−6/nt, e.g., as measured on a 1094 nt RNA. In embodiments, the in vitro premature termination rate is determined as described in Bibillo and Eickbush (2002) J Biol Chem 277(38):34836-34845 (incorporated by reference herein its entirety).


In some embodiments, the reverse transcriptase domain is able to complete at least about 30% or 50% of integrations in cells. The percent of complete integrations can be measured by dividing the number of substantially full-length integration events (e.g., genomic sites that comprise at least 98% of the expected integrated sequence) by the number of total (including substantially full-length and partial) integration events in a population of cells. In embodiments, the integrations in cells is determined (e.g., across the integration site) using long-read amplicon sequencing, e.g., as described in Karst et al. (2020) bioRxiv doi.org/10.1101/645903 (incorporated by reference herein in its entirety).


In embodiments, quantifying integrations in cells comprises counting the fraction of integrations that contain at least about 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% of the DNA sequence corresponding to the template RNA (e.g., a template RNA having a length of at least 0.05, 0.1, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.5, 2, 3, 4, or 5 kb, e.g., a length between 0.5-0.6, 0.6-0.7, 0.7-0.8, 0.8-0.9, 1.0-1.2, 1.2-1.4, 1.4-1.6, 1.6-1.8, 1.8-2.0, 2-3, 3-4, or 4-5 kb).


In some embodiments, the reverse transcriptase domain is capable of polymerizing dNTPs in vitro. In embodiments, the reverse transcriptase domain is capable of polymerizing dNTPs in vitro at a rate between 0.1-50 nt/sec (e.g., between 0.1-1, 1-10, or 10-50 nt/sec). In embodiments, polymerization of dNTPs by the reverse transcriptase domain is measured by a single-molecule assay, e.g., as described in Schwartz and Quake (2009) PNAS 106(48):20294-20299 (incorporated by reference in its entirety).


In some embodiments, the reverse transcriptase domain has an in vitro error rate (e.g., misincorporation of nucleotides) of between 1×10−3-1×10−4 or 1×10−4-1×10−5 substitutions/nt, e.g., as described in Yasukawa et al. (2017) Biochem Biophys Res Commun 492(2):147-153 (incorporated herein by reference in its entirety). In some embodiments, the reverse transcriptase domain has an error rate (e.g., misincorporation of nucleotides) in cells (e.g., HEK293T cells) of between 1×10−3-1×10−4 or 1×10−4-1×10−5 substitutions/nt, e.g., by long-read amplicon sequencing, e.g., as described in Karst et al. (2020) bioRxiv doi.org/10.1101/645903 (incorporated by reference herein in its entirety).


In some embodiments, the reverse transcriptase domain is capable of performing reverse transcription of a target RNA in vitro. In some embodiments, the reverse transcriptase requires a primer of at least 3 nucleotides to initiate reverse transcription of a template. In some embodiments, reverse transcription of the target RNA is determined by detection of cDNA from the target RNA (e.g., when provided with a ssDNA primer, e.g., which anneals to the target with at least 3, 4, 5, 6, 7, 8, 9, or 10 nt at the 3′ end), e.g., as described in Bibillo and Eickbush (2002) J Blot Chem 277(38):34836-34845 (incorporated herein by reference in its entirety).


In some embodiments, the reverse transcriptase domain performs reverse transcription at least 5 or 10 times more efficiently (e.g., by cDNA production), e.g., when converting its RNA template to cDNA, for example, as compared to an RNA template lacking the protein binding motif (e.g., a 3′ UTR). In embodiments, efficiency of reverse transcription is measured as described in Yasukawa et al. (2017) Biochem Biophys Res Commun 492(2):147-153 (incorporated by reference herein in its entirety).


In some embodiments, the reverse transcriptase domain specifically binds a specific RNA template with higher frequency (e.g., about 5 or 10-fold higher frequency) than any endogenous cellular RNA, e.g., when expressed in cells (e.g., HEK293T cells). In embodiments, frequency of specific binding between the reverse transcriptase domain and the template RNA are measured by CLIP-seq, e.g., as described in Lin and Miles (2019) Nucleic Acids Res 47(11):5490-5501 (incorporated herein by reference in its entirety).


Template Nucleic Acid Binding Domain

The gene modifying polypeptide typically contains regions capable of associating with the template nucleic acid (e.g., template RNA). In some embodiments, the template nucleic acid binding domain is an RNA binding domain. In some embodiments, the RNA binding domain is a modular domain that can associate with RNA molecules containing specific signatures, e.g., structural motifs. In other embodiments, the template nucleic acid binding domain (e.g., RNA binding domain) is contained within the reverse transcription domain, e.g., the reverse transcriptase-derived component has a known signature for RNA preference.


In other embodiments, the template nucleic acid binding domain (e.g., RNA binding domain) is contained within the target DNA binding domain. For example, in some embodiments, the DNA binding domain is a CRISPR-associated protein that recognizes the structure of a template nucleic acid (e.g., template RNA) comprising a gRNA. In some embodiments, a gene modifying polypeptide comprises a DNA-binding domain comprising a CRISPR-associated protein that associates with a gRNA scaffold that allows the DNA-binding domain to bind a target genomic DNA sequence. In some embodiments, the gRNA scaffold and gRNA spacer is comprised within the template nucleic acid (e.g., template RNA), thus the DNA-binding domain is also the template nucleic acid binding domain. In some embodiments, the polypeptide possesses RNA binding function in multiple domains, e.g., can bind a gRNA structure in a CRISPR-associated DNA binding domain and an additional sequence or structure in a reverse transcriptase domain.


In some embodiments, the RNA binding domain is capable of binding to a template RNA with greater affinity than a reference RNA binding domain. In some embodiments, the reference RNA binding domain is an RNA binding domain from Cas9 of S. pyogenes. In some embodiments, the RNA binding domain is capable of binding to a template RNA with an affinity between 100 pM-10 nM (e.g., between 100 pM-1 nM or 1 nM-10 nM). In some embodiments, the affinity of a RNA binding domain for its template RNA is measured in vitro, e.g., by thermophoresis, e.g., as described in Asmari et al. Methods 146:107-119 (2018) (incorporated by reference herein in its entirety). In some embodiments, the affinity of a RNA binding domain for its template RNA is measured in cells (e.g., by FRET or CLIP-Seq).


In some embodiments, the RNA binding domain is associated with the template RNA in vitro at a frequency at least about 5-fold or 10-fold higher than with a scrambled RNA. In some embodiments, the frequency of association between the RNA binding domain and the template RNA or scrambled RNA is measured by CLIP-seq, e.g., as described in Lin and Miles (2019) Nucleic Acids Res 47(11):5490-5501 (incorporated by reference herein in its entirety). In some embodiments, the RNA binding domain is associated with the template RNA in cells (e.g., in HEK293T cells) at a frequency at least about 5-fold or 10-fold higher than with a scrambled RNA. In some embodiments, the frequency of association between the RNA binding domain and the template RNA or scrambled RNA is measured by CLIP-seq, e.g., as described in Lin and Miles (2019), supra.


In some embodiments, an RT domain (e.g., as listed in Table 6) comprises one or more mutations as listed in Table 2A below. In some embodiment, an RT domain as listed in Table 6 comprises one, two, three, four, five, or six of the mutations listed in the corresponding row of Table 2A below.









TABLE 2A







Exemplary RT domain mutations (relative to corresponding wild-type


sequences as listed in the corresponding row of Table 6)








RT Domain Name
Mutation(s)
















AVIRE_P03360








AVIRE_P03360_3mut
D200N
G330P
L605W





AVIRE_P03360_3mutA
D200N
G330P
L605W
T306K
W313F



BAEVM_P10272








BAEVM_P10272_3mut
D198N
E328P
L602W





BAEVM_P10272_3mutA
D198N
E328P
L602W
T304K
W311F



BLVAU_P25059








BLVAU_P25059_2mut
E159Q
G286P






BLVJ_P03361








BLVJ_P03361_2mut
E159Q
L524W






BLVJ_P03361_2mutB
E159Q
L524W
197P





FFV_O93209
D21N







FFV_O93209_2mut
D21N
T293N
T419P





FFV_O93209_2mutA
D21N
T293N
T419P
L393K




FFV_O93209-Pro








FFV_O93209-Pro_2mut
T207N
T333P






FFV_O93209-Pro_2mutA
T207N
T333P
L307K





FLV_P10273








FLV_P10273_3mut
D199N
L602W






FLV_P10273_3mutA
D199N
L602W
T305K
W312F




FOAMV_P14350
D24N







FOAMV_P14350_2mut
D24N
T296N
S420P





FOAMV_P14350_2mutA
D24N
T296N
S420P
L396K




FOAMV_P14350-Pro








FOAMV_P14350-Pro_2mut
T207N
S331P






FOAMV_P14350-Pro_2mutA
T207N
S331P
L307K





GALV_P21414








GALV_P21414_3mut
D198N
E328P
L600W





GALV_P21414_3mutA
D198N
E328P
L600W
T304K
W311F



HTL1A_P03362








HTL1A_P03362_2mut
E152Q
R279P






HTL1A_P03362_2mutB
E152Q
R279P
L90P





HTL1C_P14078








HTL1C_P14078_2mut
E152Q
R279P






HTL1L_P0C211








HTL1L_P0C211_2mut
E149Q
L527W






HTL1L_P0C211_2mutB
E149Q
L527W
L87P





HTL32_Q0R5R2








HTL32_Q0R5R2_2mut
E149Q
L526W






HTL32_Q0R5R2_2mutB
E149Q
L526W
L87P





HTL3P_Q4U0X6








HTL3P_Q4U0X6_2mut
E149Q
L526W






HTL3P_Q4U0X6_2mutB
E149Q
L526W
L87P





HTLV2_P03363_2mut
E147Q
G274P






JSRV_P31623








JSRV_P31623_2mutB
A100P







KORV_Q9TTC1
D32N







KORV_Q9TTC1_3mut
D32N
D322N
E452P
L724W




KORV_Q9TTC1_3mutA
D32N
D322N
E452P
L724W
T428K
W435F


KORV_Q9TTC1-Pro








KORV_Q9TTC1-Pro_3mut
D231N
E361P
L633W





KORV_Q9TTC1-Pro_3mutA
D231N
E361P
L633W
T337K
W344F



MLVAV_P03356








MLVAV_P03356_3mut
D200N
T330P
L603W





MLVAV_P03356_3mutA
D200N
T330P
L603W
T306K
W313F



MLVBM_Q7SVK7








MLVBM_Q7SVK7








MLVBM_Q7SVK7_3mut
D200N
T330P
L603W





MLVBM_Q7SVK7_3mut
D200N
T330P
L603W





MLVBM_Q7SVK7_3mutA_WS
D199N
T329P
L602W
T305K
W312F



MLVBM_Q7SVK7_3mutA_WS
D199N
T329P
L602W
T305K
W312F



MLVCB_P08361








MLVCB_P08361_3mut
D200N
T330P
L603W





MLVCB_P08361_3mutA
D200N
T330P
L603W
T306K
W313F



MLVF5_P26810








MLVF5_P26810_3mut
D200N
T330P
L603W





MLVF5_P26810_3mutA
D200N
T330P
L603W
T306K
W313F



MLVFF_P26809_3mut
D200N
T330P
L603W





MLVFF_P26809_3mutA
D200N
T330P
L603W
T306K
W313F



MLVMS_P03355








MLVMS_P03355








MLVMS_P03355_3mut
D200N
T330P
L603W





MLVMS_P03355_3mut
D200N
T330P
L603W





MLVMS_P03355_3mutA_WS
D200N
T330P
L603W
T306K
W313F



MLVMS_P03355_3mutA_WS
D200N
T330P
L603W
T306K
W313F



MLVMS_P03355_PLV919
D200N
T330P
L603W
T306K
W313F
H8Y


MLVMS_P03355_PLV919
D200N
T330P
L603W
T306K
W313F
H8Y


MLVRD_P11227








MLVRD_P11227_3mut
D200N
T330P
L603W





MMTVB_P03365
D26N







MMTVB_P03365
D26N







MMTVB_P03365_2mut
D26N
G401P






MMTVB_P03365_2mut_WS
G400P







MMTVB_P03365_2mut_WS
G400P







MMTVB_P03365_2mutB
D26N
G401P
V215P





MMTVB_P03365_2mutB
D26N
G401P
V215P





MMTVB_P03365_2mutB_WS
G400P
V212P






MMTVB_P03365_2mutB_WS
G400P
V212P






MMTVB_P03365_WS








MMTVB_P03365_WS








MMTVB_P03365-Pro








MMTVB_P03365-Pro








MMTVB_P03365-Pro_2mut
G309P







MMTVB_P03365-Pro_2mut
G309P







MMTVB_P03365-Pro_2mutB
G309P
V123P






MMTVB_P03365-Pro_2mutB
G309P
V123P






MPMV_P07572








MPMV_P07572_2mutB
G289P
I103P






PERV_Q4VFZ2








PERV_Q4VFZ2








PERV_Q4VFZ2_3mut
D199N
E329P
L602W





PERV_Q4VFZ2_3mut
D199N
E329P
L602W





PERV_Q4VFZ2_3mutA_WS
D196N
E326P
L599W
T302K
W309F



PERV_Q4VFZ2_3mutA_WS
D196N
E326P
L599W
T302K
W309F



SFV1_P23074
D24N







SFV1_P23074_2mut
D24N
T296N
N420P





SFV1_P23074_2mutA
D24N
T296N
N420P
L396K




SFV1_P23074-Pro








SFV1_P23074-Pro_2mut
T207N
N331P






SFV1_P23074-Pro_2mutA
T207N
N331P
L307K





SFV3L_P27401
D24N







SFV3L_P27401_2mut
D24N
T296N
N422P





SFV3L_P27401_2mutA
D24N
T296N
N422P
L396K




SFV3L_P27401-Pro








SFV3L_P27401-Pro_2mut
T307N
N333P






SFV3L_P27401-Pro_2mutA
T307N
N333P
L307K





SFVCP_Q87040
D24N







SFVCP_Q87040_2mut
D24N
T296N
K422P





SFVCP_Q87040_2mutA
D24N
T296N
K422P
L396K




SFVCP_Q87040-Pro








SFVCP_Q87040-Pro_2mut
T207N
K333P






SFVCP_Q87040-Pro_2mutA
T207N
K333P
L307K





SMRVH_P03364








SMRVH_P03364_2mut
G288P







SMRVH_P03364_2mutB
G288P
I102P






SRV2_P51517








SRV2_P51517_2mutB
I103P







WDSV_O92815








WDSV_O92815_2mut
S183N
K312P






WDSV_O92815_2mutA
S183N
K312P
L288K
W295F




WMSV_P03359








WMSV_P03359_3mut
D198N
E328P
L600W





WMSV_P03359_3mutA
D198N
E328P
L600W
T304K
W311F



XMRV6_A1Z651








XMRV6_A1Z651_3mut
D200N
T330P
L603W





XMRV6_A1Z651_3mutA
D200N
T330P
L603W
T306K
W313F









Endonuclease Domains and DNA Binding Domains


In some embodiments, a gene modifying polypeptide possesses the function of DNA target site cleavage via an endonuclease domain. In some embodiments, a gene modifying polypeptide comprises a DNA binding domain, e.g., for binding to a target nucleic acid. In some embodiments, a domain (e.g., a Cas domain) of the gene modifying polypeptide comprises two or more smaller domains, e.g., a DNA binding domain and an endonuclease domain. It is understood that when a DNA binding domain (e.g., a Cas domain) is said to bind to a target nucleic acid sequence, in some embodiments, the binding is mediated by a gRNA.


In some embodiments, a domain has two functions. For example, in some embodiments, the endonuclease domain is also a DNA-binding domain. In some embodiments, the endonuclease domain is also a template nucleic acid (e.g., template RNA) binding domain. For example, in some embodiments, a polypeptide comprises a CRISPR-associated endonuclease domain that binds a template RNA comprising a gRNA, binds a target DNA sequence (e.g., with complementarity to a portion of the gRNA), and cuts the target DNA sequence. In some embodiments, an endonuclease domain or endonuclease/DNA-binding domain from a heterologous source can be used or can be modified (e.g., by insertion, deletion, or substitution of one or more residues) in a gene modifying system described herein.


In some embodiments, a nucleic acid encoding the endonuclease domain or endonuclease/DNA binding domain is altered from its natural sequence to have altered codon usage, e.g. improved for human cells. In some embodiments, the endonuclease element is a heterologous endonuclease element, such as a Cas endonuclease (e.g., Cas9), a type-II restriction endonuclease (e.g., FokI), a meganuclease (e.g., I-SceI), or other endonuclease domain.


In certain aspects, the DNA-binding domain of a gene modifying polypeptide described herein is selected, designed, or constructed for binding to a desired host DNA target sequence. In certain embodiments, the DNA-binding domain of the polypeptide is a heterologous DNA-binding element. In some embodiments the heterologous DNA binding element is a zinc-finger element or a TAL effector element, e.g., a zinc-finger or TAL polypeptide or functional fragment thereof. In some embodiments the heterologous DNA binding element is a sequence-guided DNA binding element, such as Cas9, Cpf1, or other CRISPR-related protein that has been altered to have no endonuclease activity. In some embodiments the heterologous DNA binding element retains endonuclease activity. In some embodiments, the heterologous DNA binding element retains partial endonuclease activity to cleave ssDNA, e.g., possesses nickase activity. In specific embodiments, the heterologous DNA-binding domain can be any one or more of Cas9, TAL domain, ZF domain, Myb domain, combinations thereof, or multiples thereof.


In some embodiments, DNA-binding domains are modified, for example by site-specific mutation, increasing or decreasing DNA-binding elements (for example, number and/or specificity of zinc fingers), etc., to alter DNA-binding specificity and affinity. In some embodiments a nucleic acid sequence encoding the DNA binding domain is altered from its natural sequence to have altered codon usage, e.g. improved for human cells. In embodiments, the DNA binding domain comprises one or more modifications relative to a wild-type DNA binding domain, e.g., a modification via directed evolution, e.g., phage-assisted continuous evolution (PACE).


In some embodiments, the DNA binding domain comprises a meganuclease domain (e.g., as described herein, e.g., in the endonuclease domain section), or a functional fragment thereof. In some embodiments, the meganuclease domain possesses endonuclease activity, e.g., double-strand cleavage and/or nickase activity. In other embodiments, the meganuclease domain has reduced activity, e.g., lacks endonuclease activity, e.g., the meganuclease is catalytically inactive. In some embodiments, a catalytically inactive meganuclease is used as a DNA binding domain, e.g., as described in Fonfara et al. Nucleic Acids Res 40(2):847-860 (2012), incorporated herein by reference in its entirety.


In some embodiments, a gene modifying polypeptide comprises a modification to a DNA-binding domain, e.g., relative to the wild-type polypeptide. In some embodiments, the DNA-binding domain comprises an addition, deletion, replacement, or modification to the amino acid sequence of the original DNA-binding domain. In some embodiments, the DNA-binding domain is modified to include a heterologous functional domain that binds specifically to a target nucleic acid (e.g., DNA) sequence of interest. In some embodiments, the functional domain replaces at least a portion (e.g., the entirety of) the prior DNA-binding domain of the polypeptide. In some embodiments, the functional domain comprises a zinc finger (e.g., a zinc finger that specifically binds to the target nucleic acid (e.g., DNA) sequence of interest. In some embodiments, the functional domain comprises a Cas domain (e.g., a Cas domain that specifically binds to the target nucleic acid (e.g., DNA) sequence of interest. In some embodiments, the Cas domain comprises a Cas9 or a mutant or variant thereof (e.g., as described herein). In embodiments, the Cas domain is associated with a guide RNA (gRNA), e.g., as described herein. In embodiments, the Cas domain is directed to a target nucleic acid (e.g., DNA) sequence of interest by the gRNA. In embodiments, the Cas domain is encoded in the same nucleic acid (e.g., RNA) molecule as the gRNA. In embodiments, the Cas domain is encoded in a different nucleic acid (e.g., RNA) molecule from the gRNA.


In some embodiments, the DNA binding domain is capable of binding to a target sequence (e.g., a dsDNA target sequence) with greater affinity than a reference DNA binding domain. In some embodiments, the reference DNA binding domain is a DNA binding domain from Cas9 of S. pyogenes. In some embodiments, the DNA binding domain is capable of binding to a target sequence (e.g., a dsDNA target sequence) with an affinity between 100 pM 10 nM (e.g., between 100 pM-1 nM or 1 nM-10 nM).


In some embodiments, the affinity of a DNA binding domain for its target sequence (e.g., dsDNA target sequence) is measured in vitro, e.g., by thermophoresis, e.g., as described in Asmari et al. Methods 146:107-119 (2018) (incorporated by reference herein in its entirety).


In embodiments, the DNA binding domain is capable of binding to its target sequence (e.g., dsDNA target sequence), e.g., with an affinity between 100 pM-10 nM (e.g., between 100 pM-1 nM or 1 nM-10 nM) in the presence of a molar excess of scrambled sequence competitor dsDNA, e.g., of about 100-fold molar excess.


In some embodiments, the DNA binding domain is found associated with its target sequence (e.g., dsDNA target sequence) more frequently than any other sequence in the genome of a target cell, e.g., human target cell, e.g., as measured by ChIP-seq (e.g., in HEK293T cells), e.g., as described in He and Pu (2010) Curr. Protoc Mol Biol Chapter 21 (incorporated herein by reference in its entirety). In some embodiments, the DNA binding domain is found associated with its target sequence (e.g., dsDNA target sequence) at least about 5-fold or 10-fold, more frequently than any other sequence in the genome of a target cell, e.g., as measured by ChIP-seq (e.g., in HEK293T cells), e.g., as described in He and Pu (2010), supra.


In some embodiments, the endonuclease domain has nickase activity and cleaves one strand of a target DNA. In some embodiments, nickase activity reduces the formation of double-stranded breaks at the target site. In some embodiments, the endonuclease domain creates a staggered nick structure in the first and second strands of a target DNA. In some embodiments, a staggered nick structure generates free 3′ overhangs at the target site. In some embodiments, free 3′ overhangs at the target site improve editing efficiency, e.g., by enhancing access and annealing of a 3′ homology region of a template nucleic acid. In some embodiments, a staggered nick structure reduces the formation of double-stranded breaks at the target site.


In some embodiments, the endonuclease domain cleaves both strands of a target DNA, e.g., results in blunt-end cleavage of a target with no ssDNA overhangs on either side of the cut-site. The amino acid sequence of an endonuclease domain of a gene modifying system described herein may be at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99% identical to the amino acid sequence of an endonuclease domain described herein, e.g., an endonuclease domain from Table 8.


In certain embodiments, the heterologous endonuclease is FokI or a functional fragment thereof. In certain embodiments, the heterologous endonuclease is a Holliday junction resolvase or homolog thereof, such as the Holliday junction resolving enzyme from Sulfolobus solfataricus—Ssol Hje (Govindaraju et al., Nucleic Acids Research 44:7, 2016). In certain embodiments, the heterologous endonuclease is the endonuclease of the large fragment of a spliceosomal protein, such as Prp8 (Mahbub et al., Mobile DNA 8:16, 2017). In certain embodiments, the heterologous endonuclease is derived from a CRISPR-associated protein, e.g., Cas9. In certain embodiments, the heterologous endonuclease is engineered to have only ssDNA cleavage activity, e.g., only nickase activity, e.g., be a Cas9 nickase, e.g., SpCas9 with D10A, H840A, or N863A mutations. Table 8 provides exemplary Cas proteins and mutations associated with nickase activity. In still other embodiments, homologous endonuclease domains are modified, for example by site-specific mutation, to alter DNA endonuclease activity. In still other embodiments, endonuclease domains are modified to reduce DNA-sequence specificity, e.g., by truncation to remove domains that confer DNA-sequence specificity or mutation to inactivate regions conferring DNA-sequence specificity.


In some embodiments, the endonuclease domain has nickase activity and does not form double-stranded breaks. In some embodiments, the endonuclease domain forms single-stranded breaks at a higher frequency than double-stranded breaks, e.g., at least 90%, 95%, 96%, 97%, 98%, or 99% of the breaks are single-stranded breaks, or less than 10%, 5%, 4%, 3%, 2%, or 1% of the breaks are double-stranded breaks. In some embodiments, the endonuclease forms substantially no double-stranded breaks. In some embodiments, the endonuclease does not form detectable levels of double-stranded breaks.


In some embodiments, the endonuclease domain has nickase activity that nicks the target site DNA of the first strand; e.g., in some embodiments, the endonuclease domain cuts the genomic DNA of the target site near to the site of alteration on the strand that will be extended by the writing domain. In some embodiments, the endonuclease domain has nickase activity that nicks the target site DNA of the first strand and does not nick the target site DNA of the second strand. For example, when a polypeptide comprises a CRISPR-associated endonuclease domain having nickase activity, in some embodiments, said CRISPR-associated endonuclease domain nicks the target site DNA strand containing the PAM site (e.g., and does not nick the target site DNA strand that does not contain the PAM site). As a further example, when a polypeptide comprises a CRISPR-associated endonuclease domain having nickase activity, in some embodiments, said CRISPR-associated endonuclease domain nicks the target site DNA strand not containing the PAM site (e.g., and does not nick the target site DNA strand that contains the PAM site).


In some other embodiments, the endonuclease domain has nickase activity that nicks the target site DNA of the first strand and the second strand. Without wishing to be bound by theory, after a writing domain (e.g., RT domain) of a polypeptide described herein polymerizes (e.g., reverse transcribes) from the heterologous object sequence of a template nucleic acid (e.g., template RNA), the cellular DNA repair machinery must repair the nick on the first DNA strand. The target site DNA now contains two different sequences for the first DNA strand: one corresponding to the original genomic DNA (e.g., having a free 5′ end) and a second corresponding to that polymerized from the heterologous object sequence (e.g., having a free 3′ end). It is thought that the two different sequences equilibrate with one another, first one hybridizing the second strand, then the other, and which sequence the cellular DNA repair apparatus incorporates into its repaired target site may be a stochastic process. Without wishing to be bound by theory, it is thought that introducing an additional nick to the second-strand may bias the cellular DNA repair machinery to adopt the heterologous object sequence-based sequence more frequently than the original genomic sequence (Anzalone et al. Nature 576:149-157 (2019)). In some embodiments, the additional nick is positioned at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, or 150 nucleotides 5′ or 3′ of the target site modification (e.g., the insertion, deletion, or substitution) or to the nick on the first strand.


Alternatively or additionally, without wishing to be bound by theory, it is thought that an additional nick to the second strand may promote second-strand synthesis. In some embodiments, where the gene modifying system has inserted or substituted a portion of the first strand, synthesis of a new sequence corresponding to the insertion/substitution in the second strand is necessary.


In some embodiments, the polypeptide comprises a single domain having endonuclease activity (e.g., a single endonuclease domain) and said domain nicks both the first strand and the second strand. For example, in such an embodiment the endonuclease domain may be a CRISPR-associated endonuclease domain, and the template nucleic acid (e.g., template RNA) comprises a gRNA spacer that directs nicking of the first strand and an additional gRNA spacer that directs nicking of the second strand. In some embodiments, the polypeptide comprises a plurality of domains having endonuclease activity, and a first endonuclease domain nicks the first strand and a second endonuclease domain nicks the second strand (optionally, the first endonuclease domain does not (e.g., cannot) nick the second strand and the second endonuclease domain does not (e.g., cannot) nick the first strand).


In some embodiments, the endonuclease domain is capable of nicking a first strand and a second strand. In some embodiments, the first and second strand nicks occur at the same position in the target site but on opposite strands. In some embodiments, the second strand nick occurs in a staggered location, e.g., upstream or downstream, from the first nick. In some embodiments, the endonuclease domain generates a target site deletion if the second strand nick is upstream of the first strand nick. In some embodiments, the endonuclease domain generates a target site duplication if the second strand nick is downstream of the first strand nick. In some embodiments, the endonuclease domain generates no duplication and/or deletion if the first and second strand nicks occur in the same position of the target site. In some embodiments, the endonuclease domain has altered activity depending on protein conformation or RNA-binding status, e.g., which promotes the nicking of the first or second strand (e.g., as described in Christensen et al. PNAS 2006; incorporated by reference herein in its entirety).


In some embodiments, the endonuclease domain comprises a meganuclease, or a functional fragment thereof. In some embodiments, the endonuclease domain comprises a homing endonuclease, or a functional fragment thereof. In some embodiments, the endonuclease domain comprises a meganuclease from the LAGLIDADG (SEQ ID NO: 25693), GIY-YIG, HNH, His-Cys Box, or PD-(D/E) XK families, or a functional fragment or variant thereof, e.g., which possess conserved amino acid motifs, e.g., as indicated in the family names. In some embodiments, the endonuclease domain comprises a meganuclease, or fragment thereof, chosen from, e.g., I-SmaMI (Uniprot F7WD42), I-Seel (Uniprot P03882), I-Anil (Uniprot P03880), I-Dmol (Uniprot P21505), I-CreI (Uniprot P05725), I-Teel (Uniprot P13299), I-OnuI (Uniprot Q4VWW5), or I-Bmol (Uniprot Q9ANR6). In some embodiments, the meganuclease is naturally monomeric, e.g., I-Seel, I-Teel, or dimeric, e.g., I-CreI, in its functional form. For example, the LAGLIDADG (SEQ ID NO: 25693) meganucleases with a single copy of the LAGLIDADG (SEQ ID NO: 25693) motif generally form homodimers, whereas members with two copies of the LAGLIDADG (SEQ ID NO: 25693) motif are generally found as monomers. In some embodiments, a meganuclease that normally forms as a dimer is expressed as a fusion, e.g., the two subunits are expressed as a single ORF and, optionally, connected by a linker, e.g., an I-CreI dimer fusion (Rodriguez-Fornes et al. Gene Therapy 2020; incorporated by reference herein in its entirety). In some embodiments, a meganuclease, or a functional fragment thereof, is altered to favor nickase activity for one strand of a double-stranded DNA molecule, e.g., I-Scel (K1221 and/or K223I) (Niu et al. J Mol Biol 2008), I-Anil (K227M) (McConnell Smith et al. PNAS 2009), I-Dmol (Q42A and/or K120M) (Molina et al. J Biol Chem 2015). In some embodiments, a meganuclease or functional fragment thereof possessing this preference for single-strand cleavage is used as an endonuclease domain, e.g., with nickase activity. In some embodiments, an endonuclease domain comprises a meganuclease, or a functional fragment thereof, which naturally targets or is engineered to target a safe harbor site, e.g., an I-CreI targeting SH6 site (Rodriguez-Fomes et al., supra). In some embodiments, an endonuclease domain comprises a meganuclease, or a functional fragment thereof, with a sequence tolerant catalytic domain, e.g., I-Teel recognizing the minimal motif CNNNG (Kleinstiver et al. PNAS 2012). In some embodiments, a target sequence tolerant catalytic domain is fused to a DNA binding domain, e.g., to direct activity, e.g., by fusing I-Teel to: (i) zinc fingers to create Tev-ZFEs (Kleinstiver et al. PNAS 2012), (ii) other meganucleases to create MegaTevs (Wolfs et al. Nucleic Acids Res 2014), and/or (iii) Cas9 to create TevCas9 (Wolfs et al. PNAS 2016).


In some embodiments, the endonuclease domain comprises a restriction enzyme, e.g., a Type IIS or Type TIP restriction enzyme. In some embodiments, the endonuclease domain comprises a Type IIS restriction enzyme, e.g., FokI, or a fragment or variant thereof. In some embodiments, the endonuclease domain comprises a Type TIP restriction enzyme, e.g., PvuII, or a fragment or variant thereof. In some embodiments, a dimeric restriction enzyme is expressed as a fusion such that it functions as a single chain, e.g., a FokI dimer fusion (Minczuk et al. Nucleic Acids Res 36(12):3926-3938 (2008)).


The use of additional endonuclease domains is described, for example, in Guha and Edgell Int J Mol Sci 18(22):2565 (2017), which is incorporated herein by reference in its entirety.


In some embodiments, a gene modifying polypeptide comprises a modification to an endonuclease domain, e.g., relative to a wild-type Cas protein. In some embodiments, the endonuclease domain comprises an addition, deletion, replacement, or modification to the amino acid sequence of the wild-type Cas protein. In some embodiments, the endonuclease domain is modified to include a heterologous functional domain that binds specifically to and/or induces endonuclease cleavage of a target nucleic acid (e.g., DNA) sequence of interest. In some embodiments, the endonuclease domain comprises a zinc finger. In embodiments, the endonuclease domain comprising the Cas domain is associated with a guide RNA (gRNA), e.g., as described herein. In some embodiments, the endonuclease domain is modified to include a functional domain that does not target a specific target nucleic acid (e.g., DNA) sequence. In embodiments, the endonuclease domain comprises a FokI domain.


In some embodiments, the endonuclease domain is associated with the target dsDNA in vitro at a frequency at least about 5-fold or 10-fold higher than with a scrambled dsDNA. In some embodiments, the endonuclease domain is associated with the target dsDNA in vitro at a frequency at least about 5-fold or 10-fold higher than with a scrambled dsDNA, e.g., in a cell (e.g., a HEK293T cell). In some embodiments, the frequency of association between the endonuclease domain and the target DNA or scrambled DNA is measured by ChIP-seq, e.g., as described in He and Pu (2010) Curr. Protoc Mol Biol Chapter 21 (incorporated by reference herein in its entirety).


In some embodiments, the endonuclease domain can catalyze the formation of a nick at a target sequence, e.g., to an increase of at least about 5-fold or 10-fold relative to a non-target sequence (e.g., relative to any other genomic sequence in the genome of the target cell). In some embodiments, the level of nick formation is determined using NickSeq, e.g., as described in Elacqua et al. (2019) bioRxiv doi.org/10.1101/867937 (incorporated herein by reference in its entirety).


In some embodiments, the endonuclease domain is capable of nicking DNA in vitro. In embodiments, the nick results in an exposed base. In embodiments, the exposed base can be detected using a nuclease sensitivity assay, e.g., as described in Chaudhry and Weinfeld (1995) Nucleic Acids Res 23(19):3805-3809 (incorporated by reference herein in its entirety). In embodiments, the level of exposed bases (e.g., detected by the nuclease sensitivity assay) is increased by at least 10%, 50%, or more relative to a reference endonuclease domain. In some embodiments, the reference endonuclease domain is an endonuclease domain from Cas9 of S. pyogenes.


In some embodiments, the endonuclease domain is capable of nicking DNA in a cell. In embodiments, the endonuclease domain is capable of nicking DNA in a HEK293T cell. In embodiments, an unrepaired nick that undergoes replication in the absence of Rad51 results in increased NHEJ rates at the site of the nick, which can be detected, e.g., by using a Rad51 inhibition assay, e.g., as described in Bothmer et al. (2017) Nat Commun 8:13905 (incorporated by reference herein in its entirety). In embodiments, NHEJ rates are increased above 0-5%. In embodiments, NHEJ rates are increased to 20-70% (e.g., between 30%-60% or 40-50%), e.g., upon Rad51 inhibition.


In some embodiments, the endonuclease domain releases the target after cleavage. In some embodiments, release of the target is indicated indirectly by assessing for multiple turnovers by the enzyme, e.g., as described in Yourik at al. RNA 25(1):35-44 (2019) (incorporated herein by reference in its entirety) and shown in FIG. 2. In some embodiments, the kexp of an endonuclease domain is 1×10−3−1×10−5 min-1 as measured by such methods.


In some embodiments, the endonuclease domain has a catalytic efficiency (kcat/Km) greater than about 1×108 s−1 M−1 in vitro. In embodiments, the endonuclease domain has a catalytic efficiency greater than about 1×105, 1×106, 1×107, or 1×108, s−1 M−1 in vitro. In embodiments, catalytic efficiency is determined as described in Chen et al. (2018) Science 360(6387):436-439 (incorporated herein by reference in its entirety). In some embodiments, the endonuclease domain has a catalytic efficiency (kcat/Km) greater than about 1×108 s−1 M−1 in cells. In embodiments, the endonuclease domain has a catalytic efficiency greater than about 1×105, 1×106, 1×107, or 1×108 s−1 M−1 in cells.


Gene modifying polypeptides comprising Cas domains In some embodiments, a gene modifying polypeptide described herein comprises a Cas domain. In some embodiments, the Cas domain can direct the gene modifying polypeptide to a target site specified by a gRNA spacer, thereby modifying a target nucleic acid sequence in “cis”. In some embodiments, a gene modifying polypeptide is fused to a Cas domain. In some embodiments, a gene modifying polypeptide comprises a CRISPR/Cas domain (also referred to herein as a CRISPR-associated protein). In some embodiments, a CRISPR/Cas domain comprises a protein involved in the clustered regulatory interspaced short palindromic repeat (CRISPR) system, e.g., a Cas protein, and optionally binds a guide RNA, e.g., single guide RNA (sgRNA).


CRISPR systems are adaptive defense systems originally discovered in bacteria and archaea. CRISPR systems use RNA-guided nucleases termed CRISPR-associated or “Cas” endonucleases (e.g., Cas9 or Cpf1) to cleave foreign DNA. For example, in a typical CRISPR-Cas system, an endonuclease is directed to a target nucleotide sequence (e.g., a site in the genome that is to be sequence-edited) by sequence-specific, non-coding “guide RNAs” that target single- or double-stranded DNA sequences. Three classes (I-III) of CRISPR systems have been identified. The class II CRISPR systems use a single Cas endonuclease (rather than multiple Cas proteins). One class II CRISPR system includes a type II Cas endonuclease such as Cas9, a CRISPR RNA (“crRNA”), and a trans-activating crRNA (“tracrRNA”). The crRNA contains a “spacer” sequence, a typically about 20-nucleotide RNA sequence that corresponds to a target DNA sequence (“protospacer”). In the wild-type system, and in some engineered systems, crRNA also contains a region that binds to the tracrRNA to form a partially double-stranded structure that is cleaved by RNase III, resulting in a crRNA/tracrRNA hybrid molecule. A crRNA/tracrRNA hybrid then directs the Cas endonuclease to recognize and cleave a target DNA sequence. A target DNA sequence is generally adjacent to a “protospacer adjacent motif” (“PAM”) that is specific for a given Cas endonuclease and required for cleavage activity at a target site matching the spacer of the crRNA. CRISPR endonucleases identified from various prokaryotic species have unique PAM sequence requirements, e.g., as listed for exemplary Cas enzymes in Table 7; examples of PAM sequences include 5′-NGG (Streptococcus pyogenes), 5′-NNAGAA (Streptococcus thermophilus CRISPR1), 5′-NGGNG (Streptococcus thermophilus CRISPR3), and 5″-NNNGATT (Neisseria meningiditis). Some endonucleases, e.g., Cas9 endonucleases, are associated with G-rich PAM sites, e.g., 5′-NGG, and perform blunt-end cleaving of the target DNA at a location 3 nucleotides upstream from (5′ from) the PAM site. Another class II CRISPR system includes the type V endonuclease Cpf1, which is smaller than Cas9; examples include AsCpfl (from Acidaminococcus sp.) and LbCpfl (from Lachnospiraceae sp.). Cpf1-associated CRISPR arrays are processed into mature crRNAs without the requirement of a tracrRNA; in other words, a Cpf1 system, in some embodiments, comprises only Cpf1 nuclease and a crRNA to cleave a target DNA sequence. Cpf1 endonucleases, are typically associated with T-rich PAM sites, e.g., 5′-TTN. Cpf1 can also recognize a 5″-CTA PAM motif. Cpf1 typically cleaves a target DNA by introducing an offset or staggered double-strand break with a 4- or 5-nucleotide 5′ overhang, for example, cleaving a target DNA with a 5-nucleotide offset or staggered cut located 18 nucleotides downstream from (3′ from) from a PAM site on the coding strand and 23 nucleotides downstream from the PAM site on the complimentary strand; the 5-nucleotide overhang that results from such offset cleavage allows more precise genome editing by DNA insertion by homologous recombination than by insertion at blunt-end cleaved DNA. See, e.g., Zetsche et al. (2015) Cell, 163:759-771.


A variety of CRISPR associated (Cas) genes or proteins can be used in the technologies provided by the present disclosure and the choice of Cas protein will depend upon the particular conditions of the method. Specific examples of Cas proteins include class II systems including Cas1, Cas2, Cas3, Cas4, Cas5, Cash, Cas7, Cas8, Cas9, Cas10, Cpf1, C2C1, or C2C3. In some embodiments, a Cas protein, e.g., a Cas9 protein, may be from any of a variety of prokaryotic species. In some embodiments a particular Cas protein, e.g., a particular Cas9 protein, is selected to recognize a particular protospacer-adjacent motif (PAM) sequence. In some embodiments, a DNA-binding domain or endonuclease domain includes a sequence targeting polypeptide, such as a Cas protein, e.g., Cas9. In certain embodiments a Cas protein, e.g., a Cas9 protein, may be obtained from a bacteria or archaea or synthesized using known methods. In certain embodiments, a Cas protein may be from a gram-positive bacteria or a gram-negative bacteria. In certain embodiments, a Cas protein may be from a Streptococcus (e.g., a S. pyogenes, or a S. thermophilus), a Francisella (e.g., an F. novicida), a Staphylococcus (e.g., an S. aureus), an Acidaminococcus (e.g., an Acidaminococcus sp. BV3L6), a Neisseria (e.g., an N. meningitidis), a Cryptococcus, a Corynebacterium, a Haemophilus, a Eubacterium, a Pasteurella, a Prevotella, a Veillonella, or a Marinobacter.


In some embodiments, a gene modifying polypeptide may comprise the amino acid sequence of SEQ ID NO: 4000 below, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto. In embodiments, the amino acid sequence of SEQ ID NO: 4000 below, or the sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto, is positioned at the N-terminal end of the gene modifying polypeptide. In embodiments, the amino acid sequence of SEQ ID NO: 4000 below, or the sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto, is positioned within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, or 30 amino acids of the N-terminal end of the gene modifying polypeptide.









Exemplary N-terminal NLS-Cas9 domain


(SEQ ID NO: 4000)


MPAAKRVKLDGGDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTD





RHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNE





MAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLR





KKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLV





QTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFG





NLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADL





FLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALV





RQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEEL





LVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREK





IEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQ





SFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAF





LSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNA





SLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTY





AHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFA





NRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQ





TVKVVDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIK





ELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVD





HIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPSEEVVKKMKNYWRQLLNA





KLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRM





NTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL





NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFY





SNIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSM





PQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTV





AYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKE





VKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLA





SHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDK





VLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTST





KEVLDATLIHQSITGLYETRIDLSQLGGDGG 






In some embodiments, a gene modifying polypeptide may comprise the amino acid sequence of SEQ ID NO: 4001 below, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto. In embodiments, the amino acid sequence of SEQ ID NO: 4001 below, or the sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto, is positioned at the C-terminal end of the gene modifying polypeptide. In embodiments, the amino acid sequence of SEQ ID NO: 4001 below, or the sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto, is positioned within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, or 30 amino acids of the C-terminal end of the gene modifying polypeptide.









Exemplary C-terminal sequence comprising an NLS


(SEQ ID NO: 4001)


AGKRTADGSEFEKRTADGSEFESPKKKAKVE 





Exemplary benchmarking sequence


(SEQ ID NO: 4002)


MPAAKRVKLDGGDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTD





RHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNE





MAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLR





KKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLV





QTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFG





NLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADL





FLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALV





RQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEEL





LVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREK





IEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQ





SFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAF





LSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNA





SLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTY





AHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFA





NRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQ





TVKVVDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIK





ELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVD





HIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPSEEVVKKMKNYWRQLLNA





KLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRM





NTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL





NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFY





SNIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSM





PQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTV





AYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKE





VKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLA





SHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDK





VLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTST





KEVLDATLIHQSITGLYETRIDLSQLGGDGGSGGSSGGSSGSETPGTSES





ATPESSGGSSGGSSGGTLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWA





ETGGMGLAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQ





GILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYN





LLSGLPPSHQWYTVLDLKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLT





WTRLPQGFKNSPTLFNEALHRDLADFRIQHPDLILLQYVDDLLLAATSEL





DCQQGTRALLQTLGNLGYRASAKKAQICQKQVKYLGYLLKEGQRWLTEAR





KETVMGQPTPKTPRQLREFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLF





NWGPDQQKAYQEIKQALLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQK





LGPWRRPVAYLSKKLDPVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVI





LAPHAVEALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNPATLL





PLPEEGLQHNCLDILAEAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQR





KAGAAVTTETEVIWAKALPAGTSAQRAELIALTQALKMAEGKKLNVYTDS





RYAFATAHIHGEIYRRRGWLTSEGKEIKNKDEILALLKALFLPKRLSIIH





CPGHQKGHSAEARGNRMADQAARKAAITETPDTSTLLIENSSPSGGSKRT





ADGSEFEAGKRTADGSEFEKRTADGSEFESPKKKAKVE






In some embodiments, a gene modifying polypeptide may comprise a Cas domain as listed in Table 7 or 8, or a functional fragment thereof, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity thereto.









TABLE 7







CRISPR/Cas Proteins, Species, and Mutations
















 

 
Muta-







Muta-
tions 







tions
to







to
make







alter
cata-





#

PAM
lyti- 



En-

of

recog-
cally


Name
zyme
Species
AAs
PAM
nition
dead





FnCas9
Cas9

Franci-
1629
5′-
Wt
D11A/





sella


NGG-3′

H969A/





novicida




N995A





FnCas9
Cas9

Franci-
1629
5′-
E1369R/
D11A/


RHA


sella


YG-3′
E1449H/
H969A/





novicida



R1556A
N995A





SaCas9
Cas9

Staphylo-
1053
5′-
Wt
D10A/





coccus


NNGRRT-

H557A





aureus


3′







SaCas9
Cas9

Staphylo-
1053
5′-
E782K/
D10A/


KKH


coccus


NNNRRT-
N968K/
H557A





aureus


3′
R1015H






SpCas9
Cas9

Strepto-
1368
5′-
Wt
D10A/





coccus


NGG-3′

D839A/





pyogenes




H840A/








N863A





SpCas9
Cas9

Strepto-
1368
5′-
D1135V/
D10A/


VQR


coccus


NGA-3′
R1335Q/
D839A/





pyogenes



T1337R
H840A/








N863A





AsCpf1
Cpf1

Acidamin-
1307
5′-
S542R/
E993A


RR


ococcus


TYCV-3′
K607R





sp. BV3L6









AsCpf1
Cpf1

Acidamin-
1307
5′-
S542R/
E993A


RVR


ococcus


TATV-3′
K548V/





sp. BV3L6


N552R






FnCpf1
Cpf1

Franci-
1300
5′-
Wt
D917A/





sella


NTTN-3′

E1006A/





novicida




D1255A





NmCas9
Cas9

Neisseria

1082
5′-
Wt
D16A/





meningi-

NNNGATT-

D587A/





tidis


3′

H588A/








N611A
















TABLE 8







Amino Acid Sequences of CRISPR/Cas Proteins, Species, and Mutations
















SEQ
Nick-
Nick-
Nick-



Parental

ID
ase
ase
ase


Variant
Host(s)
Protein Sequence
NO:
(HNH)
(HNH)
(RuvC)





Nme2Cas9

Neisseria

MAAFKPNPINYILGLDIGIASVGWAMVEIDEEENPIRLIDLGVRVFERAEVPK
9,001
N611A
H588A
D16A




meningitidis

TGDSLAMARRLARSVRRLTRRRAHRLLRARRLLKREGVLQAADFDENGLIKS








LPNTPWQLRAAALDRKLTPLEWSAVLLHLIKHRGYLSQRKNEGETADKELG








ALLKGVANNAHALQTGDFRTPAELALNKFEKESGHIRNQRGDYSHTFSRKD








LQAELILLFEKQKEFGNPHVSGGLKEGIETLLMTQRPALSGDAVQKMLGHCT








FEPAEPKAAKNTYTAERFIWLTKLNNLRILEQGSERPLTDTERATLMDEPYRK








SKLTYAQARKLLGLEDTAFFKGLRYGKDNAEASTLMEMKAYHAISRALEKEG








LKDKKSPLNLSSELQDEIGTAFSLFKTDEDITGRLKDRVQPEILEALLKHISFDKF








VQISLKALRRIVPLMEQGKRYDEACAEIYGDHYGKKNTEEKIYLPPIPADEIRN








PVVLRALSQARKVINGVVRRYGSPARIHIETAREVGKSFKDRKEIEKRQEENR








KDREKAAAKFREYFPNFVGEPKSKDILKLRLYEQQHGKCLYSGKEINLVRLNE








KGYVEIDHALPFSRTWDDSFNNKVLVLGSENQNKGNQTPYEYFNGKDNSR








EWQEFKARVETSRFPRSKKQRILLQKFDEDGFKECNLNDTRYVNRFLCQFVA








DHILLTGKGKRRVFASNGQITNLLRGFWGLRKVRAENDRHHALDAVVVACS








TVAMQQKITRFVRYKEMNAFDGKTIDKETGKVLHQKTHFPQPWEFFAQEV








MIRVFGKPDGKPEFEEADTPEKLRTLLAEKLSSRPEAVHEYVTPLFVSRAPNR








KMSGAHKDTLRSAKRFVKHNEKISVKRVWLTEIKLADLENMVNYKNGREIEL








YEALKARLEAYGGNAKQAFDPKDNPFYKKGGQLVKAVRVEKTQESGVLLNK








KNAYTIADNGDMVRVDVFCKVDKKGKNQYFIVPIYAWQVAENILPDIDCKG








YRIDDSYTFCFSLHKYDLIAFQKDEKSKVEFAYYINCDSSNGRFYLAWHDKGS








KEQQFRISTQNLVLIQKYQVNELGKEIRPCRLKKRPPVR









PpnCas9

Pasteurella

MQNNPLNYILGLDLGIASIGWAVVEIDEESSPIRLIDVGVRTFERAEVAKTGE
9,002
N605A
H582A
D13A




pneumotropica

SLALSRRLARSSRRLIKRRAERLKKAKRLLKAEKILHSIDEKLPINVWQLRVKGL








KEKLERQEWAAVLLHLSKHRGYLSQRKNEGKSDNKELGALLSGIASNHQML








QSSEYRTPAEIAVKKFQVEEGHIRNQRGSYTHTFSRLDLLAEMELLFQRQAEL








GNSYTSTTLLENLTALLMWQKPALAGDAILKMLGKCTFEPSEYKAAKNSYSA








ERFVWLTKLNNLRILENGTERALNDNERFALLEQPYEKSKLTYAQVRAMLAL








SDNAIFKGVRYLGEDKKTVESKTTLIEMKFYHQIRKTLGSAELKKEWNELKGN








SDLLDEIGTAFSLYKTDDDICRYLEGKLPERVLNALLENLNFDKFIQLSLKALHQ








ILPLMLQGQRYDEAVSAIYGDHYGKKSTETTRLLPTIPADEIRNPVVLRTLTQA








RKVINAVVRLYGSPARIHIETAREVGKSYQDRKKLEKQQEDNRKQRESAVKK








FKEMFPHFVGEPKGKDILKMRLYELQQAKCLYSGKSLELHRLLEKGYVEVDH








ALPFSRTWDDSFNNKVLVLANENQNKGNLTPYEWLDGKNNSERWQHFVV








RVQTSGFSYAKKQRILNHKLDEKGFIERNLNDTRYVARFLCNFIADNMLLVG








KGKRNVFASNGQITALLRHRWGLQKVREQNDRHHALDAVVVACSTVAMQ








QKITRFVRYNEGNVFSGERIDRETGEIIPLHFPSPWAFFKENVEIRIFSENPKLE








LENRLPDYPQYNHEWVQPLFVSRMPTRKMTGQGHMETVKSAKRLNEGLS








VLKVPLTQLKLSDLERMVNRDREIALYESLKARLEQFGNDPAKAFAEPFYKKG








GALVKAVRLEQTQKSGVLVRDGNGVADNASMVRVDVFTKGGKYFLVPIYT








WQVAKGILPNRAATQGKDENDWDIMDEMATFQFSLCQNDLIKLVTKKKTI








FGYFNGLNRATSNINIKEHDLDKSKGKLGIYLEVGVKLAISLEKYQVDELGKNI








RPCRPTKRQHVR









SauCas9

Staphylococcus

MKRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGA
9,003
N580A
H557A
D10A




aureus

RRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSA








ALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKK








DGEVRGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYE








GPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLN








NLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTST








GKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSE








LTQEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKV








DLSQQKEIPTTLVDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNS








KDAQKMINEMQKRNRQTNERIEEIIRTTGKENAKYLIEKIKLHDMQEGKCLYS








LEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQEENSKKGNRTPFQ








YLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFSVQKDFINRNL








VDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERNKG








YKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQ








EYKEIFITPHQIKHIKDFKDYKYSHRVDKKPNRELINDTLYSTRKDDKGNTLIVN








NLNGLYDKDNDKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNPL








YKYYEETGNYLTKYSKKDNGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVKL








SLKPYRFDVYLDNGVYKFVTVKNLDVIKKENYYEVNSKCYEEAKKLKKISNQA








EFIASFYNNDLIKINGELYRVIGVNNDLLNRIEVNMIDITYREYLENMNDKRPP








RIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG









SauCas9-

Staphylococcus

MKRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGA
9,004
N580A
H557A
D10A


KKH

aureus

RRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSA








ALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKK








DGEVRGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYE








GPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLN








NLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTST








GKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSE








LTQEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKV








DLSQQKEIPTTLVDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNS








KDAQKMINEMQKRNRQTNERIEEIIRTTGKENAKYLIEKIKLHDMQEGKCLYS








LEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQEENSKKGNRTPFQ








YLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFSVQKDFINRNL








VDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERNKG








YKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQ








EYKEIFITPHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIV








NNLNGLYDKDNDKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNP








LYKYYEETGNYLTKYSKKDNGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVK








LSLKPYRFDVYLDNGVYKFVTVKNLDVIKKENYYEVNSKCYEEAKKLKKISNQ








AEFIASFYKNDLIKINGELYRVIGVNNDLLNRIEVNMIDITYREYLENMNDKRP








PHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG









SauriCas9

Staphylococcus

MQENQQKQNYILGLDIGITSVGYGLIDSKTREVIDAGVRLFPEADSENNSNR
9,005
N588A
H565A
D15A




auricularis

RSKRGARRLKRRRIHRLNRVKDLLADYQMIDLNNVPKSTDPYTIRVKGLREPL








TKEEFAIALLHIAKRRGLHNISVSMGDEEQDNELSTKQQLQKNAQQLQDKY








VCELQLERLTNINKVRGEKNRFKTEDFVKEVKQLCETQRQYHNIDDQFIQQY








IDLVSTRREYFEGPGNGSPYGWDGDLLKWYEKLMGRCTYFPEELRSVKYAYS








ADLFNALNDLNNLVVTRDDNPKLEYYEKYHIIENVFKQKKNPTLKQIAKEIGV








QDYDIRGYRITKSGKPQFTSFKLYHDLKNIFEQAKYLEDVEMLDEIAKILTIYQ








DEISIKKALDQLPELLTESEKSQIAQLTGYTGTHRLSLKCIHIVIDELWESPENQ








MEIFTRLNLKPKKVEMSEIDSIPTTLVDEFILSPVVKRAFIQSIKVINAVINRFGL








PEDIIIELAREKNSKDRRKFINKLQKQNEATRKKIEQLLAKYGNTNAKYMIEKI








KLHDMQEGKCLYSLEAIPLEDLLSNPTHYEVDHIIPRSVSFDNSLNNKVLVKQ








SENSKKGNRTPYQYLSSNESKISYNQFKQHILNLSKAKDRISKKKRDMLLEER








DINKFEVQKEFINRNLVDTRYATRELSNLLKTYFSTHDYAVKVKTINGGFTNH








LRKVWDFKKHRNHGYKHHAEDALVIANADFLFKTHKALRRTDKILEQPGLE








VNDTTVKVDTEEKYQELFETPKQVKNIKQFRDFKYSHRVDKKPNRQLINDTL








YSTREIDGETYVVQTLKDLYAKDNEKVKKLFTERPQKILMYQHDPKTFEKLM








TILNQYAEAKNPLAAYYEDKGEYVTKYAKKGNGPAIHKIKYIDKKLGSYLDVS








NKYPETQNKLVKLSLKSFRFDIYKCEQGYKMVSIGYLDVLKKDNYYYIPKDKYE








AEKQKKKIKESDLFVGSFYYNDLIMYEDELFRVIGVNSDINNLVELNMVDITY








KDFCEVNNVTGEKRIKKTIGKRVVLIEKYTTDILGNLYKTPLPKKPQLIFKRGEL









SauriCas9-

Staphylococcus

MQENQQKQNYILGLDIGITSVGYGLIDSKTREVIDAGVRLFPEADSENNSNR
9,006
N588A
H565A
D15A


KKH

auricularis

RSKRGARRLKRRRIHRLNRVKDLLADYQMIDLNNVPKSTDPYTIRVKGLREPL








TKEEFAIALLHIAKRRGLHNISVSMGDEEQDNELSTKQQLQKNAQQLQDKY








VCELQLERLTNINKVRGEKNRFKTEDFVKEVKQLCETQRQYHNIDDQFIQQY








IDLVSTRREYFEGPGNGSPYGWDGDLLKWYEKLMGRCTYFPEELRSVKYAYS








ADLFNALNDLNNLVVTRDDNPKLEYYEKYHIIENVFKQKKNPTLKQIAKEIGV








QDYDIRGYRITKSGKPQFTSFKLYHDLKNIFEQAKYLEDVEMLDEIAKILTIYQ








DEISIKKALDQLPELLTESEKSQIAQLTGYTGTHRLSLKCIHIVIDELWESPENQ








MEIFTRLNLKPKKVEMSEIDSIPTTLVDEFILSPVVKRAFIQSIKVINAVINRFGL








PEDIIIELAREKNSKDRRKFINKLQKQNEATRKKIEQLLAKYGNTNAKYMIEKI








KLHDMQEGKCLYSLEAIPLEDLLSNPTHYEVDHIIPRSVSFDNSLNNKVLVKQ








SENSKKGNRTPYQYLSSNESKISYNQFKQHILNLSKAKDRISKKKRDMLLEER








DINKFEVQKEFINRNLVDTRYATRELSNLLKTYFSTHDYAVKVKTINGGFTNH








LRKVWDFKKHRNHGYKHHAEDALVIANADFLFKTHKALRRTDKILEQPGLE








VNDTTVKVDTEEKYQELFETPKQVKNIKQFRDFKYSHRVDKKPNRKLINDTL








YSTREIDGETYVVQTLKDLYAKDNEKVKKLFTERPQKILMYQHDPKTFEKLM








TILNQYAEAKNPLAAYYEDKGEYVTKYAKKGNGPAIHKIKYIDKKLGSYLDVS








NKYPETQNKLVKLSLKSFRFDIYKCEQGYKMVSIGYLDVLKKDNYYYIPKDKYE








AEKQKKKIKESDLFVGSFYKNDLIMYEDELFRVIGVNSDINNLVELNMVDITY








KDFCEVNNVTGEKHIKKTIGKRVVLIEKYTTDILGNLYKTPLPKKPQLIFKRGEL









ScaCas9-

Streptococcus

MEKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTNRKSIKKNLMGALL
9,007
N872A
H849A
D10A


Sc++

canis

FDSGETAEATRLKRTARRRYTRRKNRIRYLQEIFANEMAKLDDSFFQRLEESF








LVEEDKKNERHPIFGNLADEVAYHRNYPTIYHLRKKLADSPEKADLRLIYLALA








HIIKFRGHFLIEGKLNAENSDVAKLFYQLIQTYNQLFEESPLDEIEVDAKGILSA








RLSKSKRLEKLIAVFPNEKKNGLFGNIIALALGLTPNFKSNFDLTEDAKLQLSKD








TYDDDLDELLGQIGDQYADLFSAAKNLSDAILLSDILRSNSEVTKAPLSASMV








KRYDEHHQDLALLKTLVRQQFPEKYAEIFKDDTKNGYAGYVGADKKLRKRS








GKLATEEEFYKFIKPILEKMDGAEELLAKLNRDDLLRKQRTFDNGSIPHQIHLK








ELHAILRRQEEFYPFLKENREKIEKILTFRIPYYVGPLARGNSRFAWLTRKSEEA








ITPWNFEEVVDKGASAQSFIERMTNFDEQLPNKKVLPKHSLLYEYFTVYNEL








TKVKYVTERMRKPEFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS








VEIIGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIE








ERLKTYAHLFDDKVMKQLKRRHYTGWGRLSRKMINGIRDKQSGKTILDFLKS








DGFSNRNFMQLIHDDSLTFKEEIEKAQVSGQGDSLHEQIADLAGSPAIKKGIL








QTVKIVDELVKVMGHKPENIVIEMARENQTTTKGLQQSRERKKRIEEGIKELE








SQILKENPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVP








QSFIKDDSIDNKVLTRSVENRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQ








RKFDNLTKAERGGLSEADKAGFIKRQLVETRQITKHVARILDSRMNTKRDKN








DKPIREVKVITLKSKLVSDFRKDFQLYKVRDINNYHHAHDAYLNAVVGTALIK








KYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKRFFYSNIMNFFKTEVKL








ANGEIRKRPLIETNGETGEVVWNKEKDFATVRKVLAMPQVNIVKKTEVQTG








GFSKESILSKRESAKLIPRKKGWDTRKYGGFGSPTVAYSILVVAKVEKGKAKKL








KSVKVLVGITIMEKGSYEKDPIGFLEAKGYKDIKKELIFKLPKYSLFELENGRRR








MLASAKELQKANELVLPQHLVRLLYYTQNISATTGSNNLGYIEQHREEFKEIF








EKIIDFSEKYILKNKVNSNLKSSFDEQFAVSDSILLSNSFVSLLKYTSFGASGGFT








FLDLDVKQGRLRYQTVTEVLDATLIYQSITGLYETRTDLSQLGGD









SpyCas9

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,008
N863A
H840A
D10A




pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








GELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAF








KYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,009
N863A
H840A
D10A


NG

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








IRPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








RFLQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPRAF








KYFDTTIDRKVYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,010
N863A
H840A
D10A


SpRY

pyogenes

DSGETAERTRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








IRPKRNSDKLIARKKDWDPKKYGGFLWPTVAYSVLVVAKVEKGKSKKLKSVK








ELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLAS








AKQLQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDE








IIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTRLGAPRAF








KYFDTTIDPKQYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









St1Cas9

Streptococcus

MSDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQG
9,011
N622A
H599A
D9A




thermophilus

RRLARRKKHRRVRLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFI








ALKNMVKHRGISYLDDASDDGNSSVGDYAQIVKENSKQLETKTPGQIQLER








YQTYGQLRGDFTVEKDGKKHRLINVFPTSAYRSEALRILQTQQEFNPQITDEF








INRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGILIGKCTFYPDEF








RAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGPAK








LFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETL








DKLAYVLTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGW








HNFSVKLMMELIPELYETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIY








NPVVAKSVRQAIKIVNAAIKEYGDFDNIVIEMARETNEDDEKKAIQKIQKAN








KDEKDAAMLKAANQYNGKAELPHSVFHGHKQLATKIRLWHQQGERCLYT








GKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKGQRTPYQ








ALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLV








DTRYASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYH








HHAVDALIIAASSQLNLWKKQKNTLVSYSEDQLLDIETGELISDDEYKESVFK








APYQHFVDTLKSKEFEDSILFSYQVDSKFNRKISDATIYATRQAKVGKDKADE








TYVLGKIKDIYTQDGYDAFMKIYKKDKSKFLMYRHDPQTFEKVIEPILENYPN








KQINEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLKYYDSKLGNHIDIT








PKDSNNKVVLQSVSPWRADVYFNKTTGKYEILGLKYADLQFEKGTGTYKISQ








EKYNDIKKKEGVDSDSEFKFTLYKNDLLLVKDTETKEQQLFRFLSRTMPKQKH








YVELKPYDKQKFEGGEALIKVLGNVANSGQCKKGLGKSNISIYKVRTDVLGN








QHIIKNEGDKPKLDF









BlatCas9

Brevibacillus

MAYTMGIDVGIASCGWAIVDLERQRIIDIGVRTFEKAENPKNGEALAVPRRE
9,012
N607A
H584A
D8A




laterosporus

ARSSRRRLRRKKHRIERLKHMFVRNGLAVDIQHLEQTLRSQNEIDVWQLRV








DGLDRMLTQKEWLRVLIHLAQRRGFQSNRKTDGSSEDGQVLVNVTENDRL








MEEKDYRTVAEMMVKDEKFSDHKRNKNGNYHGVVSRSSLLVEIHTLFETQ








RQHHNSLASKDFELEYVNIWSAQRPVATKDQIEKMIGTCTFLPKEKRAPKAS








WHFQYFMLLQTINHIRITNVQGTRSLNKEEIEQVVNMALTKSKVSYHDTRKI








LDLSEEYQFVGLDYGKEDEKKKVESKETIIKLDDYHKLNKIFNEVELAKGETWE








ADDYDTVAYALTFFKDDEDIRDYLQNKYKDSKNRLVKNLANKEYTNELIGKV








STLSFRKVGHLSLKALRKIIPFLEQGMTYDKACQAAGFDFQGISKKKRSVVLP








VIDQISNPVVNRALTQTRKVINALIKKYGSPETIHIETARELSKTFDERKNITKD








YKENRDKNEHAKKHLSELGIINPTGLDIVKYKLWCEQQGRCMYSNQPISFER








LKESGYTEVDHIIPYSRSMNDSYNNRVLVMTRENREKGNQTPFEYMGNDT








QRWYEFEQRVTTNPQIKKEKRQNLLLKGFTNRRELEMLERNLNDTRYITKYL








SHFISTNLEFSPSDKKKKVVNTSGRITSHLRSRWGLEKNRGQNDLHHAMDAI








VIAVTSDSFIQQVTNYYKRKERRELNGDDKFPLPWKFFREEVIARLSPNPKEQ








IEALPNHFYSEDELADLQPIFVSRMPKRSITGEAHQAQFRRVVGKTKEGKNIT








AKKTALVDISYDKNGDFNMYGRETDPATYEAIKERYLEFGGNVKKAFSTDLH








KPKKDGTKGPLIKSVRIMENKTLVHPVNKGKGVVYNSSIVRTDVFQRKEKYY








LLPVYVTDVTKGKLPNKVIVAKKGYHDWIEVDDSFTFLFSLYPNDLIFIRQNPK








KKISLKKRIESHSISDSKEVQEIHAYYKGVDSSTAAIEFIIHDGSYYAKGVGVQN








LDCFEKYQVDILGNYFKVKGEKRLELETSDSNHKGKDVNSIKSTSR









cCas9-v16

Staphylococcus

MKRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGA
9,013
N580A
H557A
D10A




aureus

RRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSA








ALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKK








DGEVRGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYE








GPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLN








NLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTST








GKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSE








LTQEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKV








DLSQQKEIPTTLVDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNS








KDAQKMINEMQKRNRQTNERIEEIIRTTGKENAKYLIEKIKLHDMQEGKCLYS








LEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQEENSKKGNRTPFQ








YLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFSVQKDFINRNL








VDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERNKG








YKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQ








EYKEIFITPHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIV








NNLNGLYDKDNDKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNP








LYKYYEETGNYLTKYSKKDNGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVK








LSLKPYRFDVYLDNGVYKFVTVKNLDVIKKENYYEVNSKCYEEAKKLKKISNQ








AEFIASFYKNDLIKINGELYRVIGVNSDKNNLIEVNMIDITYREYLENMNDKRP








PHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG









cCas9-v17

Staphylococcus

MKRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGA
9,014
N580A
H557A
D10A




aureus

RRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSA








ALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKK








DGEVRGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYE








GPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLN








NLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTST








GKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSE








LTQEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKV








DLSQQKEIPTTLVDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNS








KDAQKMINEMQKRNRQTNERIEEIIRTTGKENAKYLIEKIKLHDMQEGKCLYS








LEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQEENSKKGNRTPFQ








YLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFSVQKDFINRNL








VDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERNKG








YKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQ








EYKEIFITPHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIV








NNLNGLYDKDNDKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNP








LYKYYEETGNYLTKYSKKDNGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVK








LSLKPYRFDVYLDNGVYKFVTVKNLDVIKKENYYEVNSKCYEEAKKLKKISNQ








AEFIASFYKNDLIKINGELYRVIGVNNSTRNIVELNMIDITYREYLENMNDKRP








PHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG









cCas9-v21

Staphylococcus

MKRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGA
9,015
N580A
H557A
D10A




aureus

RRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSA








ALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKK








DGEVRGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYE








GPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLN








NLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTST








GKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSE








LTQEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKV








DLSQQKEIPTTLVDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNS








KDAQKMINEMQKRNRQTNERIEEIIRTTGKENAKYLIEKIKLHDMQEGKCLYS








LEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQEENSKKGNRTPFQ








YLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFSVQKDFINRNL








VDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERNKG








YKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQ








EYKEIFITPHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIV








NNLNGLYDKDNDKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNP








LYKYYEETGNYLTKYSKKDNGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVK








LSLKPYRFDVYLDNGVYKFVTVKNLDVIKKENYYEVNSKCYEEAKKLKKISNQ








AEFIASFYKNDLIKINGELYRVIGVNSDDRNIIELNMIDITYREYLENMNDKRP








PHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG









cCas9-v42

Staphylococcus

MKRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGA
9,016
N580A
H557A
D10A




aureus

RRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSA








ALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKK








DGEVRGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYE








GPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLN








NLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTST








GKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSE








LTQEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKV








DLSQQKEIPTTLVDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNS








KDAQKMINEMQKRNRQTNERIEEIIRTTGKENAKYLIEKIKLHDMQEGKCLYS








LEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQEENSKKGNRTPFQ








YLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFSVQKDFINRNL








VDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERNKG








YKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQ








EYKEIFITPHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIV








NNLNGLYDKDNDKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNP








LYKYYEETGNYLTKYSKKDNGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVK








LSLKPYRFDVYLDNGVYKFVTVKNLDVIKKENYYEVNSKCYEEAKKLKKISNQ








AEFIASFYKNDLIKINGELYRVIGVNNNRLNKIELNMIDITYREYLENMNDKRP








PHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG









CdiCas9

Corynebac-
MKYHVGIDVGTFSVGLAAIEVDDAGMPIKTLSLVSHIHDSGLDPDEIKSAVT
9,017
N597A
H573A
D8A




terium

RLASSGIARRTRRLYRRKRRRLQQLDKFIQRQGWPVIELEDYSDPLYPWKVR








diphtheriae

AELAASYIADEKERGEKLSVALRHIARHRGWRNPYAKVSSLYLPDGPSDAFK








AIREEIKRASGQPVPETATVGQMVTLCELGTLKLRGEGGVLSARLQQSDYAR








EIQEICRMQEIGQELYRKIIDVVFAAESPKGSASSRVGKDPLQPGKNRALKAS








DAFQRYRIAALIGNLRVRVDGEKRILSVEEKNLVFDHLVNLTPKKEPEWVTIA








EILGIDRGQLIGTATMTDDGERAGARPPTHDTNRSIVNSRIAPLVDWWKTA








SALEQHAMVKALSNAEVDDFDSPEGAKVQAFFADLDDDVHAKLDSLHLPV








GRAAYSEDTLVRLTRRMLSDGVDLYTARLQEFGIEPSWTPPTPRIGEPVGNP








AVDRVLKTVSRWLESATKTWGAPERVIIEHVREGFVTEKRAREMDGDMRR








RAARNAKLFQEMQEKLNVQGKPSRADLWRYQSVQRQNCQCAYCGSPITF








SNSEMDHIVPRAGQGSTNTRENLVAVCHRCNQSKGNTPFAIWAKNTSIEG








VSVKEAVERTRHWVTDTGMRSTDFKKFTKAVVERFQRATMDEEIDARSME








SVAWMANELRSRVAQHFASHGTTVRVYRGSLTAEARRASGISGKLKFFDGV








GKSRLDRRHHAIDAAVIAFTSDYVAETLAVRSNLKQSQAHRQEAPQWREFT








GKDAEHRAAWRVWCQKMEKLSALLTEDLRDDRVVVMSNVRLRLGNGSA








HKETIGKLSKVKLSSQLSVSDIDKASSEALWCALTREPGFDPKEGLPANPERHI








RVNGTHVYAGDNIGLFPVSAGSIALRGGYAELGSSFHHARVYKITSGKKPAF








AMLRVYTIDLLPYRNQDLFSVELKPQTMSMRQAEKKLRDALATGNAEYLG








WLVVDDELVVDTSKIATDQVKAVEAELGTIRRWRVDGFFSPSKLRLRPLQM








SKEGIKKESAPELSKIIDRPGWLPAVNKLFSDGNVTVVRRDSLGRVRLESTAH








LPVTWKVQ









CjeCas9

Campylobacter

MARILAFDIGISSIGWAFSENDELKDCGVRIFTKVENPKTGESLALPRRLARSA
9,018
N582A
H559A
D8A




jejuni

RKRLARRKARLNHLKHLIANEFKLNYEDYQSFDESLAKAYKGSLISPYELRFRA








LNELLSKQDFARVILHIAKRRGYDDIKNSDDKEKGAILKAIKQNEEKLANYQS








VGEYLYKEYFQKFKENSKEFTNVRNKKESYERCIAQSFLKDELKLIFKKQREFG








FSFSKKFEEEVLSVAFYKRALKDFSHLVGNCSFFTDEKRAPKNSPLAFMFVAL








TRIINLLNNLKNTEGILYTKDDLNALLNEVLKNGTLTYKQTKKLLGLSDDYEFK








GEKGTYFIEFKKYKEFIKALGEHNLSQDDLNEIAKDITLIKDEIKLKKALAKYDLN








QNQIDSLSKLEFKDHLNISFKALKLVTPLMLEGKKYDEACNELNLKVAINEDK








KDFLPAFNETYYKDEVTNPVVLRAIKEYRKVLNALLKKYGKVHKINIELAREVG








KNHSQRAKIEKEQNENYKAKKDAELECEKLGLKINSKNILKLRLFKEQKEFCAY








SGEKIKISDLQDEKMLEIDHIYPYSRSFDDSYMNKVLVFTKQNQEKLNQTPFE








AFGNDSAKWQKIEVLAKNLPTKKQKRILDKNYKDKEQKNFKDRNLNDTRYI








ARLVLNYTKDYLDFLPLSDDENTKLNDTQKGSKVHVEAKSGMLTSALRHTW








GFSAKDRNNHLHHAIDAVIIAYANNSIVKAFSDFKKEQESNSAELYAKKISELD








YKNKRKFFEPFSGFRQKVLDKIDEIFVSKPERKKPSGALHEETFRKEEEFYQSY








GGKEGVLKALELGKIRKVNGKIVKNGDMFRVDIFKHKKTNKFYAVPIYTMDF








ALKVLPNKAVARSKKGEIKDWILMDENYEFCFSLYKDSLILIQTKDMQEPEFV








YYNAFTSSTVSLIVSKHDNKFETLSKNQKILFKNANEKEVIAKSIGIQNLKVFEK








YIVSALGEVTKAEFRQREDFKK









GeoCas9

Geobacillus

MRYKIGLDIGITSVGWAVMNLDIPRIEDLGVRIFDRAENPQTGESLALPRRLA
9,019
N605A
H582A
D8A




stearothermo-
RSARRRLRRRKHRLERIRRLVIREGILTKEELDKLFEEKHEIDVWQLRVEALDR








philus

KLNNDELARVLLHLAKRRGFKSNRKSERSNKENSTMLKHIEENRAILSSYRTV








GEMIVKDPKFALHKRNKGENYTNTIARDDLEREIRLIFSKQREFGNMSCTEEF








ENEYITIWASQRPVASKDDIEKKVGFCTFEPKEKRAPKATYTFQSFIAWEHIN








KLRLISPSGARGLTDEERRLLYEQAFQKNKITYHDIRTLLHLPDDTYFKGIVYDR








GESRKQNENIRFLELDAYHQIRKAVDKVYGKGKSSSFLPIDFDTFGYALTLFKD








DADIHSYLRNEYEQNGKRMPNLANKVYDNELIEELLNLSFTKFGHLSLKALRS








ILPYMEQGEVYSSACERAGYTFTGPKKKQKTMLLPNIPPIANPVVMRALTQA








RKVVNAIIKKYGSPVSIHIELARDLSQTFDERRKTKKEQDENRKKNETAIRQL








MEYGLTLNPTGHDIVKFKLWSEQNGRCAYSLQPIEIERLLEPGYVEVDHVIPY








SRSLDDSYTNKVLVLTRENREKGNRIPAEYLGVGTERWQQFETFVLTNKQFS








KKKRDRLLRLHYDENEETEFKNRNLNDTRYISRFFANFIREHLKFAESDDKQK








VYTVNGRVTAHLRSRWEFNKNREESDLHHAVDAVIVACTTPSDIAKVTAFY








QRREQNKELAKKTEPHFPQPWPHFADELRARLSKHPKESIKALNLGNYDDQ








KLESLQPVFVSRMPKRSVTGAAHQETLRRYVGIDERSGKIQTVVKTKLSEIKL








DASGHFPMYGKESDPRTYEAIRQRLLEHNNDPKKAFQEPLYKPKKNGEPGP








VIRTVKIIDTKNQVIPLNDGKTVAYNSNIVRVDVFEKDGKYYCVPVYTMDIM








KGILPNKAIEPNKPYSEWKEMTEDYTFRFSLYPNDLIRIELPREKTVKTAAGEE








INVKDVFVYYKTIDSANGGLELISHDHRFSLRGVGSRTLKRFEKYQVDVLGNI








YKVRGEKRVGLASSAHSKPGKTIRPLQSTRD









iSpyMac

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,020
N863A
H840A
D10A


Cas9
spp.
DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRKLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLKREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEIQTVGQNGG








LFDDNPKSPLEVTPSKLVPLKKELNPKKYGGYQKPTTAYPVLLITDTKQLIPISV








MNKKQFEQNPVKFLRDRGYQQVGKNDFIKLPKYTLVDIGDGIKRLWASSKEI








HKGNQLVVSKKSQILLYHAHHLDSDLSNDYLQNHNQQFDVLFNEIISFSKKC








KLGKEHIQKIENVYSNKKNSASIEELAESFIKLLGFTQLGATSPFNFLGVKLNQ








KQYKGKKDYILPCTEGTLIRQSITGLYETRVDLSKIGEDSGGSGGSKRTADGSE








FES









NmeCas9

Neisseria

MAAFKPNSINYILGLDIGIASVGWAMVEIDEEENPIRLIDLGVRVFERAEVPK
9,021
N611A
H588A
D16A




meningitidis

TGDSLAMARRLARSVRRLTRRRAHRLLRTRRLLKREGVLQAANFDENGLIKS








LPNTPWQLRAAALDRKLTPLEWSAVLLHLIKHRGYLSQRKNEGETADKELG








ALLKGVAGNAHALQTGDFRTPAELALNKFEKESGHIRNQRSDYSHTFSRKDL








QAELILLFEKQKEFGNPHVSGGLKEGIETLLMTQRPALSGDAVQKMLGHCTF








EPAEPKAAKNTYTAERFIWLTKLNNLRILEQGSERPLTDTERATLMDEPYRKS








KLTYAQARKLLGLEDTAFFKGLRYGKDNAEASTLMEMKAYHAISRALEKEGL








KDKKSPLNLSPELQDEIGTAFSLFKTDEDITGRLKDRIQPEILEALLKHISFDKFV








QISLKALRRIVPLMEQGKRYDEACAEIYGDHYGKKNTEEKIYLPPIPADEIRNP








VVLRALSQARKVINGVVRRYGSPARIHIETAREVGKSFKDRKEIEKRQEENRK








DREKAAAKFREYFPNFVGEPKSKDILKLRLYEQQHGKCLYSGKEINLGRLNEK








GYVEIDHALPFSRTWDDSFNNKVLVLGSENQNKGNQTPYEYFNGKDNSRE








WQEFKARVETSRFPRSKKQRILLQKFDEDGFKERNLNDTRYVNRFLCQFVA








DRMRLTGKGKKRVFASNGQITNLLRGFWGLRKVRAENDRHHALDAVVVA








CSTVAMQQKITRFVRYKEMNAFDGKTIDKETGEVLHQKTHFPQPWEFFAQ








EVMIRVFGKPDGKPEFEEADTLEKLRTLLAEKLSSRPEAVHEYVTPLFVSRAP








NRKMSGQGHMETVKSAKRLDEGVSVLRVPLTQLKLKDLEKMVNREREPKL








YEALKARLEAHKDDPAKAFAEPFYKYDKAGNRTQQVKAVRVEQVQKTGVW








VRNHNGIADNATMVRVDVFEKGDKYYLVPIYSWQVAKGILPDRAVVQGKD








EEDWQLIDDSFNFKFSLHPNDLVEVITKKARMFGYFASCHRGTGNINIRIHD








LDHKIGKNGILEGIGVKTALSFQKYQIDELGKEIRPCRLKKRPPVR









ScaCas9

Streptococcus

MEKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTNRKSIKKNLMGALL
9,022
N872A
H849A
D10A




canis

FDSGETAEATRLKRTARRRYTRRKNRIRYLQEIFANEMAKLDDSFFQRLEESF








LVEEDKKNERHPIFGNLADEVAYHRNYPTIYHLRKKLADSPEKADLRLIYLALA








HIIKFRGHFLIEGKLNAENSDVAKLFYQLIQTYNQLFEESPLDEIEVDAKGILSA








RLSKSKRLEKLIAVFPNEKKNGLFGNIIALALGLTPNFKSNFDLTEDAKLQLSKD








TYDDDLDELLGQIGDQYADLFSAAKNLSDAILLSDILRSNSEVTKAPLSASMV








KRYDEHHQDLALLKTLVRQQFPEKYAEIFKDDTKNGYAGYVGIGIKHRKRTT








KLATQEEFYKFIKPILEKMDGAEELLAKLNRDDLLRKQRTFDNGSIPHQIHLKE








LHAILRRQEEFYPFLKENREKIEKILTFRIPYYVGPLARGNSRFAWLTRKSEEAI








TPWNFEEVVDKGASAQSFIERMTNFDEQLPNKKVLPKHSLLYEYFTVYNELT








KVKYVTERMRKPEFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSV








EIIGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEE








RLKTYAHLFDDKVMKQLKRRHYTGWGRLSRKMINGIRDKQSGKTILDFLKS








DGFSNRNFMQLIHDDSLTFKEEIEKAQVSGQGDSLHEQIADLAGSPAIKKGIL








QTVKIVDELVKVMGHKPENIVIEMARENQTTTKGLQQSRERKKRIEEGIKELE








SQILKENPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVP








QSFIKDDSIDNKVLTRSVENRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQ








RKFDNLTKAERGGLSEADKAGFIKRQLVETRQITKHVARILDSRMNTKRDKN








DKPIREVKVITLKSKLVSDFRKDFQLYKVRDINNYHHAHDAYLNAVVGTALIK








KYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKRFFYSNIMNFFKTEVKL








ANGEIRKRPLIETNGETGEVVWNKEKDFATVRKVLAMPQVNIVKKTEVQTG








GFSKESILSKRESAKLIPRKKGWDTRKYGGFGSPTVAYSILVVAKVEKGKAKKL








KSVKVLVGITIMEKGSYEKDPIGFLEAKGYKDIKKELIFKLPKYSLFELENGRRR








MLASATELQKANELVLPQHLVRLLYYTQNISATTGSNNLGYIEQHREEFKEIF








EKIIDFSEKYILKNKVNSNLKSSFDEQFAVSDSILLSNSFVSLLKYTSFGASGGFT








FLDLDVKQGRLRYQTVTEVLDATLIYQSITGLYETRTDLSQLGGD









ScaCas9-

Streptococcus

MEKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTNRKSIKKNLMGALL
9,023
N872A
H849A
D10A


HiFi-Sc++

canis

FDSGETAEATRLKRTARRRYTRRKNRIRYLQEIFANEMAKLDDSFFQRLEESF








LVEEDKKNERHPIFGNLADEVAYHRNYPTIYHLRKKLADSPEKADLRLIYLALA








HIIKFRGHFLIEGKLNAENSDVAKLFYQLIQTYNQLFEESPLDEIEVDAKGILSA








RLSKSKRLEKLIAVFPNEKKNGLFGNIIALALGLTPNFKSNFDLTEDAKLQLSKD








TYDDDLDELLGQIGDQYADLFSAAKNLSDAILLSDILRSNSEVTKAPLSASMV








KRYDEHHQDLALLKTLVRQQFPEKYAEIFKDDTKNGYAGYVGADKKLRKRS








GKLATEEEFYKFIKPILEKMDGAEELLAKLNRDDLLRKQRTFDNGSIPHQIHLK








ELHAILRRQEEFYPFLKENREKIEKILTFRIPYYVGPLARGNSRFAWLTRKSEEA








ITPWNFEEVVDKGASAQSFIERMTNFDEQLPNKKVLPKHSLLYEYFTVYNEL








TKVKYVTERMRKPEFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS








VEIIGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIE








ERLKTYAHLFDDKVMKQLKRRHYTGWGRLSRKMINGIRDKQSGKTILDFLKS








DGFSNANFMQLIHDDSLTFKEEIEKAQVSGQGDSLHEQIADLAGSPAIKKGIL








QTVKIVDELVKVMGHKPENIVIEMARENQTTTKGLQQSRERKKRIEEGIKELE








SQILKENPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVP








QSFIKDDSIDNKVLTRSVENRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQ








RKFDNLTKAERGGLSEADKAGFIKRQLVETRQITKHVARILDSRMNTKRDKN








DKPIREVKVITLKSKLVSDFRKDFQLYKVRDINNYHHAHDAYLNAVVGTALIK








KYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKRFFYSNIMNFFKTEVKL








ANGEIRKRPLIETNGETGEVVWNKEKDFATVRKVLAMPQVNIVKKTEVQTG








GFSKESILSKRESAKLIPRKKGWDTRKYGGFGSPTVAYSILVVAKVEKGKAKKL








KSVKVLVGITIMEKGSYEKDPIGFLEAKGYKDIKKELIFKLPKYSLFELENGRRR








MLASAKELQKANELVLPQHLVRLLYYTQNISATTGSNNLGYIEQHREEFKEIF








EKIIDFSEKYILKNKVNSNLKSSFDEQFAVSDSILLSNSFVSLLKYTSFGASGGFT








FLDLDVKQGRLRYQTVTEVLDATLIYQSITGLYETRTDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,024
N863A
H840A
D10A


3var-NRRH

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MVKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEE








FYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGIIPHQIHLGELHAILRRQ








GDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRLRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRN








FMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVV








DELVKVMGGHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQI








LKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKGNSDKLIARKKDWDPKKYGGFNSPTAAYSVLVVAKVEKGKSKKLKSVK








ELLGITIMERSSFEKNPIGFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLAS








AGVLHKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDE








IIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGVPAA








FKYFDTTIDKKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,025
N863A
H840A
D10A


3var-NRTH

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MVKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEE








FYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGIIPHQIHLGELHAILRRQ








GDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRLRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRN








FMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVV








DELVKVMGGHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQI








LKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKGNSDKLIARKKDWDPKKYGGFNSPTVAYSVLVVAKVEKGKSKKLKSVK








ELLGITIMERSSFEKNPIGFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLAS








ASVLHKGNELALPSKYVNFLYLASHYEKLKGSSEDNKQKQLFVEQHKHYLDEI








IEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGASAAF








KYFDTTIGRKLYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,026
N863A
H840A
D10A


3var-NRCH

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MVKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEE








FYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGIIPHQIHLGELHAILRRQ








GDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRLRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRN








FMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVV








DELVKVMGGHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQI








LKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKGNSDKLIARKKDWDPKKYGGFNSPTVAYSVLVVAKVEKGKSKKLKSVK








ELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLAS








AGVLQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDE








IIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAA








FKYFDTTINRKQYNTTKEVLDATLIRQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,027
N863A
H840A
D10A


HF1

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








GELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAF








KYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,028
N863A
H840A
D10A


QQR1

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








RELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADAQLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAF








KYFDTTFKQKQYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,029
N863A
H840A
D10A


SpG

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFLWPTVAYSVLVVAKVEKGKSKKLKSVK








ELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLAS








AKQLQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDE








IIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAA








FKYFDTTIDRKQYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,030
N863A
H840A
D10A


VQR

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








GELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAF








KYFDTTIDRKQYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,031
N863A
H840A
10A


VRER

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQ








EDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFE








EVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKV








VDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQ








ILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








RELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAF








KYFDTTIDRKEYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,032
N863A
H840A
D10A


xCas

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDTKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKLYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGIIPHQIHLGELHAILRRQE








DFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEK








VVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGDQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFIQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVV








DELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQI








LKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








ILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








GVLQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAF








KYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD









SpyCas9-

Streptococcus

MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLF
9,033
N863A
H840A
D10A


xCas-NG

pyogenes

DSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFL








VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAH








MIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILS








ARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDTKLQLS








KDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSAS








MIKLYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEF








YKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGIIPHQIHLGELHAILRRQE








DFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEK








VVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTE








GMRKPAFLSGDQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVED








RFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYA








HLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANR








NFIQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVV








DELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQI








LKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSF








LKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKF








DNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI








REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPK








LESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI








RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKES








IRPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKE








LLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASA








RFLQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEII








EQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPRAF








KYFDTTIDRKVYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD









St1Cas9-

Streptococcus

MSDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQG
9,034
N622A
H599A
D9A


CNRZ1066

thermophilus

RRLARRKKHRRVRLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFI








ALKNMVKHRGISYLDDASDDGNSSVGDYAQIVKENSKQLETKTPGQIQLER








YQTYGQLRGDFTVEKDGKKHRLINVFPTSAYRSEALRILQTQQEFNPQITDEF








INRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGILIGKCTFYPDEF








RAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGPAK








LFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETL








DKLAYVLTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGW








HNFSVKLMMELIPELYETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIY








NPVVAKSVRQAIKIVNAAIKEYGDFDNIVIEMARETNEDDEKKAIQKIQKAN








KDEKDAAMLKAANQYNGKAELPHSVFHGHKQLATKIRLWHQQGERCLYT








GKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKGQRTPYQ








ALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLV








DTRYASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYH








HHAVDALIIAASSQLNLWKKQKNTLVSYSEEQLLDIETGELISDDEYKESVFKA








PYQHFVDTLKSKEFEDSILFSYQVDSKFNRKISDATIYATRQAKVGKDKKDET








YVLGKIKDIYTQDGYDAFMKIYKKDKSKFLMYRHDPQTFEKVIEPILENYPNK








QMNEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLKYYDSKLLGNPIDI








TPENSKNKVVLQSLKPWRTDVYFNKATGKYEILGLKYADLQFEKGTGTYKIS








QEKYNDIKKKEGVDSDSEFKFTLYKNDLLLVKDTETKEQQLFRFLSRTLPKQK








HYVELKPYDKQKFEGGEALIKVLGNVANGGQCIKGLAKSNISIYKVRTDVLG








NQHIIKNEGDKPKLDF









St1Cas9-

Streptococcus

MSDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQG
9,035
N622A
H599A
D9A


LMG1831

thermophilus

RRLARRKKHRRVRLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFI








ALKNMVKHRGISYLDDASDDGNSSVGDYAQIVKENSKQLETKTPGQIQLER








YQTYGQLRGDFTVEKDGKKHRLINVFPTSAYRSEALRILQTQQEFNPQITDEF








INRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGILIGKCTFYPDEF








RAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGPAK








LFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETL








DKLAYVLTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGW








HNFSVKLMMELIPELYETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIY








NPVVAKSVRQAIKIVNAAIKEYGDFDNIVIEMARETNEDDEKKAIQKIQKAN








KDEKDAAMLKAANQYNGKAELPHSVFHGHKQLATKIRLWHQQGERCLYT








GKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKGQRTPYQ








ALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLV








DTRYASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYH








HHAVDALIIAASSQLNLWKKQKNTLVSYSEEQLLDIETGELISDDEYKESVFKA








PYQHFVDTLKSKEFEDSILFSYQVDSKFNRKISDATIYATRQAKVGKDKKDET








YVLGKIKDIYTQDGYDAFMKIYKKDKSKFLMYRHDPQTFEKVIEPILENYPNK








QMNEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLKYYDSKLLGNPIDI








TPENSKNKVVLQSLKPWRTDVYFNKNTGKYEILGLKYADLQFEKKTGTYKISQ








EKYNGIMKEEGVDSDSEFKFTLYKNDLLLVKDTETKEQQLFRFLSRTMPNVK








YYVELKPYSKDKFEKNESLIEILGSADKSGRCIKGLGKSNISIYKVRTDVLGNQH








IIKNEGDKPKLDF









St1Cas9-

Streptococcus

MSDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQG
9,036
N622A
H599A
D9A


MTH17CL3

thermophilus

RRLARRKKHRRVRLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFI






96

ALKNMVKHRGISYLDDASDDGNSSVGDYAQIVKENSKQLETKTPGQIQLER








YQTYGQLRGDFTVEKDGKKHRLINVFPTSAYRSEALRILQTQQEFNPQITDEF








INRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGILIGKCTFYPDEF








RAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGPAK








LFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETL








DKLAYVLTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGW








HNFSVKLMMELIPELYETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIY








NPVVAKSVRQAIKIVNAAIKEYGDFDNIVIEMARETNEDDEKKAIQKIQKAN








KDEKDAAMLKAANQYNGKAELPHSVFHGHKQLATKIRLWHQQGERCLYT








GKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKGQRTPYQ








ALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLV








DTRYASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYH








HHAVDALIIAASSQLNLWKKQKNTLVSYSEDQLLDIETGELISDDEYKESVFK








APYQHFVDTLKSKEFEDSILFSYQVDSKFNRKISDATIYATRQAKVGKDKADE








TYVLGKIKDIYTQDGYDAFMKIYKKDKSKFLMYRHDPQTFEKVIEPILENYPN








KQINEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLKYYDSKLGNHIDIT








PKDSNNKVVLQSLKPWRTDVYFNKNTGKYEILGLKYSDMQFEKGTGKYSISK








EQYENIKVREGVDENSEFKFTLYKNDLLLLKDSENGEQILLRFTSRNDTSKHYV








ELKPYNRQKFEGSEYLIKSLGTVAKGGQCIKGLGKSNISIYKVRTDVLGNQHII








KNEGDKPKLDF









St1Cas9-

Streptococcus

MSDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQG
9,037
N622A
H599A
D9A


TH1477

thermophilus

RRLARRKKHRRVRLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFI








ALKNMVKHRGISYLDDASDDGNSSVGDYAQIVKENSKQLETKTPGQIQLER








YQTYGQLRGDFTVEKDGKKHRLINVFPTSAYRSEALRILQTQQEFNPQITDEF








INRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGILIGKCTFYPDEF








RAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGPAK








LFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETL








DKLAYVLTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGW








HNFSVKLMMELIPELYETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIY








NPVVAKSVRQAIKIVNAAIKEYGDFDNIVIEMARETNEDDEKKAIQKIQKAN








KDEKDAAMLKAANQYNGKAELPHSVFHGHKQLATKIRLWHQQGERCLYT








GKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKGQRTPYQ








ALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLV








DTRYASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYH








HHAVDALIIAASSQLNLWKKQKNTLVSYSEDQLLDIETGELISDDEYKESVFK








APYQHFVDTLKSKEFEDSILFSYQVDSKFNRKISDATIYATRQAKVGKDKADE








TYVLGKIKDIYTQDGYDAFMKIYKKDKSKFLMYRHDPQTFEKVIEPILENYPN








KQINEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLKYYDSKLGNHIDIT








PKDSNNKVVLQSLKPWRTDVYFNKNTGKYEILGLKYSDMQFEKGTGKYSISK








EQYENIKVREGVDENSEFKFTLYKNDLLLLKDSENGEQILLRFTSRNDTSKHYV








ELKPYNRQKFEGSEYLIKSLGTVVKGGRCIKGLGKSNISIYKVRTDVLGNQHIIK








NEGDKPKLDF









SRGN3.1

Staphylococcus

MNQKFILGLDIGITSVGYGLIDYETKNIIDAGVRLFPEANVENNEGRRSKRGS
9,038
N585A
H562A
D10A



spp.
RRLKRRRIHRLERVKLLLTEYDLINKEQIPTSNNPYQIRVKGLSEILSKDELAIAL








LHLAKRRGIHNVDVAADKEETASDSLSTKDQINKNAKFLESRYVCELQKERLE








NEGHVRGVENRFLTKDIVREAKKIIDTQMQYYPEIDETFKEKYISLVETRREYF








EGPGQGSPFGWNGDLKKWYEMLMGHCTYFPQELRSVKYAYSADLFNALN








DLNNLIIQRDNSEKLEYHEKYHIIENVFKQKKKPTLKQIAKEIGVNPEDIKGYRI








TKSGTPEFTSFKLFHDLKKVVKDHAILDDIDLLNQIAEILTIYQDKDSIVAELGQ








LEYLMSEADKQSISELTGYTGTHSLSLKCMNMIIDELWHSSMNQMEVFTYL








NMRPKKYELKGYQRIPTDMIDDAILSPVVKRTFIQSINVINKVIEKYGIPEDIIIE








LARENNSDDRKKFINNLQKKNEATRKRINEIIGQTGNQNAKRIVEKIRLHDQ








QEGKCLYSLESIPLEDLLNNPNHYEVDHIIPRSVSFDNSYHNKVLVKQSENSK








KSNLTPYQYFNSGKSKLSYNQFKQHILNLSKSQDRISKKKKEYLLEERDINKFE








VQKEFINRNLVDTRYATRELTNYLKAYFSANNMNVKVKTINGSFTDYLRKV








WKFKKERNHGYKHHAEDALIIANADFLFKENKKLKAVNSVLEKPEIETKQLDI








QVDSEDNYSEMFIIPKQVQDIKDFRNFKYSHRVDKKPNRQLINDTLYSTRKK








DNSTYIVQTIKDIYAKDNTTLKKQFDKSPEKFLMYQHDPRTFEKLEVIMKQYA








NEKNPLAKYHEETGEYLTKYSKKNNGPIVKSLKYIGNKLGSHLDVTHQFKSST








KKLVKLSIKNYRFDVYLTEKGYKFVTIAYLNVFKKDNYYYIPKDKYQELKEKKKI








KDTDQFIASFYKNDLIKLNGDLYKIIGVNSDDRNIIELDYYDIKYKDYCEINNIK








GEPRIKKTIGKKTESIEKFTTDVLGNLYLHSTEKAPQLIFKRGL









sRGN3.3

Staphylococcus

MNQKFILGLDIGITSVGYGLIDYETKNIIDAGVRLFPEANVENNEGRRSKRGS
9,039
N585A
H562A
D10A



spp.
RRLKRRRIHRLERVKLLLTEYDLINKEQIPTSNNPYQIRVKGLSEILSKDELAIAL








LHLAKRRGIHNVDVAADKEETASDSLSTKDQINKNAKFLESRYVCELQKERLE








NEGHVRGVENRFLTKDIVREAKKIIDTQMQYYPEIDETFKEKYISLVETRREYF








EGPGQGSPFGWNGDLKKWYEMLMGHCTYFPQELRSVKYAYSADLFNALN








DLNNLIIQRDNSEKLEYHEKYHIIENVFKQKKKPTLKQIAKEIGVNPEDIKGYRI








TKSGTPEFTSFKLFHDLKKVVKDHAILDDIDLLNQIAEILTIYQDKDSIVAELGQ








LEYLMSEADKQSISELTGYTGTHSLSLKCMNMIIDELWHSSMNQMEVFTYL








NMRPKKYELKGYQRIPTDMIDDAILSPVVKRTFIQSINVINKVIEKYGIPEDIIIE








LARENNSDDRKKFINNLQKKNEATRKRINEIIGQTGNQNAKRIVEKIRLHDQ








QEGKCLYSLESIPLEDLLNNPNHYEVDHIIPRSVSFDNSYHNKVLVKQSENSK








KSNLTPYQYFNSGKSKLSYNQFKQHILNLSKSQDRISKKKKEYLLEERDINKFE








VQKEFINRNLVDTRYATRELTSYLKAYFSANNMDVKVKTINGSFTNHLRKV








WRFDKYRNHGYKHHAEDALIIANADFLFKENKKLQNTNKILEKPTIENNTKK








VTVEKEEDYNNVFETPKLVEDIKQYRDYKFSHRVDKKPNRQLINDTLYSTRM








KDEHDYIVQTITDIYGKDNTNLKKQFNKNPEKFLMYQNDPKTFEKLSIIMKQ








YSDEKNPLAKYYEETGEYLTKYSKKNNGPIVKKIKLLGNKVGNHLDVTNKYEN








STKKLVKLSIKNYRFDVYLTEKGYKFVTIAYLNVFKKDNYYYIPKDKYQELKEKK








KIKDTDQFIASFYKNDLIKLNGDLYKIIGVNSDDRNIIELDYYDIKYKDYCEINNI








KGEPRIKKTIGKKTESIEKFTTDVLGNLYLHSTEKAPQLIFKRGL









In some embodiments, a Cas protein requires a protospacer adjacent motif (PAM) to be present in or adjacent to a target DNA sequence for the Cas protein to bind and/or function. In some embodiments, the PAM is or comprises, from 5′ to 3′, NGG, YG, NNGRRT, NNNRRT, NGA, TYCV, TATV, NTTN, or NNNGATT, where N stands for any nucleotide, Y stands for C or T, R stands for A or G, and V stands for A or C or G. In some embodiments, a Cas protein is a protein listed in Table 7 or 8. In some embodiments, a Cas protein comprises one or more mutations altering its PAM. In some embodiments, a Cas protein comprises E1369R, E1449H, and R1556A mutations or analogous substitutions to the amino acids corresponding to said positions. In some embodiments, a Cas protein comprises E782K, N968K, and R1015H mutations or analogous substitutions to the amino acids corresponding to said positions. In some embodiments, a Cas protein comprises D1135V, R1335Q, and T1337R mutations or analogous substitutions to the amino acids corresponding to said positions. In some embodiments, a Cas protein comprises S542R and K607R mutations or analogous substitutions to the amino acids corresponding to said positions. In some embodiments, a Cas protein comprises S542R, K548V, and N552R mutations or analogous substitutions to the amino acids corresponding to said positions. Exemplary advances in the engineering of Cas enzymes to recognize altered PAM sequences are reviewed in Collias et al Nature Communications 12:555 (2021), incorporated herein by reference in its entirety.


In some embodiments, the Cas protein is catalytically active and cuts one or both strands of the target DNA site. In some embodiments, cutting the target DNA site is followed by formation of an alteration, e.g., an insertion or deletion, e.g., by the cellular repair machinery.


In some embodiments, the Cas protein is modified to deactivate or partially deactivate the nuclease, e.g., nuclease-deficient Cas9. Whereas wild-type Cas9 generates double-strand breaks (DSBs) at specific DNA sequences targeted by a gRNA, a number of CRISPR endonucleases having modified functionalities are available, for example: a “nickase” version of Cas9 that has been partially deactivated generates only a single-strand break; a catalytically inactive Cas9 (“dCas9”) does not cut target DNA. In some embodiments, dCas9 binding to a DNA sequence may interfere with transcription at that site by steric hindrance. In some embodiments, dCas9 binding to an anchor sequence may interfere with (e.g., decrease or prevent) genomic complex (e.g., ASMC) formation and/or maintenance. In some embodiments, a DNA-binding domain comprises a catalytically inactive Cas9, e.g., dCas9. Many catalytically inactive Cas9 proteins are known in the art. In some embodiments, dCas9 comprises mutations in each endonuclease domain of the Cas protein, e.g., D10A and H840A or N863A mutations. In some embodiments, a catalytically inactive or partially inactive CRISPR/Cas domain comprises a Cas protein comprising one or more mutations, e.g., one or more of the mutations listed in Table 7. In some embodiments, a Cas protein described on a given row of Table 7 comprises one, two, three, or all of the mutations listed in the same row of Table 7. In some embodiments, a Cas protein, e.g., not described in Table 7, comprises one, two, three, or all of the mutations listed in a row of Table 7 or a corresponding mutation at a corresponding site in that Cas protein.


In some embodiments, a catalytically inactive, e.g., dCas9, or partially deactivated Cas9 protein comprises a D11 mutation (e.g., D11A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a H969 mutation (e.g., H969A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a N995 mutation (e.g., N995A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, comprises mutations at one, two, or three of positions D11, H969, and N995 (e.g., D11A, H969A, and N995A mutations) or analogous substitutions to the amino acids corresponding to said positions.


In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a D10 mutation (e.g., a D10A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a H557 mutation (e.g., a H557A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, comprises a D10 mutation (e.g., a D1OA mutation) and a H557 mutation (e.g., a H557A mutation) or analogous substitutions to the amino acids corresponding to said positions.


In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a D839 mutation (e.g., a D839A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a H840 mutation (e.g., a H840A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a N863 mutation (e.g., a N863A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, comprises a D10 mutation (e.g., D10A), a D839 mutation (e.g., D839A), a H840 mutation (e.g., H840A), and a N863 mutation (e.g., N863A) or analogous substitutions to the amino acids corresponding to said positions.


In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a E993 mutation (e.g., a E993A mutation) or an analogous substitution to the amino acid corresponding to said position.


In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a D917 mutation (e.g., a D917A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a a E1006 mutation (e.g., a E1006A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a D1255 mutation (e.g., a D1255A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, comprises a D917 mutation (e.g., D917A), a E1006 mutation (e.g., E1006A), and a D1255 mutation (e.g., D1255A) or analogous substitutions to the amino acids corresponding to said positions.


In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a D16 mutation (e.g., a D16A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a D587 mutation (e.g., a D587A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a partially deactivated Cas domain has nickase activity. In some embodiments, a partially deactivated Cas9 domain is a Cas9 nickase domain. In some embodiments, the catalytically inactive Cas domain or dead Cas domain produces no detectable double strand break formation. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a H588 mutation (e.g., a H588A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, or partially deactivated Cas9 protein comprises a N611 mutation (e.g., a N611A mutation) or an analogous substitution to the amino acid corresponding to said position. In some embodiments, a catalytically inactive Cas9 protein, e.g., dCas9, comprises a D16 mutation (e.g., D16A), a D587 mutation (e.g., D587A), a H588 mutation (e.g., H588A), and a N611 mutation (e.g., N611A) or analogous substitutions to the amino acids corresponding to said positions.


In some embodiments, a DNA-binding domain or endonuclease domain may comprise a Cas molecule comprising or linked (e.g., covalently) to a gRNA (e.g., a template nucleic acid, e.g., template RNA, comprising a gRNA).


In some embodiments, an endonuclease domain or DNA binding domain comprises a Streptococcus pyogenes Cas9 (SpCas9) or a functional fragment or variant thereof. In some embodiments, the endonuclease domain or DNA binding domain comprises a modified SpCas9. In embodiments, the modified SpCas9 comprises a modification that alters protospacer-adjacent motif (PAM) specificity. In embodiments, the PAM has specificity for the nucleic acid sequence 5′-NGT-3′. In embodiments, the modified SpCas9 comprises one or more amino acid substitutions, e.g., at one or more of positions L1111, D1135, G1218, E1219, A1322, of R1335, e.g., selected from L1111R, D1135V, G1218R, E1219F, A1322R, R1335V. In embodiments, the modified SpCas9 comprises the amino acid substitution T1337R and one or more additional amino acid substitutions, e.g., selected from L1111, D1135L, S1136R, G1218S, E1219V, D1332A, D1332S, D1332T, D1332V, D1332L, D1332K, D1332R, R1335Q, T1337, T1337L, T1337Q, T1337I, T1337V, T1337F, T1337S, T1337N, T1337K, T1337H, T1337Q, and T1337M, or corresponding amino acid substitutions thereto. In embodiments, the modified SpCas9 comprises: (i) one or more amino acid substitutions selected from D1135L, S1136R, G1218S, E1219V, A1322R, R1335Q, and T1337; and (ii) one or more amino acid substitutions selected from L1111R, G1218R, E1219F, D1332A, D1332S, D1332T, D1332V, D1332L, D1332K, D1332R, T1337L, T1337I, T1337V, T1337F, T1337S, T1337N, T1337K, T1337R, T1337H, T1337Q, and T1337M, or corresponding amino acid substitutions thereto.


In some embodiments, the endonuclease domain or DNA binding domain comprises a Cas domain, e.g., a Cas9 domain. In embodiments, the endonuclease domain or DNA binding domain comprises a nuclease-active Cas domain, a Cas nickase (nCas) domain, or a nuclease-inactive Cas (dCas) domain. In embodiments, the endonuclease domain or DNA binding domain comprises a nuclease-active Cas9 domain, a Cas9 nickase (nCas9) domain, or a nuclease-inactive Cas9 (dCas9) domain. In some embodiments, the endonuclease domain or DNA binding domain comprises a Cas9 domain of Cas9 (e.g., dCas9 and nCas9), Cas12a/Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12d/CasY, Cas12e/CasX, Cas12g, Cas12h, or Cas12i. In some embodiments, the endonuclease domain or DNA binding domain comprises a Cas9 (e.g., dCas9 and nCas9), Cas12a/Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12d/CasY, Cas12e/CasX, Cas12g, Cas12h, or Cas12i. In some embodiments, the endonuclease domain or DNA binding domain comprises an S. pyogenes or an S. thermophilus Cas9, or a functional fragment thereof. In some embodiments, the endonuclease domain or DNA binding domain comprises a Cas9 sequence, e.g., as described in Chylinski, Rhun, and Charpentier (2013) RNA Biology 10:5, 726-737; incorporated herein by reference. In some embodiments, the endonuclease domain or DNA binding domain comprises the HNH nuclease subdomain and/or the RuvC1 subdomain of a Cas, e.g., Cas9, e.g., as described herein, or a variant thereof. In some embodiments, the endonuclease domain or DNA binding domain comprises Cas12a/Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12d/CasY, Cas12e/CasX, Cas12g, Cas12h, or Cas12i. In some embodiments, the endonuclease domain or DNA binding domain comprises a Cas polypeptide (e.g., enzyme), or a functional fragment thereof. In embodiments, the Cas polypeptide (e.g., enzyme) is selected from Cas1, Cas1B, Cas2, Cas3, Cas4, Cas5, Cas5d, Cas5t, Cas5h, Cas5a, Cash, Cas7, Cas8, Cas8a, Cas8b, Cas8c, Cas9 (e.g., Csn1 or Csx12), Cas10, Cas10d, Cas12a/Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12d/CasY, Cas12e/CasX, Cas12g, Cas12h, Cas12i, Csy1, Csy2, Csy3, Csy4, Cse1, Cse2, Cse3, Cse4, Cse5e, Csc1, Csc2, Csa5, Csn1, Csn2, Csm1, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10, Csx16, CsaX, Csx3, Csx1, Csx1S, Csx11, Csf1, Csf2, CsO, Csf4, Csd1, Csd2, Cst1, Cst2, Csh1, Csh2, Csa1, Csa2, Csa3, Csa4, Csa5, Type II Cas effector proteins, Type V Cas effector proteins, Type VI Cas effector proteins, CARF, DinG, Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12b/C2c1, Cas12c/C2c3, SpCas9(K855A), eSpCas9(1.1), SpCas9-HF1, hyper accurate Cas9 variant (HypaCas9), homologues thereof, modified or engineered versions thereof, and/or functional fragments thereof. In embodiments, the Cas9 comprises one or more substitutions, e.g., selected from H840A, D10A, P475A, W476A, N477A, D1125A, W1126A, and D1127A. In embodiments, the Cas9 comprises one or more mutations at positions selected from: D10, G12, G17, E762, H840, N854, N863, H982, H983, A984, D986, and/or A987, e.g., one or more substitutions selected from D1OA, G12A, G17A, E762A, H840A, N854A, N863A, H982A, H983A, A984A, and/or D986A. In some embodiments, the endonuclease domain or DNA binding domain comprises a Cas (e.g., Cas9) sequence from Corynebacterium ulcerans, Corynebacterium diphtheria, Spiroplasma syrphidicola, Prevotella intermedia, Spiroplasma taiwanense, Streptococcus iniae, Belliella baltica, Psychroflexus torquis, Streptococcus thermophilus, Listeria innocua, Campylobacter jejuni, Neisseria meningitidis, Streptococcus pyogenes, or Staphylococcus aureus, or a fragment or variant thereof.


In some embodiments, the endonuclease domain or DNA binding domain comprises a Cpf1 domain, e.g., comprising one or more substitutions, e.g., at position D917, E1006A, D1255 or any combination thereof, e.g., selected from D917A, E1006A, D1255A, D917A/E1006A, D917A/D1255A, E1006A/D1255A, and D917A/E1006A/D1255A.


In some embodiments, the endonuclease domain or DNA binding domain comprises spCas9, spCas9-VRQR, spCas9-VRER, xCas9 (sp), saCas9, saCas9-KKH, spCas9-MQKSER, spCas9-LRKIQK, or spCas9-LRVSQL.


In some embodiments, a gene modifying polypeptide has an endonuclease domain comprising a Cas9 nickase, e.g., Cas9 H840A. In embodiments, the Cas9 H840A has the following amino acid sequence:









Cas9 nickase (H840A):


(SEQ ID NO: 11,001)


DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGAL





LFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRL





EESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADL





RLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPI





NASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPN





FKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAIL





LSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIF





FDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRK





QRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYY





VGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKN





LPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDL





LFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKII





KDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQL





KRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDS





LTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVM





GRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPV





ENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDS





IDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLT





KAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLIR





EVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKY





PKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEIT





LANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQ





TGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEK





GKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKY





SLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSPED





NEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKP





IREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQS 





ITGLYETRIDLSQLGGD






In some embodiments, a gene modifying polypeptide comprises a dCas9 sequence comprising a D10A and/or H840A mutation, e.g., the following sequence:









(SEQ ID NO: 5007)


SMDKKYSIGLAIGTNSVGWAVITDDYKVPSKKFKVLGNTDRHSIKKNLIG





ALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFH





RLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKA





DLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEEN





PINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLT





PNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDA





ILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKE





IFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL





RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIP





YYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFD





KNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIV





DLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLK





IIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKVMK





QLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHD





DSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVK





VMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEH





PVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKD





DSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDN





LTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKL





IREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIK





KYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE





ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTE





VQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKV





EKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLP





KYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSP





EDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRD





KPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIH





QSITGLYETRIDLSQLGGD






TAL Effectors and Zinc Finger Nucleases


In some embodiments, an endonuclease domain or DNA-binding domain comprises a TAL effector molecule. A TAL effector molecule, e.g., a TAL effector molecule that specifically binds a DNA sequence, typically comprises a plurality of TAL effector domains or fragments thereof, and optionally one or more additional portions of naturally occurring TAL effectors (e.g., N- and/or C-terminal of the plurality of TAL effector domains). Many TAL effectors are known to those of skill in the art and are commercially available, e.g., from Thermo Fisher Scientific.


Naturally occurring TALEs are natural effector proteins secreted by numerous species of bacterial pathogens including the plant pathogen Xanthomonas which modulates gene expression in host plants and facilitates bacterial colonization and survival. The specific binding of TAL effectors is based on a central repeat domain of tandemly arranged nearly identical repeats of typically 33 or 34 amino acids (the repeat-variable di-residues, RVD domain).


Members of the TAL effectors family differ mainly in the number and order of their repeats. The number of repeats typically ranges from 1.5 to 33.5 repeats and the C-terminal repeat is usually shorter in length (e.g., about 20 amino acids) and is generally referred to as a “half-repeat.” Each repeat of the TAL effector generally features a one-repeat-to-one-base-pair correlation with different repeat types exhibiting different base-pair specificity (one repeat recognizes one base-pair on the target gene sequence). Generally, the smaller the number of repeats, the weaker the protein-DNA interactions. A number of 6.5 repeats has been shown to be sufficient to activate transcription of a reporter gene (Scholze et al., 2010).


Repeat to repeat variations occur predominantly at amino acid positions 12 and 13, which have therefore been termed “hypervariable” and which are responsible for the specificity of the interaction with the target DNA promoter sequence, as shown in Table 9 listing exemplary repeat variable diresidues (RVD) and their correspondence to nucleic acid base targets.









TABLE 9







RVDs and Nucleic Acid Base Specificity








Target
Possible RVD Amino Acid Combinations























A
NI
NN
CI
HI
KI













G
NN
GN
SN
VN
LN
DN
QN
EN
HN
RH
NK
AN
FN





C
HD
RD
KD
ND
AD













T
NG
HG
VG
IG
EG
MG
YG
AA
EP
VA
QG
KG
RG









Accordingly, it is possible to modify the repeats of a TAL effector to target specific DNA sequences. Further studies have shown that the RVD NK can target G. Target sites of TAL effectors also tend to include a T flanking the 5′ base targeted by the first repeat, but the exact mechanism of this recognition is not known. More than 113 TAL effector sequences are known to date. Non-limiting examples of TAL effectors from Xanthomonas include, Hax2, Hax3, Hax4, AvrXa7, AvrXa10 and AvrBs3.


Accordingly, the TAL effector domain of a TAL effector molecule described herein may be derived from a TAL effector from any bacterial species (e.g., Xanthomonas species such as the African strain of Xanthomonas oryzae pv. Oryzae (Yu et al. 2011), Xanthomonas campestris pv. raphani strain 756C and Xanthomonas oryzae pv. Oryzicola strain BLS256 (Bogdanove et al. 2011). In some embodiments, the TAL effector domain comprises an RVD domain as well as flanking sequence(s) (sequences on the N-terminal and/or C-terminal side of the RVD domain) also from the naturally occurring TAL effector. It may comprise more or fewer repeats than the RVD of the naturally occurring TAL effector. The TAL effector molecule can be designed to target a given DNA sequence based on the above code and others known in the art. The number of TAL effector domains (e.g., repeats (monomers or modules)) and their specific sequence can beselected based on the desired DNA target sequence. For example, TAL effector domains, e.g., repeats, may be removed or added in order to suit a specific target sequence. In an embodiment, the TAL effector molecule of the present invention comprises between 6.5 and 33.5 TAL effector domains, e.g., repeats. In an embodiment, TAL effector molecule of the present invention comprises between 8 and 33.5 TAL effector domains, e.g., repeats, e.g., between 10 and 25 TAL effector domains, e.g., repeats, e.g., between 10 and 14 TAL effector domains, e.g., repeats.


In some embodiments, the TAL effector molecule comprises TAL effector domains that correspond to a perfect match to the DNA target sequence. In some embodiments, a mismatch between a repeat and a target base-pair on the DNA target sequence is permitted as along as it allows for the function of the polypeptide comprising the TAL effector molecule. In general, TALE binding is inversely correlated with the number of mismatches. In some embodiments, the TAL effector molecule of a polypeptide of the present invention comprises no more than 7 mismatches, 6 mismatches, 5 mismatches, 4 mismatches, 3 mismatches, 2 mismatches, or 1 mismatch, and optionally no mismatch, with the target DNA sequence. Without wishing to be bound by theory, in general the smaller the number of TAL effector domains in the TAL effector molecule, the smaller the number of mismatches will be tolerated and still allow for the function of the polypeptide comprising the TAL effector molecule. The binding affinity is thought to depend on the sum of matching repeat-DNA combinations. For example, TAL effector molecules having 25 TAL effector domains or more may be able to tolerate up to 7 mismatches.


In addition to the TAL effector domains, the TAL effector molecule of the present invention may comprise additional sequences derived from a naturally occurring TAL effector. The length of the C-terminal and/or N-terminal sequence(s) included on each side of the TAL effector domain portion of the TAL effector molecule can vary and be selected by one skilled in the art, for example based on the studies of Zhang et al. (2011). Zhang et al., have characterized a number of C-terminal and N-terminal truncation mutants in Hax3 derived TAL-effector based proteins and have identified key elements, which contribute to optimal binding to the target sequence and thus activation of transcription. Generally, it was found that transcriptional activity is inversely correlated with the length of N-terminus. Regarding the C-terminus, an important element for DNA binding residues within the first 68 amino acids of the Hax 3 sequence was identified. Accordingly, in some embodiments, the first 68 amino acids on the C-terminal side of the TAL effector domains of the naturally occurring TAL effector is included in the TAL effector molecule. Accordingly, in an embodiment, a TAL effector molecule comprises 1) one or more TAL effector domains derived from a naturally occurring TAL effector; 2) at least 70, 80, 90, 100, 110, 120, 130, 140, 150, 170, 180, 190, 200, 220, 230, 240, 250, 260, 270, 280 or more amino acids from the naturally occurring TAL effector on the N-terminal side of the TAL effector domains; and/or 3) at least 68, 80, 90, 100, 110, 120, 130, 140, 150, 170, 180, 190, 200, 220, 230, 240, 250, 260 or more amino acids from the naturally occurring TAL effector on the C-terminal side of the TAL effector domains.


In some embodiments, an endonuclease domain or DNA-binding domain is or comprises a Zn finger molecule. A Zn finger molecule comprises a Zn finger protein, e.g., a naturally occurring Zn finger protein or engineered Zn finger protein, or fragment thereof. Many Zn finger proteins are known to those of skill in the art and are commercially available, e.g., from Sigma-Aldrich.


In some embodiments, a Zn finger molecule comprises a non-naturally occurring Zn finger protein that is engineered to bind to a target DNA sequence of choice. See, for example, Beerli, et al. (2002) Nature Biotechnol. 20:135-141; Pabo, et al. (2001) Ann. Rev. Biochem. 70:313-340; Isalan, et al. (2001) Nature Biotechnol. 19:656-660; Segal, et al. (2001) Curr. Opin. Biotechnol. 12:632-637; Choo, et al. (2000) Curr. Opin. Struct. Biol. 10:411-416; U.S. Pat. Nos. 6,453,242; 6,534,261; 6,599,692; 6,503,717; 6,689,558; 7,030,215; 6,794,136; 7,067,317; 7,262,054; 7,070,934; 7,361,635; 7,253,273; and U.S. Patent Publication Nos. 2005/0064474; 2007/0218528; 2005/0267061, all incorporated herein by reference in their entireties.


An engineered Zn finger protein may have a novel binding specificity, compared to a naturally-occurring Zn finger protein. Engineering methods include, but are not limited to, rational design and various types of selection. Rational design includes, for example, using databases comprising triplet (or quadruplet) nucleotide sequences and individual Zn finger amino acid sequences, in which each triplet or quadruplet nucleotide sequence is associated with one or more amino acid sequences of zinc fingers which bind the particular triplet or quadruplet sequence. See, for example, U.S. Pat. Nos. 6,453,242 and 6,534,261, incorporated by reference herein in their entireties.


Exemplary selection methods, including phage display and two-hybrid systems, are disclosed in U.S. Pat. Nos. 5,789,538; 5,925,523; 6,007,988; 6,013,453; 6,410,248; 6,140,466; 6,200,759; and 6,242,568; as well as International Patent Publication Nos. WO 98/37186; WO 98/53057; WO 00/27878; and WO 01/88197 and GB 2,338,237. In addition, enhancement of binding specificity for zinc finger proteins has been described, for example, in International Patent Publication No. WO 02/077227.


In addition, as disclosed in these and other references, zinc finger domains and/or multi-fingered zinc finger proteins may be linked together using any suitable linker sequences, including for example, linkers of 5 or more amino acids in length. See, also, U.S. Pat. Nos. 6,479,626; 6,903,185; and 7,153,949 for exemplary linker sequences 6 or more amino acids in length. The proteins described herein may include any combination of suitable linkers between the individual zinc fingers of the protein. In addition, enhancement of binding specificity for zinc finger binding domains has been described, for example, in co-owned International Patent Publication No. WO 02/077227.


Zn finger proteins and methods for design and construction of fusion proteins (and polynucleotides encoding same) are known to those of skill in the art and described in detail in U.S. Pat. Nos. 6,140,0815; 789,538; 6,453,242; 6,534,261; 5,925,523; 6,007,988; 6,013,453; and 6,200,759; International Patent Publication Nos. WO 95/19431; WO 96/06166; WO 98/53057; WO 98/54311; WO 00/27878; WO 01/60970; WO 01/88197; WO 02/099084; WO 98/53058; WO 98/53059; WO 98/53060; WO 02/016536; and WO 03/016496.


In addition, as disclosed in these and other references, Zn finger proteins and/or multi-fingered Zn finger proteins may be linked together, e.g., as a fusion protein, using any suitable linker sequences, including for example, linkers of 5 or more amino acids in length. See, also, U.S. Pat. Nos. 6,479,626; 6,903,185; and 7,153,949 for exemplary linker sequences 6 or more amino acids in length. The Zn finger molecules described herein may include any combination of suitable linkers between the individual zinc finger proteins and/or multi-fingered Zn finger proteins of the Zn finger molecule.


In certain embodiments, the DNA-binding domain or endonuclease domain comprises a Zn finger molecule comprising an engineered zinc finger protein that binds (in a sequence-specific manner) to a target DNA sequence. In some embodiments, the Zn finger molecule comprises one Zn finger protein or fragment thereof. In other embodiments, the Zn finger molecule comprises a plurality of Zn finger proteins (or fragments thereof), e.g., 2, 3, 4, 5, 6 or more Zn finger proteins (and optionally no more than 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2 Zn finger proteins). In some embodiments, the Zn finger molecule comprises at least three Zn finger proteins. In some embodiments, the Zn finger molecule comprises four, five or six fingers. In some embodiments, the Zn finger molecule comprises 8, 9, 10, 11 or 12 fingers. In some embodiments, a Zn finger molecule comprising three Zn finger proteins recognizes a target DNA sequence comprising 9 or 10 nucleotides. In some embodiments, a Zn finger molecule comprising four Zn finger proteins recognizes a target DNA sequence comprising 12 to 14 nucleotides. In some embodiments, a Zn finger molecule comprising six Zn finger proteins recognizes a target DNA sequence comprising 18 to 21 nucleotides.


In some embodiments, a Zn finger molecule comprises a two-handed Zn finger protein. Two handed zinc finger proteins are those proteins in which two clusters of zinc finger proteins are separated by intervening amino acids so that the two zinc finger domains bind to two discontinuous target DNA sequences. An example of a two handed type of zinc finger binding protein is SIP1, where a cluster of four zinc finger proteins is located at the amino terminus of the protein and a cluster of three Zn finger proteins is located at the carboxyl terminus (see Remade, et al. (1999) EMBO Journal 18(18):5073-5084). Each cluster of zinc fingers in these proteins is able to bind to a unique target sequence and the spacing between the two target sequences can comprise many nucleotides.


Linkers


In some embodiments, a gene modifying polypeptide may comprise a linker, e.g., a peptide linker, e.g., a linker as described in Table 10. In some embodiments, a gene modifying polypeptide comprises, in an N-terminal to C-terminal direction, a Cas domain (e.g., a Cas domain of Table 8), a linker of Table 10 (or a sequence having at least 70%, 80%, 85%, 90%, 95%, or 99% identity thereto), and an RT domain (e.g., an RT domain of Table 6). In some embodiments, a gene modifying polypeptide comprises a flexible linker between the endonuclease and the RT domain, e.g., a linker comprising the amino acid sequence SGGSSGGSSGSETPGTSESATPESSGGSSGGSS (SEQ ID NO: 11,002). In some embodiments, an RT domain of a gene modifying polypeptide may be located C-terminal to the endonuclease domain. In some embodiments, an RT domain of a gene modifying polypeptide may be located N-terminal to the endonuclease domain.









TABLE 10







Exemplary linker sequences











SEQ



Amino Acid Sequence
ID NO






GGS







GGSGGS
5102






GGSGGSGGS
5103






GGSGGSGGSGGS
5104






GGSGGSGGSGGSGGS
5105






GGSGGSGGSGGSGGSGGS
5106






GGGGS
5107






GGGGSGGGGS
5108






GGGGSGGGGSGGGGS
5109






GGGGSGGGGSGGGGSGGGGS
5110






GGGGSGGGGSGGGGSGGGGSGGGGS
5111






GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS
5112






GGG







GGGG
5114






GGGGG
5115






GGGGGG
5116






GGGGGGG
5117






GGGGGGGG
5118






GSS







GSSGSS
5120






GSSGSSGSS
5121






GSSGSSGSSGSS
5122






GSSGSSGSSGSSGSS
5123






GSSGSSGSSGSSGSSGSS
5124






EAAAK
5125






EAAAKEAAAK
5126






EAAAKEAAAKEAAAK
5127






EAAAKEAAAKEAAAKEAAAK
5128






EAAAKEAAAKEAAAKEAAAKEAAAK
5129






EAAAKEAAAKEAAAKEAAAKEAAAKEAAAK
5130






PAP







PAPAP
5132






PAPAPAP
5133






PAPAPAPAP
5134






PAPAPAPAPAP
5135






PAPAPAPAPAPAP
5136






GGSGGG
5137






GGGGGS
5138






GGSGSS
5139






GSSGGS
5140






GGSEAAAK
5141






EAAAKGGS
5142






GGSPAP
5143






PAPGGS
5144






GGGGSS
5145






GSSGGG
5146






GGGEAAAK
5147






EAAAKGGG
5148






GGGPAP
5149






PAPGGG
5150






GSSEAAAK
5151






EAAAKGSS
5152






GSSPAP
5153






PAPGSS
5154






EAAAKPAP
5155






PAPEAAAK
5156






GGSGGGGSS
5157






GGSGSSGGG
5158






GGGGGSGSS
5159






GGGGSSGGS
5160






GSSGGSGGG
5161






GSSGGGGGS
5162






GGSGGGEAAAK
5163






GGSEAAAKGGG
5164






GGGGGSEAAAK
5165






GGGEAAAKGGS
5166






EAAAKGGSGGG
5167






EAAAKGGGGGS
5168






GGSGGGPAP
5169






GGSPAPGGG
5170






GGGGGSPAP
5171






GGGPAPGGS
5172






PAPGGSGGG
5173






PAPGGGGGS
5174






GGSGSSEAAAK
5175






GGSEAAAKGSS
5176






GSSGGSEAAAK
5177






GSSEAAAKGGS
5178






EAAAKGGSGSS
5179






EAAAKGSSGGS
5180






GGSGSSPAP
5181






GGSPAPGSS
5182






GSSGGSPAP
5183






GSSPAPGGS
5184






PAPGGSGSS
5185






PAPGSSGGS
5186






GGSEAAAKPAP
5187






GGSPAPEAAAK
5188






EAAAKGGSPAP
5189






EAAAKPAPGGS
5190






PAPGGSEAAAK
5191






PAPEAAAKGGS
5192






GGGGSSEAAAK
5193






GGGEAAAKGSS
5194






GSSGGGEAAAK
5195






GSSEAAAKGGG
5196






EAAAKGGGGSS
5197






EAAAKGSSGGG
5198






GGGGSSPAP
5199






GGGPAPGSS
5200






GSSGGGPAP
5201






GSSPAPGGG
5202






PAPGGGGSS
5203






PAPGSSGGG
5204






GGGEAAAKPAP
5205






GGGPAPEAAAK
5206






EAAAKGGGPAP
5207






EAAAKPAPGGG
5208






PAPGGGEAAAK
5209






PAPEAAAKGGG
5210






GSSEAAAKPAP
5211






GSSPAPEAAAK
5212






EAAAKGSSPAP
5213






EAAAKPAPGSS
5214






PAPGSSEAAAK
5215






PAPEAAAKGSS
5216






AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
5217



AKEAAAKEAAAKA







GGGGSEAAAKGGGGS
5218






EAAAKGGGGSEAAAK
5219






SGSETPGTSESATPES
5220






GSAGSAAGSGEF
5221






SGGSSGGSSGSETPGTSESATPESSGGSSGGSS
5222









In some embodiments, a linker of a gene modifying polypeptide comprises a motif chosen from: (SGGS)n (SEQ ID NO: 5025), (GGGS)n (SEQ ID NO: 5026), (GGGGS)n (SEQ ID NO: 5027), (G)n, (EAAAK)n (SEQ ID NO: 5028), (GGS)n, or (XP)n.


Gene Modifying Polypeptide Selection by Pooled Screening


Candidate gene modifying polypeptides may be screened to evaluate a candidate's gene editing ability. For example, an RNA gene modifying system designed for the targeted editing of a coding sequence in the human genome may be used. In certain embodiments, such a gene modifying system may be used in conjunction with a pooled screening approach.


For example, a library of gene modifying polypeptide candidates and a template guide RNA (tgRNA) may be introduced into mammalian cells to test the candidates' gene editing abilities by a pooled screening approach. In specific embodiments, a library of gene modifying polypeptide candidates is introduced into mammalian cells followed by introduction of the tgRNA into the cells.


Representative, non-limiting examples of mammalian cells that may be used in screening include HEK293T cells, U2OS cells, HeLa cells, HepG2 cells, Huh? cells, K562 cells, or iPS cells.


A gene modifying polypeptide candidate may comprise 1) a Cas-nuclease, for example a wild-type Cas nuclease, e.g., a wild-type Cas9 nuclease, a mutant Cas nuclease, e.g., a Cas nickase, for example, a Cas9 nickase such as a Cas9 N863A nickase, or a Cas nuclease selected from Table 7 or Table 8, 2) a peptide linker, e.g., a sequence from Table D or Table 10, that may exhibit varying degrees of length, flexibility, hydrophobicity, and/or secondary structure; and 3) a reverse transcriptase (RT), e.g. an RT domain from Table D or Table 6. A gene modifying polypeptide candidate library comprises: a plurality of different gene modifying polypeptide candidates that differ from each other with respect to one, two or all three of the Cas nuclease, peptide linker or RT domain components, or a plurality of nucleic acid expression vectors that encode such gene modifying polypeptide candidates.


For screening of gene modifying polypeptide candidates, a two-component system may be used that comprises a gene modifying polypeptide component and a tgRNA component. A gene modifying component may comprise, for example, an expression vector, e.g., an expression plasmid or lentiviral vector, that encodes a gene modifying polypeptide candidate, for example, comprises a human codon-optimized nucleic acid that encodes a gene modifying polypeptide candidate, e.g., a Cas-linker-RT fusion as described above. In a particular embodiment, a lentiviral cassette is utilized that comprises: (i) a promoter for expression in mammalian cells, e.g., a CMV promoter; (ii) a gene modifying library candidate, e.g. a Cas-linker-RT fusion comprising a Cas nuclease of Table 7 or Table 8, a peptide linker of Table 10, and an RT of Table 6, for example a Cas-linker-RT fusion as in Table D; (iii) a self-cleaving polypeptide, e.g., a T2A peptide; (iv) a marker enabling selection in mammalian cells, e.g., a puromycin resistance gene; and (v) a termination signal, e.g., a poly A tail.


The tgRNA component may comprise a tgRNA or expression vector, e.g., an expression plasmid, that produces the tgRNA, for example, utilizes a U6 promoter to drive expression of the tgRNA, wherein the tgRNA is a non-coding RNA sequence that is recognized by Cas and localizes it to the genomic locus of interest, and that also templates reverse transcription of the desired edit into the genome by the RT domain.


To prepare a pool of cells expressing gene modifying polypeptide library candidates, mammalian cells, e.g., HEK293T or U2OS cells, may be transduced with pooled gene modifying polypeptide candidate expression vector preparations, e.g., lentiviral preparations, of the gene modifying candidate polypeptide library. In a particular embodiment, lentiviral plasmids are utilized, and HEK293 Lenti-X cells are seeded in 15 cm plates (12×106 cells) prior to lentiviral plasmid transfection. In such an embodiment, lentiviral plasmid transfection may be performed using the Lentiviral Packaging Mix (Biosettia) and transfection of the plasmid DNA for the gene modifying candidate library is performed the following day using Lipofectamine 2000 and Opti-MEM media according to the manufacturer's protocol. In such an embodiment, extracellular DNA may be removed by a full media change the next day and virus-containing media may be harvested 48 hours after. Lentiviral media may be concentrated using Lenti-X Concentrator (TaKaRa Biosciences) and 5 mL lentiviral aliquots may be made and stored at −80° C. Lentiviral titering is performed by enumerating colony forming units post-selection, e.g., post Puromycin selection.


For monitoring gene editing of a target DNA, mammalian cells, e.g., HEK293T or U2OS cells, carrying a target DNA may be utilized. In other embodiments for monitoring gene editing of a target DNA, mammalian cells, e.g., HEK293T or U2OS cells, carrying a target DNA genomic landing pad may be utilized. In particular embodiments, the target DNA genomic landing pad may comprise a gene to be edited for treatment of a disease or disorder of interest. In other particular embodiments, the target DNA is a gene sequence that expresses a protein that exhibits detectable characteristics that may be monitored to determine whether gene editing has occurred. For example, in certain embodiments, a blue fluorescence protein (BFP)- or green fluorescence protein (GFP)-expressing genomic landing pad is utilized. In certain embodiments, mammalian cells, e.g., HEK293T or U2OS cells, comprising a target DNA, e.g., a target DNA genomic landing pad, are seeded in culture plates at 500×-3000× cells per gene modifying library candidate and transduced at a 0.2-0.3 multiplicity of infection (MOI) to minimize multiple infections per cell. Puromycin (2.5 ug/mL) may be added 48 hours post infection to allow for selection of infected cells. In such an embodiment, cells may be kept under puromycin selection for at least 7 days and then scaled up for tgRNA introduction, e.g., tgRNA electroporation.


To ascertain whether gene editing occurs, mammalian cells containing a target DNA to be edited may be infected with gene modifying polypeptide library candidates then transfected with tgRNA designed for use in editing of the target DNA. Subsequently, the cells may be analyzed to determine whether editing of the target locus has occurred according to the designed outcome, or whether no editing or imperfect editing has occurred, e.g., by using cell sorting and sequence analysis.


In a particular embodiment, to ascertain whether genome editing occurs, BFP- or GFP-expressing mammalian cells, e.g., HEK293T or U2OS cells, may be infected with gene modifying library candidates and then transfected or electroporated with tgRNA plasmid or RNA, e.g., by electroporation of 250,000 cells/well with 200 ng of a tgRNA plasmid designed to convert BFP-to-GFP or GFP-to-BFP, at a cell count ensuring >250×-1000× coverage per library candidate. In such an embodiment, the genome-editing capacity of the various constructs in this assay may be assessed by sorting the cells by Fluorescence-Activated Cell Sorting (FACS) for expression of the color-converted fluorescent protein (FP) at 4-10 days post-electroporation. Cells are sorted and harvested as distinct populations of unedited cells (exhibiting original florescence protein signal), edited cells (exhibiting converted fluorescence protein signal), and imperfect edit (exhibiting no florescence protein signal) cells. A sample of unsorted cells may also be harvested as the input population to determine candidate enrichment during analysis.


To determine which gene modifying library candidates exhibit genome-editing capacity in an assay, genomic DNA (gDNA) is harvested from the sorted cell populations, and analyzed by sequencing the gene modifying library candidates in each population. Briefly, gene modifying candidates may be amplified from the genome using primers specific to the gene modifying polypeptide expression vector, e.g., the lentiviral cassette, amplified in a second round of PCR to dilute genomic DNA, and then sequenced, for example, sequenced by a next-generation sequencing platform. After quality control of sequencing reads, reads of at least about 1500 nucleotides and generally no more than about 3200 nucleotides are mapped to the gene modifying polypeptide library sequences and those containing a minimum of about an 80% match to a library sequence are considered to be successfully aligned to a given candidate for purposes of this pooled screen. In order to identify candidates capable of performing gene editing in the assay, e.g., the BFP-to-GFP or GFP-to-BFP edit, the read count of each library candidate in the edited population is compared to its read count in the initial, unsorted population.


For purposes of pooled screening, gene modifying candidates with genome-editing capacity are identified based on enrichment in the edited (converted FP) population relative to unsorted (input) cells. In some embodiments, an enrichment of at least 1.0, 1.5, 2.0, 2.5, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, or at least 100-fold over the input indicates potentially useful gene editing activity, e.g., at least 2-fold enrichment. In some embodiments, the enrichment is converted to a log-value by taking the log base 2 of the enrichment ratio. In some embodiments, a log 2 enrichment score of at least 0, 1, 2, 3, 4, 5, 5.5, 6.0, 6.2, 6.3, 6.4, 6.5, or at least 6.6 indicates potentially useful gene editing activity, e.g., a log 2 enrichment score of at least 1.0. In particular embodiments, enrichment values observed for gene modifying candidates may be compared to enrichment values observed under similar conditions utilizing a reference, e.g., Element ID No: 17380.


In some embodiments, multiple tgRNAs may be used to screen the gene modifying candidate library. In particular embodiments, a plurality of tgRNAs may be utilized to optimize template/Cas-linker-RT fusion pairs, e.g., for gene editing of particular target genes, for example, gene targets for the treatment of disease. In specific embodiments, a pooled approach to screening gene modifying candidates may be performed using a multiplicity of different tgRNAs in an arrayed format.


In some embodiments, multiple types of edits, e.g., insertions, substitutions, and/or deletions of different lengths, may be used to screen the gene modifying candidate library.


In some embodiments, multiple target sequences, e.g., different fluorescent proteins, may be used to screen the gene modifying candidate library. In some embodiments, multiple target sequences, e.g., different fluorescent proteins, may be used to screen the gene modifying candidate library. In some embodiments, multiple cell types, e.g., HEK293T or U20S, may be used to screen the gene modifying candidate library. The person of ordinary skill in the art will appreciate that a given candidate may exhibit altered editing capacity or even the gain or loss of any observable or useful activity across different conditions, including tgRNA sequence (e.g., nucleotide modifications, PBS length, RT template length), target sequence, target location, type of edit, location of mutation relative to the first-strand nick of the gene modifying polypeptide, or cell type. Thus, in some embodiments, gene modifying library candidates are screened across multiple parameters, e.g., with at least two distinct tgRNAs in at least two cell types, and gene editing activity is identified by enrichment in any single condition. In other embodiments, a candidate with more robust activity across different tgRNA and cell types is identified by enrichment in at least two conditions, e.g., in all conditions screened. For clarity, candidates found to exhibit little to no enrichment under any given condition are not assumed to be inactive across all conditions and may be screened with different parameters or reconfigured at the polypeptide level, e.g., by swapping, shuffling, or evolving domains (e.g., RT domain), linkers, or other signals (e.g., NLS).


Sequences of Exemplary Cas9-Linker-RT Fusions


In some embodiments, a gene modifying polypeptide comprises a linker sequence and an RT sequence. In some embodiments, a gene modifying polypeptide comprises a linker sequence as listed in Table D, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises the amino acid sequence of an RT domain as listed in Table D, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


In some embodiments, a gene modifying polypeptide comprises a linker sequence as listed in Table D, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto; and the amino acid sequence of an RT domain as listed in Table D, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises: (i) a linker sequence as listed in a row of Table D, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto; and (ii) the amino acid sequence of an RT domain as listed in the same row of Table D, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


Exemplary Gene Modifying Polypeptides

In some embodiments, a gene modifying polypeptide (e.g., a gene modifying polypeptide that is part of a system described herein) comprises an amino acid sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 80% identity thereto. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 90% identity thereto. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 95% identity thereto. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 1-7743. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises an amino acid sequence of any one of SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In some embodiments, a gene modifying polypeptide comprises an amino acid sequence as listed in Table A1, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In some embodiments, a gene modifying polypeptide comprises an amino acid sequence as listed in Table T1, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises a linker comprising a linker sequence as listed in Table T1, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises an RT domain comprising an RT domain sequence as listed in Table T1, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises: (i) a linker comprising a linker sequence as listed in a row of Table T1, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto; and (ii) an RT domain comprising an RT domain sequence as listed in the same row of Table T1, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.









TABLE T1







Selection of exemplary gene modifying polypeptides










SEQ ID NO:





for Full

SEQ ID



Polypeptide

NO: of



Sequence
Linker Sequence
linker
RT name





1372
AEAAAKEAAAKEAAAK
15,401
AVIRE_



EAAAKALEAEAAAKEA

P03360_



AAKEAAAKEAAAKA

3mutA





1197
AEAAAKEAAAKEAAAK
15,402
FLV_



EAAAKALEAEAAAKEA

P10273_



AAKEAAAKEAAAKA

3mutA





2784
AEAAAKEAAAKEAAAK
15,403
MLVMS_



EAAAKALEAEAAAKEA

P03355_



AAKEAAAKEAAAKA

3mutA_WS





 647
AEAAAKEAAAKEAAAK
15,404
SFV3L_



EAAAKALEAEAAAKEA

P27401_



AAKEAAAKEAAAKA

2mutA









In some embodiments, a gene modifying polypeptide comprises an amino acid sequence as listed in Table T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises a linker comprising a linker sequence as listed in Table T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises an RT domain comprising an RT domain sequence as listed in Table T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, a gene modifying polypeptide comprises: (i) a linker comprising a linker sequence as listed in a row of Table T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto; and (ii) an RT domain comprising an RT domain sequence as listed in the same row of Table T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.









TABLE T2







Selection of exemplary gene modifying polypeptides










SEQ ID NO:





for Full

SEQ ID



Polypeptide

NO: of



Sequence
Linker Sequence
linker
RT name





2311
GGGGSGGGGSGGGGSGGGGS
15,405
MLVCB_P08361_3mutA





1373
GGGGGGGGSGGGGSGGGGSGGGGSGGGGS
15,406
AVIRE_P03360_3mutA





2644
GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS
15,407
MLVMS_P03355_PLV919





2304
GSSGSSGSSGSSGSSGSS
15,408
MLVCB_P08361_3mutA





2325
EAAAKEAAAKEAAAKEAAAK
15,409
MLVCB_P08361_3mutA





2322
EAAAKEAAAKEAAAKEAAAKEAAAKEAAAK
15,410
MLVCB_P08361_3mutA





2187
PAPAPAPAPAP
15,411
MLVBM_Q7SVK7_3mut





2309
PAPAPAPAPAPAP
15,412
MLVCB_P08361_3mutA





2534
PAPAPAPAPAPAP
15,413
MLVFF_P26809_3mutA





2797
PAPAPAPAPAPAP
15,414
MLVMS_P03355_3mutA_WS





3084
PAPAPAPAPAPAP
15,415
MLVMS_P03355_3mutA_WS





2868
PAPAPAPAPAPAP
15,416
MLVMS_P03355_PLV919





 126
EAAAKGGG
15,417
PERV_Q4VFZ2_3mut





 306
EAAAKGGG
15,418
PERV_Q4VFZ2_3mut





1410
PAPGGG
15,419
AVIRE_P03360_3mutA





 804
GGGGSSGGS
15,420
WMSV_P03359_3mut





1937
GGGGGSEAAAK
15,421
BAEVM_P10272_3mutA





2721
GGGEAAAKGGS
15,422
MLVMS_P03355_3mut





3018
GGGEAAAKGGS
15,423
MLVMS_P03355_3mut





1018
GGGEAAAKGGS
15,424
XMRV6_A1Z651_3mutA





2317
GGSGGGPAP
15,425
MLVCB_P08361_3mutA





2649
PAPGGSGGG
15,426
MLVMS_P03355_PLV919





2878
PAPGGSGGG
15,427
MLVMS_P03355_PLV919





 912
GGSEAAAKPAP
15,428
WMSV_P03359_3mutA





2338
GGSPAPEAAAK
15,429
MLVCB_P08361_3mutA





2527
GGSPAPEAAAK
15,430
MLVFF_P26809_3mutA





 141
EAAAKGGSPAP
15,431
PERV_Q4VFZ2_3mut





 341
EAAAKGGSPAP
15,432
PERV_Q4VFZ2_3mut





2315
EAAAKPAPGGS
15,433
MLVCB_P08361_3mutA





3080
EAAAKPAPGGS
15,434
MLVMS_P03355_3mutA_WS





2688
GGGGSSEAAAK
15,435
MLVMS_P03355_PLV919





2885
GGGGSSEAAAK
15,436
MLVMS_P03355_PLV919





2810
GSSGGGEAAAK
15,437
MLVMS_P03355_3mutA_WS





3057
GSSGGGEAAAK
15,438
MLVMS_P03355_3mutA_WS





1861
GSSEAAAKGGG
15,439
MLVAV_P03356_3mutA





3056
GSSGGGPAP
15,440
MLVMS_P03355_3mutA_WS





1038
GSSPAPGGG
15,441
XMRV6_A1Z651_3mutA





2308
PAPGGGGSS
15,442
MLVCB_P08361_3mutA





1672
GGGEAAAKPAP
15,443
KORV_Q9TTC1-Pro_3mutA





2526
GGGEAAAKPAP
15,444
MLVFF_P26809_3mutA





1938
GGGPAPEAAAK
15,445
BAEVM_P10272_3mutA





2641
GSSEAAAKPAP
15,446
MLVMS_P03355_PLV919





2891
GSSEAAAKPAP
15,447
MLVMS_P03355_PLV919





1225
GSSPAPEAAAK
15,448
FLV_P10273_3mutA





2839
GSSPAPEAAAK
15,449
MLVMS_P03355_3mutA_WS





3127
GSSPAPEAAAK
15,450
MLVMS_P03355_3mutA_WS





2798
PAPGSSEAAAK
15,451
MLVMS_P03355_3mutA_WS





3091
PAPGSSEAAAK
15,452
MLVMS_P03355_3mutA_WS





1372
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,453
AVIRE_P03360_3mutA



AKEAAAKEAAAKA







1197
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,454
FLV_P10273_3mutA



AKEAAAKEAAAKA







2611
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,455
MLVMS_P03355_PLV919



AKEAAAKEAAAKA







2784
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,456
MLVMS_P03355_3mutA_WS



AKEAAAKEAAAKA







 480
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,457
SFV1_P23074_2mutA



AKEAAAKEAAAKA







 647
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,458
SFV3L_P27401_2mutA



AKEAAAKEAAAKA







1006
AEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAA
15,459
XMRV6_A1Z651_3mutA



AKEAAAKEAAAKA







2518
SGSETPGTSESATPES
15,460
MLVFF_P26809_3mutA









Subsequences of Exemplary Gene Modifying Polypeptides

In some embodiments, the gene modifying polypeptide comprises, in N-terminal to C-terminal order, one or more (e.g., 1, 2, 3, 4, 5, or all 6) of an N-terminal methionine residue, a first nuclear localization signal (NLS), a DNA binding domain, a linker, an RT domain, and/or a second NLS. In some embodiments, a gene modifying polypeptide comprises, in N-terminal to C-terminal order, a NLS (e.g., a first NLS), a DNA binding domain, a linker, and an RT domain, wherein the linker and RT domain are the linker and RT domain of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said linker and RT domain. In some embodiments, a gene modifying polypeptide comprises, in N-terminal to C-terminal order, a DNA binding domain, a linker, an RT domain, and an NLS (e.g., a second NLS) wherein the linker and RT domain are the linker and RT domain of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said linker and RT domain. In some embodiments, a gene modifying polypeptide comprises, in N-terminal to C-terminal order, a first NLS, a DNA binding domain, a linker, an RT domain, and a second NLS, wherein the linker and RT domain are the linker and RT domain of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said linker and RT domain. In some embodiments, the gene modifying polypeptide further comprises an N-terminal methionine residue.


In some embodiments, the gene modifying polypeptide comprises, in N-terminal to C-terminal order, one or more (e.g., 1, 2, 3, 4, 5, or all 6) of an N-terminal methionine residue, a first nuclear localization signal (NLS) (e.g., of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743 and/or as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto), a DNA binding domain (e.g., a Cas domain, e.g., a SpyCas9 domain, e.g., as listed in Table 8, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto; or a DNA binding domain of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743 and/or as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto), a linker (e.g., of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743 and/or as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto), an RT domain (e.g., of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743 and/or as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto), and a second NLS (e.g., of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743 and/or as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto). In some embodiments, the gene modifying polypeptide further comprises (e.g., C-terminal to the second NLS) a T2A sequence and/or a puromycin sequence (e.g., of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743 and/or as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto). In some embodiments, a nucleic acid encoding a gene modifying polypeptide (e.g., as described herein) encodes a T2A sequence, e.g., wherein the T2A sequence is situated between a region encoding the gene modifying polypeptide and a second region, wherein the second region optionally encodes a selectable marker, e.g., puromycin.


In certain embodiments, the first NLS comprises a first NLS sequence of a gene modifying polypeptide having an amino acid sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the first NLS comprises a first NLS sequence of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the first NLS sequence comprises a C-myc NLS. In certain embodiments, the first NLS comprises the amino acid sequence PAAKRVKLD (SEQ ID NO: 11,095), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the gene modifying polypeptide further comprises a spacer sequence between the first NLS and the DNA binding domain. In certain embodiments, the spacer sequence between the first NLS and the DNA binding domain comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In certain embodiments, the spacer sequence between the first NLS and the DNA binding domain comprises the amino acid sequence GG.


In certain embodiments, the DNA binding domain comprises a DNA binding domain of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the DNA binding domain comprises a DNA binding domain of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the DNA binding domain comprises a Cas domain (e.g., as listed in Table 8). In certain embodiments, the DNA binding domain comprises the amino acid sequence of a SpyCas9 polypeptide (e.g., as listed in Table 8, e.g., a Cas9 N863A polypeptide), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the DNA binding domain comprises the amino acid sequence:









(SEQ ID NO: 11,096)


DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIG





ALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSF





FHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDS





TDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYN





QLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNL





IALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADL





FLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKA





LVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDG





TEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFL





KDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEV





VDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKY





VTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS





VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTL





FEDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDK





QSGKTILDFLKSDGFANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLH





EHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPENIVIEMARENQT





TQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQ





NGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARG





KSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDK





AGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSK





LVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPKLESEFV





YGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI





RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGF





SKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGK





SKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKY





SLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSP





EDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKH





RDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDA





TLIHQSITGLYETRIDLSQLGGD,







or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the gene modifying polypeptide further comprises a spacer sequence between the DNA binding domain and the linker. In certain embodiments, the spacer sequence between the DNA binding domain and the linker comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In certain embodiments, the spacer sequence between the DNA binding domain and the linker comprises the amino acid sequence GG.


In certain embodiments, the linker comprises a linker sequence of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the linker comprises a linker sequence of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the linker comprises an amino acid sequence as listed in Table D or 10, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the gene modifying polypeptide further comprises a spacer sequence between the linker and the RT domain. In certain embodiments, the spacer sequence between the linker and the RT domain comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In certain embodiments, the spacer sequence between the linker and the RT domain comprises the amino acid sequence GG.


In certain embodiments, the RT domain comprises a RT domain sequence of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the RT domain comprises a RT domain sequence of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the RT domain comprises an amino acid sequence as listed in Table D or 6, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain has a length of about 400-500, 500-600, 600-700, 700-800, 800-900, or 900-1000 amino acids.


In certain embodiments, the gene modifying polypeptide further comprises a spacer sequence between the RT domain and the second NLS. In certain embodiments, the spacer sequence between the RT domain and the second NLS comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In certain embodiments, the spacer sequence between the RT domain and the second NLS comprises the amino acid sequence AG.


In certain embodiments, the second NLS comprises a second NLS sequence of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743. In certain embodiments, the second NLS comprises a second NLS sequence of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2. In certain embodiments, the second NLS sequence comprises a plurality of partial NLS sequences. In embodiments, the NLS sequence, e.g., the second NLS sequence, comprises a first partial NLS sequence, e.g., comprising the amino acid sequence KRTADGSEFE (SEQ ID NO: 11,097), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In embodiments, the NLS sequence, e.g., the second NLS sequence, comprises a second partial NLS sequence. In embodiments, the NLS sequence, e.g., the second NLS sequence, comprises an SV40A5 NLS, e.g., a bipartite SV40A5 NLS, e.g., comprising the amino acid sequence KRTADGSEFESPKKKAKVE (SEQ ID NO: 11,098), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the NLS sequence, e.g., the second NLS sequence, comprises the amino acid sequence KRTADGSEFEKRTADGSEFESPKKKAKVE (SEQ ID NO: 11,099), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the gene modifying polypeptide further comprises a spacer sequence between the second NLS and the T2A sequence and/or puromycin sequence. In certain embodiments, the spacer sequence between the second NLS and the T2A sequence and/or puromycin sequence comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In certain embodiments, the spacer sequence between the second NLS and the T2A sequence and/or puromycin sequence comprises the amino acid sequence GSG.


Linkers and RT Domains

In some embodiments, the gene modifying polypeptide comprises a linker (e.g., as described herein) and an RT domain (e.g., as described herein). In certain embodiments, the gene modifying polypeptide comprises, in N-terminal to C-terminal order, a linker (e.g., as described herein) and an RT domain (e.g., as described herein).


In certain embodiments, the linker comprises a linker sequence as listed in Table 10, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the linker comprises a linker sequence of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the linker comprises a linker sequence of any one of SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the linker comprises a linker sequence of any one of SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the linker comprises a linker sequence of an exemplary gene modifying polypeptide listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the RT domain comprises an RT domain sequence as listed in Table 6, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the RT domain comprises an RT domain sequence of an exemplary gene modifying polypeptide listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In some embodiments, a gene modifying polypeptide comprises a portion of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion.


In some embodiments, a gene modifying polypeptide comprises a linker of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said linker. In some embodiments, a gene modifying polypeptide comprises a linker of a gene modifying polypeptide of any one of SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said linker. In some embodiments, a gene modifying polypeptide comprises a linker of a gene modifying polypeptide of any one of SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said linker. In some embodiments, a gene modifying polypeptide comprises a linker of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2, or a linker comprising an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In some embodiments, a gene modifying polypeptide comprises an RT domain of a gene modifying polypeptide of any one of SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said RT domain. In some embodiments, a gene modifying polypeptide comprises an RT domain of a gene modifying polypeptide of any one of SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity said RT domain. In some embodiments, a gene modifying polypeptide comprises an RT domain of a gene modifying polypeptide of any one of SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity said RT domain. In some embodiments, a gene modifying polypeptide comprises an RT domain of a gene modifying polypeptide as listed in any of Tables A1, T1, or T2, or an RT domain comprising an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise the amino acid sequences of a linker and RT domain (or amino acid sequences having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto) of a gene modifying polypeptide having the amino acid sequence of any one of SEQ ID NOs: 1-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise amino acid sequences of a linker and RT domain having at least 80% identity to the linker and RT domains of any one of SEQ ID NOs: 1-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise amino acid sequences of a linker and RT domain having at least 90% identity to the linker and RT domains of any one of SEQ ID NOs: 1-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise amino acid sequences of a linker and RT domain having at least 95% identity to the linker and RT domains of any one of SEQ ID NOs: 1-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise amino acid sequences of a linker and RT domain having at least 99% identity to the linker and RT domains of any one of SEQ ID NOs: 1-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise the amino acid sequences of a linker and RT domain (or amino acid sequences having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto) of a gene modifying polypeptide having the amino acid sequence of any one of SEQ ID NOs: 6001-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise the amino acid sequences of a linker and RT domain (or amino acid sequences having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto) of a gene modifying polypeptide having the amino acid sequence of any one of SEQ ID NOs: 4501-4541. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise the amino acid sequences of a linker and RT domain (or amino acid sequences having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto) from a single row of any of Tables A1, T1, or T2 (e.g., from a single exemplary gene modifying polypeptide as listed in any of Tables A1, T1, or T2).


In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise the amino acid sequences of a linker and RT domain (or amino acid sequences having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto) from two different amino acid sequences selected from SEQ ID NOs: 1-7743. In certain embodiments, the linker and the RT domain of a gene modifying polypeptide comprise the amino acid sequences of a linker and RT domain (or amino acid sequences having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto) from different rows of any of Tables A1, T1, or T2.


In certain embodiments, the gene modifying polypeptide further comprises a first NLS (e.g., a 5′ NLS), e.g., as described herein. In certain embodiments, the gene modifying polypeptide further comprises a second NLS (e.g., a 3′ NLS), e.g., as described herein. In certain embodiments, the gene modifying polypeptide further comprises an N-terminal methionine residue.


RT Families and Mutants In certain embodiments, a gene modifying polypeptide comprises the amino acid sequence of an RT domain sequence from a family selected from: AVIRE, BAEVM, FFV, FLY, FOAMY, GALV, KORV, MLVAV, MLVBM, MLVCB, MLVFF, MLVMS, PERV, SFV1, SFV3L, WMSV, XMRV6, BLVAU, BLVJ, HTL1A, HTL1C, HTL1L, HTL32, HTL3P, HTLV2, JSRV, MLVFS, MLVRD, MMTVB, MPMV, SFVCP, SMRVH, SRV1, SRV2, and WDSV. In certain embodiments, a gene modifying polypeptide comprises the amino acid sequence of an RT domain sequence from a family selected from: AVIRE, BAEVM, FFV, FLY, FOAMY, GALV, KORV, MLVAV, MLVBM, MLVCB, MLVFF, MLVMS, PERV, SFV1, SFV3L, WMSV, and XMRV6.


In certain embodiments, a gene modifying polypeptide comprises comprises the amino acid sequence of an RT domain sequence from an MLVMS RT domain. In embodiments, the amino acid sequence of an RT domain sequence comprises one or more point mutations as listed in column 1 of Table M1, or a point mutation corresponding thereto. In embodiments, the amino acid sequence of an RT domain sequence comprises one or more point mutations as listed in column 3 of Table M1 (Gen1 MLVMS), or a point mutation corresponding thereto. In embodiments, the amino acid sequence of an RT domain sequence comprises one or more point mutations at an amino acid position of the RT domain as listed in columns 1 and 2 of Table M2, or an amino acid position corresponding thereto.


In certain embodiments, a gene modifying polypeptide comprises the amino acid sequence of an RT domain sequence from an AVIRE RT domain. In embodiments, the amino acid sequence of an RT domain sequence comprises one or more point mutations as listed in column 2 of Table M1, or a point mutation corresponding thereto. In embodiments, the amino acid sequence of an RT domain sequence comprises one or more point mutations as listed in column 4 of Table M1 (Gen2 AVIRE), or a point mutation corresponding thereto. In embodiments, the amino acid sequence of an RT domain sequence comprises one or more point mutations at an amino acid position of the RT domain as listed in columns 3 and 4 of Table M2, or an amino acid position corresponding thereto. In certain embodiments, the RT domain comprises an IENSSP (e.g., at the C-terminus).









TABLE M1







Exemplary point mutations in MLVMS and


AVIRE RT domains










RT-linker

Gen1
Gen2


filing
Corresponding
MLVMS
AVIRE


(MLVMS)
AVIRE
(PLV4921)
(PLV10990)







H8Y



P51L
Q51L




S67R
T67R




E67K
E67K




E69K
E69K




T197A
T197A




D200N
D200N
D200N
D200N


H204R
N204R




E302K
E302K






T306K
T306K


F309N
Y309N




W313F
W313F
W313F
W313F


T330P
G330P
T330P
G330P


L435G
T436G




N454K
N455K




D524G
D526G




E562Q
E564Q




D583N
D585N




H594Q
H596Q




L603W
L605W
L603W
L605W


D653N
D655N




L671P
L673P













IENSSP at C-term
















TABLE M2







Positions that can be mutated in exemplary


MLVMS and AVIRE RT domains


WT residue & position













MLVMS

AVIRE



MLVMS
position
AVIRE
position



aa
# *
aa
# *
















H
8
Y
8



P
51
Q
51



S
67
T
67



E
69
E
69



T
197
T
197



D
200
D
200



H
204
N
204



E
302
E
302



T
306
T
306



F
309
Y
309



W
313
W
313



T
330
G
330



L
435
T
436



N
454
N
455



D
524
D
526



E
562
E
564



D
583
D
585



H
594
H
596



L
603
L
605



D
653
D
655



L
671
S
673










In certain embodiments, a gene modifying polypeptide comprises a gamma retrovirus derived RT domain. In certain embodiments, the gamma retrovirus-derived RT domain of a gene modifying polypeptide comprises the amino acid sequence of an RT domain sequence from a family selected from: AVIRE, BAEVM, FFV, FLY, FOAMY, GALV, KORV, MLVAV,


MLVBM, MLVCB, MLVFF, MLVMS, PERV, SFV1, SFV3L, WMSV, and XMRV6. In some embodiments, the gamma retrovirus-derived RT domain of a gene modifying polypeptide is not derived from PERV. In some embodiments, said RT includes one, two, three, four, five, six or more mutations shown in Table 2A and corresponding to mutations D200N, L603W, T330P, D524G, E562Q, D583N, P51L, S67R, E67K, T197A, H204R, E302K, F309N, W313F, L435G, N454K, H594Q, L671P, E69K, or D653N in the RT domain of murine leukemia virus reverse transcriptase. In some embodiments, the gene modifying polypeptide further comprises a linker having at least 99% identity to a linker domains of any one of SEQ ID NOs: 1-7743. In some embodiments, the gene modifying polypeptide further comprises a linker having at least 99% or 100% identity to SEQ ID NO: 5217 or SEQ ID NO:11,041.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of an AVIRE RT (e.g., an AVIRE P03360 sequence, e.g., SEQ ID NO: 8001), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of an AVIRE RT further comprising one, two, three, four, or five mutations selected from the group consisting of D200N, G330P, L605W, T306K, and W313F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an AVIRE RT further comprising one, two, or three mutations selected from the group consisting of D200N, G330P, and L605W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a BAEVM RT (e.g., an BAEVM_P10272 sequence, e.g., SEQ ID NO: 8004), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a BAEVM RT further comprising one, two, three, four, or five mutations selected from the group consisting of D198N, E328P, L602W, T304K, and W311F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a BAEVM RT further comprising one, two, or three mutations selected from the group consisting of D198N, E328P, and L602W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of an FFV RT (e.g., an FFV 093209 sequence, e.g., SEQ ID NO: 8012), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of an FFV RT further comprising one, two, three, or four mutations selected from the group consisting of D21N, T293N, T419P, and L393K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FFV RT further comprising one, two, or three mutations selected from the group consisting of D21N, T293N, and T419P, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FFV RT further comprising the mutation D21N. In some embodiments, the RT domain comprises the amino acid sequence of an FFV RT further comprising one, two, or three mutations selected from the group consisting of T207N, T333P, and L307K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FFV RT further comprising one or two mutations selected from the group consisting of T207N and T333P, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of an FLV RT (e.g., an FLV P10273 sequence, e.g., SEQ ID NO: 8019), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of an FLV RT further comprising one, two, three, or four mutations selected from the group consisting of D199N, L602W, T305K, and W312F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FLV RT further comprising one or two mutations selected from the group consisting of D199N and L602W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a FOAMV RT (e.g., an FOAMV P14350 sequence, e.g., SEQ ID NO: 8021), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of an FOAMV RT further comprising one, two, three, or four mutations selected from the group consisting of D24N, T296N, S420P, and L396K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FOAMV RT further comprising one, two, or three mutations selected from the group consisting of D24N, T296N, and S420P, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FOAMV RT further comprising the mutation D24N, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FOAMV RT further comprising one, two, or three mutations selected from the group consisting of T207N, S331P, and L307K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of an FOAMV RT further comprising one or two mutations selected from the group consisting of T207N and S331P, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a GALV RT (e.g., an GALV P21414 sequence, e.g., SEQ ID NO: 8027), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a GALV RT further comprising one, two, three, four, or five mutations selected from the group consisting of D198N, E328P, L600W, T304K, and W311F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a GALV RT further comprising one, two, or three mutations selected from the group consisting of D198N, E328P, and L600W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a KORV RT (e.g., an KORV_Q9TTC1 sequence, e.g., SEQ ID NO: 8047), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a GALV RT further comprising one, two, three, four, five, or six mutations selected from the group consisting of D32N, D322N, E452P, L274W, T428K, and W435F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a GALV RT further comprising one, two, three, or four mutations selected from the group consisting of D32N, D322N, E452P, and L274W, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a GALV RT further comprising the mutation D32N. In some embodiments, the RT domain comprises the amino acid sequence of a KORV RT further comprising one, two, three, four, or five mutations selected from the group consisting of D231N, E361P, L633W, T337K, and W344F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a KORV RT further comprising one, two, or three mutations selected from the group consisting of D231N, E361P, and L633W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a MLVAV RT (e.g., an MLVAV_P03356 sequence, e.g., SEQ ID NO: 8053), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a MLVAV RT further comprising one, two, three, four, or five mutations selected from the group consisting of D200N, T330P, L603W, T306K, and W313F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a MLVAV RT further comprising one, two, or three mutations selected from the group consisting of D200N, T330P, and L603W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a MLVBM RT (e.g., an MLVBM_Q7SVK7 sequence, e.g., SEQ ID NO: 8056), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a MLVBM RT further comprising one, two, three, four, or five mutations selected from the group consisting of D199N, T329P, L602W, T305K, and W312F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a MLVBM RT further comprising one, two, and three mutations selected from the group consisting of D200N, T330P, and L603W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a MLVCB RT (e.g., an MLVCB_P08361 sequence, e.g., SEQ ID NO: 8062), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a MLVCB RT further comprising one, two, three, four, or five mutations selected from the group consisting of D200N, T330P, L603W, T306K, and W313F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a MLVCB RT further comprising one, two, and three mutations selected from the group consisting of D200N, T330P, and L603W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a MLVFF RT, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a MLVFF RT further comprising one, two, three, four, or five mutations selected from the group consisting of D200N, T330P, L603W, T306K, and W313F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a MLVFF RT further comprising one, two, and three mutations selected from the group consisting of D200N, T330P, and L603W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a MLVMS RT (e.g., an MLVMS reference sequence, e.g., SEQ ID NO: 8137; or an MLVMS P03355 sequence, e.g., SEQ ID NO: 8070), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a MLVMS RT further comprising one, two, three, four, five, or six mutations selected from the group consisting of D200N, T330P, L603W, T306K, W313F, and H8Y, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a MLVMS RT further comprising one, two, three, four, or five mutations selected from the group consisting of D200N, T330P, L603W, T306K, and W313F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a MLVMS RT further comprising one, two, or three mutations selected from the group consisting of D200N, T330P, and L603W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a PERV RT (e.g., an PERV Q4VFZ2 sequence, e.g., SEQ ID NO: 8099), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a PERV RT further comprising one, two, three, four, or five mutations selected from the group consisting of D196N, E326P, L599W, T302K, and W309F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a PERV RT further comprising one, two, or three mutations selected from the group consisting of D196N, E326P, and L599W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a SFV1 RT (e.g., an SFV1_P23074 sequence, e.g., SEQ ID NO: 8105), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a SFV1 RT further comprising one, two, three, or four mutations selected from the group consisting of D24N, T296N, N420P, and L396K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a SFV1 RT further comprising one, two, or three mutations selected from the group consisting of D24N, T296N, and N420P, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a SFV1 RT further comprising the D24N, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a SFV3L RT (e.g., an SFV3L P27401 sequence, e.g., SEQ ID NO: 8111), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a SFV3L RT further comprising one, two, three, or four mutations selected from the group consisting of D24N, T296N, N422P, and L396K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a SFV3L RT further comprising one, two, or three mutations selected from the group consisting of D24N, T296N, and N422P, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a SFV3L RT further comprising the mutation D24N, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a SFV3L RT further comprising one, two, or three mutations selected from the group consisting of T307N, N333P, and L307K, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a SFV3L RT further comprising one or two mutations selected from the group consisting of T307N and N333P, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a WMSV RT (e.g., an WMSV P03359 sequence, e.g., SEQ ID NO: 8131), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a WMSV RT further comprising one, two, three, four, or five mutations selected from the group consisting of D198N, E328P, L600W, T304K, and W311F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a WMSV RT further comprising one, two, or three mutations selected from the group consisting of D198N, E328P, and L600W, or a corresponding position in a homologous RT domain.


In embodiments, the RT domain comprises the amino acid sequence of an RT domain of a XMRV6 RT (e.g., an XMRV6_A1Z651 sequence, e.g., SEQ ID NO: 8134), or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the RT domain comprises the amino acid sequence of a XMRV6 RT further comprising one, two, three, four, or five mutations selected from the group consisting of D200N, T330P, L603W, T306K, and W313F, or a corresponding position in a homologous RT domain. In some embodiments, the RT domain comprises the amino acid sequence of a XMRV6 RT further comprising one, two, or three mutations selected from the group consisting of D200N, T330P, and L603W, or a corresponding position in a homologous RT domain.


In certain embodiments, the RT domain of a gene modifying polypeptide comprises the amino acid sequence of an RT domain of an AVIRE RT, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In embodiments, the RT domain comprises the amino acid sequence of an RT domain comprised in a sequence listed in column 1 of Table A5, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the gene modifying polypeptide further comprises a linker having at least 99% or 100% identity to SEQ ID NO: 5217 or SEQ ID NO:11,041.


In certain embodiments, the RT domain of a gene modifying polypeptide comprises the amino acid sequence of an RT domain of an MLVMS RT, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In embodiments, the RT domain comprises the amino acid sequence of an RT domain comprised in a sequence listed in any of columns 2-6 of Table A5, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the gene modifying polypeptide further comprises a linker having at least 99% or 100% identity to SEQ ID NO: 5217 or SEQ ID NO:11,041.









TABLE A5







Exemplary gene modifying polypeptides comprising


an AVIRE RT domain or an MLVMS RT domain.














AVIRE SEQ















ID NOS:
MLVMS SEQ ID NOS:


















  1
2704
3007
3038
2638
2930



  2
2706
3007
3038
2639
2930



  3
2708
3008
3039
2639
2931



  4
2709
3008
3039
2640
2931



  5
2709
3009
3040
2640
2932



  6
2710
3010
3040
2641
2932



  7
2957
3010
3041
2641
2933



  9
2957
3011
3041
2642
2933



 10
2958
3012
3042
2642
2934



 12
2959
3012
3042
2643
2934



 13
2960
3013
3043
2643
2935



 14
2962
3013
3043
2644
2935



6076
6042
3014
3044
2644
2936



6143
6068
3014
3044
2645
2936



6200
6097
3015
3045
2645
2937



6254
6136
3015
3045
2646
2937



6274
6156
3016
3046
2646
2938



6315
6215
3016
3046
2647
2938



6328
6216
3017
3047
2647
2939



6337
6301
3018
3047
2648
2939



6403
6352
3018
3048
2648
2940



6420
6365
3019
3048
2649
2940



6440
6411
3019
3049
2649
2941



6513
6436
3020
3049
2650
2941



6552
6458
3020
3050
2650
2942



6613
6459
3021
3051
2651
2942



6671
6524
3021
3051
2651
2943



6822
6562
3022
3052
2652
2943



6840
6563
3023
3052
2652
2944



6884
6699
3023
3053
2653
2945



6907
6865
3024
3053
2653
2945



6970
7022
3024
3054
2654
2946



7025
7037
3025
3054
2655
2946



7052
7088
3025
3055
2655
2947



7078
7116
3026
3055
2656
2947



7243
7175
3026
3056
2656
2948



7253
7200
3027
3056
2657
2948



7318
7206
3027
3057
2657
2949



7379
7277
3028
3057
2658
2949



7486
7294
3028
3058
2658
2950



7524
7330
3029
3058
2659
2950



7668
7411
3030
3059
2659
2951



7680
7455
3030
3059
2660
2951



7720
7477
3031
3060
2660
2952



1137
7511
3031
3060
2661
2952



1138
7538
3032
3061
2661
2953



1139
7559
3032
3061
2662
2953



1140
7560
3033
3062
2662
2954



1141
7593
3033
3062
2663
2954



1142
7594
3034
3063
2663
2955



1143
7607
3034
3063
2664
2955



1144
7623
6025
3064
2664
6485



1145
7638
6041
3064
2665
6486



1146
7717
6043
3065
2665
6504



1147
7731
6098
3065
2666
6505



1148
7732
6099
3066
2666
6595



1149
2711
6180
3066
2667
6596



1150
2711
6182
3067
2667
6751



1151
2712
6237
3067
2668
6752



1152
2712
6238
3068
2668
6777



1153
2713
6311
3068
2669
6778



1154
2713
6312
3069
2669
7172



1155
2714
6578
3069
2670
7174



1156
2714
6579
3070
2670
7313



1157
2715
6663
3070
2671
7314



1158
2715
6664
3071
2671




1159
2716
6708
3071
2672




1160
2716
6709
3072
2672




1161
2717
6809
3072
2673




1162
2717
6831
3073
2673




1163
2718
6832
3073
2674




1164
2718
6864
3074
2674




1165
2719
6866
3074
2675




1166
2719
7089
3075
2675




1167
2720
7157
3075
2676




6015
2720
7159
3076
2676




6029
2721
7173
3076
2677




6045
2721
7176
3077
2677




6077
2722
7293
3077
2678




6129
2722
7295
3078
2678




6144
2723
7343
3078
2679




6164
2723
7393
3079
2680




6201
2724
7394
3079
2680




6227
2724
7425
3080
2681




6244
2725
7426
3080
2681




6250
2725
7444
3081
2682




6264
2726
7445
3081
2682




6289
2726
7476
3082
2683




6304
2727
7478
3082
2683




6316
2727
7496
3083
2684




6384
2728
7497
3083
2684




6421
2728
7537
3084
2685




6441
2729
7539
3084
2685




6492
2729
2780
3085
2686




6514
2730
2780
3085
2686




6530
2730
2781
3086
2687




6569
2731
2781
3086
2687




6584
2731
2782
3087
2688




6621
2732
2782
3087
2688




6651
2732
2783
3088
2689




6659
2733
2783
3088
2689




6683
2734
2784
3089
2690




6703
2734
2784
3089
2690




6727
2735
2785
3090
2691




6732
2735
2785
3090
2692




6745
2736
2786
3091
2692




6755
2736
2786
3091
2693




6784
2737
2787
3092
2693




6817
2737
2787
3092
2694




6823
2738
2788
3093
2694




6841
2739
2788
3093
2695




6871
2740
2789
3094
2695




6885
2740
2789
3095
2696




6898
2741
2790
3095
2696




6908
2741
2790
3096
2697




6933
2742
2791
3096
2697




6971
2742
2791
3097
2698




7009
2743
2792
3097
2698




7018
2743
2792
3098
2699




7045
2744
2793
3098
2699




7053
2744
2793
3099
2700




7068
2745
2794
3099
2700




7079
2745
2794
3100
2701




7096
2746
2795
3100
2701




7104
2746
2795
3101
2702




7122
2747
2796
3101
2702




7151
2747
2796
3102
2703




7163
2748
2797
3102
2703




7181
2748
2797
3103
2862




7244
2749
2798
3103
2862




7273
2750
2798
3104
2863




7319
2750
2799
3104
2863




7336
2751
2799
3105
2864




7380
2751
2800
3105
2864




7402
2752
2800
3106
2865




7462
2752
2801
3106
2865




7487
2753
2801
3107
2866




7525
2753
2802
3107
2866




7569
2754
2802
3108
2867




7626
2754
2803
3108
2867




7689
2755
2803
3109
2868




7707
2755
2804
3109
2868




7721
2756
2804
3110
2869




1371
2756
2805
3110
2869




1372
2757
2805
3111
2870




1373
2758
2806
3111
2870




1374
2758
2806
3112
2871




1375
2759
2807
3112
2871




1376
2759
2807
3113
2872




1377
2760
2808
3113
2872




1378
2760
2808
3114
2873




1379
2761
2809
3114
2873




1380
2761
2809
3115
2874




1381
2762
2810
3115
2874




1382
2762
2810
3116
2875




1383
2763
2811
3116
2875




1384
2763
2811
3117
2876




1385
2764
2812
3117
2876




1386
2764
2812
3118
2877




1387
2765
2813
3118
2877




1388
2765
2813
3119
2878




1389
2766
2814
3119
2878




1390
2766
2814
3120
2879




1391
2767
2815
3120
2879




1392
2767
2815
3121
2880




1393
2768
2816
3121
2880




1394
2768
2816
3122
2881




1395
2769
2817
3122
2881




1396
2769
2817
3123
2882




1397
2770
2818
3123
2882




1398
2770
2818
3124
2883




1399
2771
2819
3124
2883




1400
2771
2819
3125
2884




1401
2772
2820
3125
2884




1402
2773
2820
3126
2885




1403
2773
2821
3126
2885




1404
2774
2821
3127
2886




1405
2774
2822
3127
2886




1406
2775
2822
3128
2887




1407
2775
2823
3128
2887




1408
2776
2823
3129
2888




1409
2776
2824
3129
2888




1410
2777
2824
3130
2889




1411
2777
2825
3130
2889




1412
2778
2825
3131
2890




1413
2779
2826
3131
2890




1414
2779
2826
3132
2891




1415
2965
2827
3133
2891




1416
2965
2827
3133
2892




1417
2966
2828
3134
2893




1418
2966
2828
3134
2893




1419
2967
2829
3135
2894




1420
2968
2829
3135
2894




1421
2968
2830
3136
2895




1422
2969
2830
3136
2895




1423
2969
2831
6181
2896




1424
2970
2831
6183
2896




1425
2970
2832
6284
2897




1426
2971
2832
6285
2897




1427
2971
2833
6760
2898




1428
2972
2833
6761
2898




1429
2972
2834
7036
2899




1430
2973
2834
7038
2899




1431
2974
2835
7158
2900




1432
2974
2835
7160
2900




1433
2975
2836
2610
2901




1434
2976
2836
2610
2901




1435
2976
2837
2611
2902




1436
2977
2837
2611
2902




1437
2977
2838
2612
2903




1439
2978
2838
2612
2903




1440
2978
2839
2613
2904




1441
2979
2839
2613
2904




1442
2979
2840
2614
2905




1443
2980
2840
2614
2905




1444
2980
2841
2615
2906




1445
2981
2841
2615
2906




1446
2981
2842
2616
2907




1447
2982
2842
2616
2907




6001
2982
2843
2617
2908




6030
2983
2843
2617
2908




6078
2983
2844
2618
2909




6108
2984
2844
2618
2909




6130
2985
2845
2619
2910




6165
2985
2845
2619
2910




6265
2986
2846
2620
2911




6275
2987
2846
2620
2911




6305
2987
2847
2621
2912




6329
2988
2847
2621
2912




6370
2988
2848
2622
2913




6385
2989
2848
2622
2913




6404
2989
2849
2623
2914




6531
2990
2849
2623
2914




6585
2990
2850
2624
2915




6622
2991
2850
2624
2915




6652
2991
2851
2625
2916




6733
2992
2851
2625
2916




6756
2992
2852
2626
2917




6765
2993
2852
2626
2917




6798
2993
2853
2627
2918




6824
2994
2853
2627
2919




6972
2994
2854
2628
2919




7046
2995
2854
2628
2920




7054
2995
2855
2629
2920




7069
2996
2855
2629
2921




7080
2996
2856
2630
2921




7105
2997
2856
2630
2922




7123
2998
2857
2631
2922




7143
2998
2857
2631
2923




7152
2999
2858
2632
2923




7204
2999
2858
2632
2924




7320
3001
2859
2633
2924




7351
3001
2859
2633
2925




7381
3002
2860
2634
2925




7403
3002
2860
2634
2926




7438
3003
2861
2635
2926




7488
3003
2861
2635
2927




7500
3004
3035
2636
2927




7526
3004
3036
2636
2928




7588
3005
3036
2637
2928




7612
3005
3037
2637
2929




7627
3006
3037
2638
2929










Systems

In an aspect, the disclosure relates to a system comprising nucleic acid molecule encoding a gene modifying polypeptide (e.g., as described herein) and a template nucleic acid (e.g., a template RNA, e.g., as described herein). In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises one or more silent mutations in the coding region (e.g., in the sequence encoding the RT domain) relative to a nucleic acid molecule as described herein. In certain embodiments, the system further comprises a gRNA (e.g., a gRNA that binds to a polypeptide that induces a nick, e.g., in the opposite strand of the target DNA bound by the gene modifying polypeptide).


In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide encodes a polypeptide having an amino acid sequence selected from SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide encodes a polypeptide having an amino acid sequence selected from SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide encodes a polypeptide having an amino acid sequence selected from SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide encodes a polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding a portion of an amino acid sequence selected from SEQ ID NOs: 1-7743, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding a portion of an amino acid sequence selected from SEQ ID NOs: 6001-7743, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding a portion of an amino acid sequence selected from SEQ ID NOs: 4501-4541, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding a portion of a polypeptide listed in any of Tables A1, T1, or T2, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion.


In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the linker of an amino acid sequence selected from SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the linker of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the linker of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the linker of a polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the RT domain of an amino acid sequence selected from SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the RT domain of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the RT domain of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the nucleic acid molecule encoding the gene modifying polypeptide comprises a sequence encoding the RT domain of a polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In an aspect, the disclosure relates to a system comprising a gene modifying polypeptide (e.g., as described herein) and a template nucleic acid (e.g., a template RNA, e.g., as described herein).


In certain embodiments, the gene modifying polypeptide comprises a polypeptide having an amino acid sequence selected from SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a polypeptide having an amino acid sequence selected from SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a polypeptide having an amino acid sequence selected from SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the gene modifying polypeptide comprises a portion of an amino acid sequence selected from SEQ ID NOs: 1-7743, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion. In certain embodiments, the gene modifying polypeptide comprises a portion of an amino acid sequence selected from SEQ ID NOs: 6001-7743, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion. In certain embodiments, the gene modifying polypeptide comprises a portion of an amino acid sequence selected from SEQ ID NOs: 4501-4541, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion. In certain embodiments, the gene modifying polypeptide comprises a portion of a polypeptide listed in any of Tables A1, T1, or T2, wherein the portion comprises a linker and RT domain, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity to said portion.


In certain embodiments, the gene modifying polypeptide comprises the linker of an amino acid sequence selected from SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a sequence encoding the linker of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a sequence encoding the linker of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises the linker of a polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.


In certain embodiments, the gene modifying polypeptide comprises the RT domain of an amino acid sequence selected from SEQ ID NOs: 1-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a sequence encoding the RT domain of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 6001-7743, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises a sequence encoding the RT domain of a polypeptide having an amino acid sequence selected from SEQ ID NOs: 4501-4541, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto. In certain embodiments, the gene modifying polypeptide comprises the RT domain of a polypeptide as listed in any of Tables A1, T1, or T2, or an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity thereto.










Lengthy table referenced here




US20240084334A1-20240314-T00001


Please refer to the end of the specification for access instructions.






Localization Sequences for Gene Modifying Systems

In certain embodiments, a gene editor system RNA further comprises an intracellular localization sequence, e.g., a nuclear localization sequence (NLS). In some embodiments, a gene modifying polypeptide comprises an NLS as comprised in SEQ ID NO: 4000 and/or SEQ ID NO: 4001, or an NLS having an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


The nuclear localization sequence may be an RNA sequence that promotes the import of the RNA into the nucleus. In certain embodiments the nuclear localization signal is located on the template RNA. In certain embodiments, the gene modifying polypeptide is encoded on a first RNA, and the template RNA is a second, separate, RNA, and the nuclear localization signal is located on the template RNA and not on an RNA encoding the gene modifying polypeptide. While not wishing to be bound by theory, in some embodiments, the RNA encoding the gene modifying polypeptide is targeted primarily to the cytoplasm to promote its translation, while the template RNA is targeted primarily to the nucleus to promote insertion into the genome. In some embodiments the nuclear localization signal is at the 3′ end, 5′ end, or in an internal region of the template RNA. In some embodiments the nuclear localization signal is 3′ of the heterologous sequence (e.g., is directly 3′ of the heterologous sequence) or is 5′ of the heterologous sequence (e.g., is directly 5′ of the heterologous sequence). In some embodiments the nuclear localization signal is placed outside of the 5′ UTR or outside of the 3′ UTR of the template RNA. In some embodiments the nuclear localization signal is placed between the 5′ UTR and the 3′ UTR, wherein optionally the nuclear localization signal is not transcribed with the transgene (e.g., the nuclear localization signal is an anti-sense orientation or is downstream of a transcriptional termination signal or polyadenylation signal). In some embodiments the nuclear localization sequence is situated inside of an intron. In some embodiments a plurality of the same or different nuclear localization signals are in the RNA, e.g., in the template RNA. In some embodiments the nuclear localization signal is less than 5, 10, 25, 50, 75, 100, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900 or 1000 bp in length. Various RNA nuclear localization sequences can be used. For example, Lubelsky and Ulitsky, Nature 555 (107-111), 2018 describe RNA sequences which drive RNA localization into the nucleus. In some embodiments, the nuclear localization signal is a SINE-derived nuclear RNA localization (SIRLOIN) signal. In some embodiments the nuclear localization signal binds a nuclear-enriched protein. In some embodiments the nuclear localization signal binds the HNRNPK protein. In some embodiments the nuclear localization signal is rich in pyrimidines, e.g., is a C/T rich, C/U rich, C rich, T rich, or U rich region. In some embodiments the nuclear localization signal is derived from a long non-coding RNA. In some embodiments the nuclear localization signal is derived from MALAT1 long non-coding RNA or is the 600 nucleotide M region of MALAT1 (described in Miyagawa et al., RNA 18, (738-751), 2012). In some embodiments the nuclear localization signal is derived from BORG long non-coding RNA or is a AGCCC motif (described in Zhang et al., Molecular and Cellular Biology 34, 2318-2329 (2014). In some embodiments the nuclear localization sequence is described in Shukla et al., The EAIBO Journal e98452 (2018). In some embodiments the nuclear localization signal is derived from a retrovirus.


In some embodiments, a polypeptide described herein comprises one or more (e.g., 2, 3, 4, 5) nuclear targeting sequences, for example a nuclear localization sequence (NLS). In some embodiments, the NLS is a bipartite NLS. In some embodiments, an NLS facilitates the import of a protein comprising an NLS into the cell nucleus. In some embodiments, the NLS is fused to the N-terminus of a gene modifying polypeptide as described herein. In some embodiments, the NLS is fused to the C-terminus of the gene modifying polypeptide. In some embodiments, the NLS is fused to the N-terminus or the C-terminus of a Cas domain. In some embodiments, a linker sequence is disposed between the NLS and the neighboring domain of the gene modifying polypeptide.


In some embodiments, an NLS comprises the amino acid sequence MDSLLMNRRKFLYQFKNVRWAKGRRETYLC (SEQ ID NO: 5009), PKKRKVEGADKRTADGSEFESPKKKRKV(SEQ ID NO: 5010), RKSGKIAAIWKRPRKPKKKRKV (SEQ ID NO: 5011) KRTADGSEFESPKKKRKV(SEQ ID NO: 5012), KKTELQTTNAENKTKKL (SEQ ID NO: 5013), or KRGINDRNFWRGENGRKTR (SEQ ID NO: 5014), KRPAATKKAGQAKKKK (SEQ ID NO: 5015), PAAKRVKLD (SEQ ID NO:4644), KRTADGSEFEKRTADGSEFESPKKKAKVE (SEQ ID NO: 4649), KRTADGSEFE (SEQ ID NO: 4650), KRTADGSEFESPKKKAKVE (SEQ ID NO: 4651), AGKRTADGSEFEKRTADGSEFESPKKKAKVE (SEQ ID NO: 4001), or a functional fragment or variant thereof. Exemplary NLS sequences are also described in PCT/EP2000/011690, the contents of which are incorporated herein by reference for their disclosure of exemplary nuclear localization sequences. In some embodiments, an NLS comprises an amino acid sequence as disclosed in Table 11. An NLS of this table may be utilized with one or more copies in a polypeptide in one or more locations in a polypeptide, e.g., 1, 2, 3 or more copies of an NLS in an N-terminal domain, between peptide domains, in a C-terminal domain, or in a combination of locations, in order to improve subcellular localization to the nucleus. Multiple unique sequences may be used within a single polypeptide. Sequences may be naturally monopartite or bipartite, e.g., having one or two stretches of basic amino acids, or may be used as chimeric bipartite sequences. Sequence references correspond to UniProt accession numbers, except where indicated as SeqNLS for sequences mined using a subcellular localization prediction algorithm (Lin et al BMC Bioinformat 13:157 (2012), incorporated herein by reference in its entirety).









TABLE 11







Exemplary nuclear localization


signals for use in gene modifying systems











SEQ


Sequence
Sequence References
ID No.





AHFKISGEKRPSTDPGKKAK
Q76IQ7
5223


NPKKKKKKDP







AHRAKKMSKTHA
P21827
5224





ASPEYVNLPINGNG
SeqNLS
5225





CTKRPRW
O88622, Q86W56, Q9QYM2, O02776
5226





DKAKRVSRNKSEKKRR
O15516, Q5RAK8, Q91YB2, Q91YB0,
5227



Q8QGQ6, O08785, Q9WVS9, Q6YGZ4






EELRLKEELLKGIYA
Q9QY16, Q9UHL0, Q2TBP1, Q9QY15
5228





EEQLRRRKNSRLNNTG
G5EFF5
5229





EVLKVIRTGKRKKKAWKR
SeqNLS
5230


MVTKVC







HHHHHHHHHHHHQPH
Q63934, G3V7L5, Q12837
5231





HKKKHPDASVNFSEFSK
P10103, Q4R844, P12682, B0CM99,
5232



A9RA84, Q6YKA4, P09429, P63159,




Q08IE6, P63158, Q9YH06, B1MTB0






HKRTKK
Q2R2D5
5233





IINGRKLKLKKSRRRSSQTS
SeqNLS
5234


NNSFTSRRS







KAEQERRK
Q8LH59
5235





KEKRKRREELFIEQKKRK
SeqNLS
5236





KKGKDEWFSRGKKP
P30999
5237





KKGPSVQKRKKT
Q6ZN17
5238





KKKTVINDLLHYKKEK
SeqNLS, P32354
5239





KKNGGKGKNKPSAKIKK
SeqNLS
5240





KKPKWDDFKKKKK
Q15397, Q8BKS9, Q562C7
5241





KKRKKD
SeqNLS, Q91Z62, Q1A730, Q969P5,
5242



Q2KHT6, Q9CPU7






KKRRKRRRK
SeqNLS
5243





KKRRRRARK
Q9UMS6, D4A702, Q91YE8
5244





KKSKRGR
Q9UBS0
5245





KKSRKRGS
B4FG96
5246





KKSTALSRELGKIMRRR
SeqNLS, P32354
5247





KKSYQDPEIIAHSRPRK
Q9U7C9
5248





KKTGKNRKLKSKRVKTR
Q9Z301, O54943, Q8K3T2
5249





KKVSIAGQSGKLWRWKR
Q6YUL8
5250





KKYENVVIKRSPRKRGRPR
SeqNLS
5251


K







KNKKRK
SeqNLS
5252





KPKKKR
SeqNLS
5253





KRAMKDDSHGNSTSPKRRK
Q0E671
5254





KRANSNLVAAYEKAKKK
P23508
5255





KRASEDTTSGSPPKKSSAGP
Q9BZZ5, Q5R644
5256


KR







KRFKRRWMVRKMKTKK
SeqNLS
5257





KRGLNSSFETSPKKVK
Q8IV63
5258





KRGNSSIGPNDLSKRKQRK
SeqNLS
5259


K







KRIHSVSLSQSQIDPSKKVK
SeqNLS
5260


RAK







KRKGKLKNKGSKRKK
O15381
5261





KRRRRRRREKRKR
Q96GM8
5262





KRSNDRTYSPEEEKQRRA
Q91ZF2
5263





KRTVATNGDASGAHRAKK
SeqNLS
5264


MSK







KRVYNKGEDEQEHLPKGKK
SeqNLS
5265


R







KSGKAPRRRAVSMDNSNK
Q9WVH4, O43524
5266





KVNFLDMSLDDIIIYKELE
Q9P127
5267





KVQHRIAKKTTRRRR
Q9DXE6
5268





LSPSLSPL
Q9Y261, P32182, P35583
5269





MDSLLMNRRKFLYQFKNVR
Q9GZX7
5270


WAKGRRETYLC







MPQNEYIELHRKRYGYRLD
SeqNLS
5271


YHEKKRKKESREAHERSKK




AKKMIGLKAKLYHK







MVQLRPRASR
SeqNLS
5272





NNKLLAKRRKGGASPKDDP
Q965G5
5273


MDDIK







NYKRPMDGTYGPPAKRHEG
O14497, A2BH40
5274


E







PDTKRAKLDSSETTMVKKK
SeqNLS
5275





PEKRTKI
SeqNLS
5276





PGGRGKKK
Q719N1, Q9UBP0, A2VDN5
5277





PGKMDKGEHRQERRDRPY
Q01844, Q61545
5278





PKKGDKYDKTD
Q45FA5
5279





PKKKSRK
O35914, Q01954
5280





PKKNKPE
Q22663
5281





PKKRAKV
P04295, P89438
5282





PKPKKLKVE
P55263, P55262, P55264, Q64640
5283





PKRGRGR
Q9FYS5, Q43386
5284





PKRRLVDDA
P0C797
5285





PKRRRTY
SeqNLS
5286





PLFKRR
A8X6H4, Q9TXJ0
5287





PLRKAKR
Q86WB0, Q5R8V9
5288





PPAKRKCIF
Q6AZ28, O75928, Q8C5D8
5289





PPARRRRL
Q8NAG6
5290





PPKKKRKV
Q3L6L5, P03070, P14999, P03071
5291





PPNKRMKVKH
Q8BN78
5292





PPRIYPQLPSAPT
P0C799
5293





PQRSPFPKSSVKR
SeqNLS
5294





PRPRKVPR
P0C799
5295





PRRRVQRKR
SeqNLS, Q5R448, Q5TAQ9
5296





PRRVRLK
Q58DJ0, P56477, Q13568
5297





PSRKRPR
Q62315, Q5F363, Q92833
5298





PSSKKRKV
SeqNLS
5299





PTKKRVK
P07664
5300





QRPGPYDRP
SeqNLS
5301





RGKGGKGLGKGGAKRHRK
SeqNLS
5302





RKAGKGGGGHKTTKKRSA
B4FG96
5303


KDEKVP







RKIKLKRAK
A1L3G9
5304





RKIKRKRAK
B9X187
5305





RKKEAPGPREELRSRGR
O35126, P54258, Q5IS70, P54259
5306





RKKRKGK
SeqNLS, Q29243, Q62165, Q28685,
5307



O18738, Q9TSZ6, Q14118






RKKRRQRRR
P04326, P69697, P69698, P05907,
5308



P20879, P04613, P19553, P0C1J9,




P20893, P12506, P04612, Q73370,




P0C1K0, P05906, P35965, P04609,




P04610, P04614, P04608, P05905






RKKSIPLSIKNLKRKHKRKK
Q9C0C9
5309


NKITR







RKLVKPKNTKMKTKLRTNP
Q14190
5310


Y







RKRLILSDKGQLDWKK
SeqNLS, Q91Z62, Q1A730, Q2KHT6,
5311



Q9CPU7






RKRLKSK
Q13309
5312





RKRRVRDNM
Q8QPH4, Q809M7, A8C8X1, Q2VNC5,
5313



Q38SQ0, O89749, Q6DNQ9, Q809L9,




Q0A429, Q20NV3, P16509, P16505,




Q6DNQ5, P16506, Q6XT06, P26118,




Q2ICQ2, Q2RCG8, Q0A2D0, Q0A2H9,




Q9IQ46, Q809M3, Q6J847, Q6J856,




B4URE4, A4GCM7, Q0A440, P26120,




P16511,






RKRSPKDKKEKDLDGAGKR
Q7RTP6
5314


RKT







RKRTPRVDGQTGENDMNK
O94851
5315


RRRK







RLPVRRRRRR
P04499, P12541, P03269, P48313,
5316



P03270






RLRFRKPKSK
P69469
5317





RQQRKR
Q14980
5318





RRDLNSSFETSPKKVK
Q8K3G5
5319





RRDRAKLR
Q9SLB8
5320





RRGDGRRR
Q80WE1, Q5R9B4, Q06787, P35922
5321





RRGRKRKAEKQ
Q812D1, Q5XXA9, Q99JF8, Q8MJG1,
5322



Q66T72, O75475






RRKKRR
Q0VD86, Q58DS6, Q5R6G2, Q9ERI5,
5323



Q6AYK2, Q6NYC1






RRKRSKSEDMDSVESKRRR
Q7TT18
5324





RRKRSR
Q99PU7, D3ZHS6, Q92560, A2VDM8
5325





RRPKGKTLQKRKPK
Q6ZN17
5326





RRRGFERFGPDNMGRKRK
Q63014, Q9DBR0
5327





RRRGKNKVAAQNCRK
SeqNLS
5328





RRRKRR
Q5FVH8, Q6MZT1, Q08DH5, Q8BQP9
5329





RRRQKQKGGASRRR
SeqNLS
5330





RRRREGPRARRRR
P08313, P10231
5331





RRTIRLKLVYDKCDRSCKIQ
SeqNLS
5332


KKNRNKCQYCRFHKCLSVG




MSHNAIRFGRMPRSEKAKL




KAE







RRVPQRKEVSRCRKCRK
Q5RJN4, Q32L09, Q8CAK3, Q9NUL5
5333





RVGGRRQAVECIEDLLNEP
P03255
5334


GQPLDLSCKRPRP







RVVKLRIAP
P52639, Q8JMN0
5335





RVVRRR
P70278
5336





SKRKTKISRKTR
Q5RAY1, O00443
5337





SYVKTVPNRTRTYIKL
P21935
5338





TGKNEAKKRKIA
P52739, Q8K3J5, Q5RAU9
5339





TLSPASSPSSVSCPVIPASTD
SeqNLS
5340


ESPGSALNI







VSKKQRTGKKIH
P52739, Q8K3J5, Q5RAU9
5341





SPKKKRKVE

5342





KRTAD GSEFE SPKKKRKVE

5343





PAAKRVKLD

5344





PKKKRKV

5345





MDSLLMNRRKFLYQFKNVR

5346


WAKGRRETYLC







SPKKKRKVEAS

5347





MAPKKKRKVGIHRGVP

5348





KRTADGSEFEKRTADGSEFE

5349


SPKKKAKVE







KRTADGSEFE

5350





KRTADGSEFESPKKKAKVE

5351





AGKRTADGSEFEKRTADGS

4001


EFESPKKKAKVE









In some embodiments, the NLS is a bipartite NLS. A bipartite NLS typically comprises two basic amino acid clusters separated by a spacer sequence (which may be, e.g., about 10 amino acids in length). A monopartite NLS typically lacks a spacer. An example of a bipartite NLS is the nucleoplasmin NLS, having the sequence KR[PAATKKAGQA]KKKK (SEQ ID NO: 5015), wherein the spacer is bracketed. Another exemplary bipartite NLS has the sequence PKKKRKVEGADKRTADGSEFESPKKKRKV (SEQ ID NO: 5016). Exemplary NLSs are described in International Application WO2020051561, which is herein incorporated by reference in its entirety, including for its disclosures regarding nuclear localization sequences.


In certain embodiments, a gene editor system polypeptide (e.g., a gene modifying polypeptide as described herein) further comprises an intracellular localization sequence, e.g., a nuclear localization sequence and/or a nucleolar localization sequence. The nuclear localization sequence and/or nucleolar localization sequence may be amino acid sequences that promote the import of the protein into the nucleus and/or nucleolus, where it can promote integration of heterologous sequence into the genome. In certain embodiments, a gene editor system polypeptide (e.g., (e.g., a gene modifying polypeptide as described herein) further comprises a nucleolar localization sequence. In certain embodiments, the gene modifying polypeptide is encoded on a first RNA, and the template RNA is a second, separate, RNA, and the nucleolar localization signal is encoded on the RNA encoding the gene modifying polypeptide and not on the template RNA. In some embodiments, the nucleolar localization signal is located at the N-terminus, C-terminus, or in an internal region of the polypeptide. In some embodiments, a plurality of the same or different nucleolar localization signals are used. In some embodiments, the nuclear localization signal is less than 5, 10, 25, 50, 75, or 100 amino acids in length. Various polypeptide nucleolar localization signals can be used. For example, Yang et al., Journal of Biomedical Science 22, 33 (2015), describe a nuclear localization signal that also functions as a nucleolar localization signal. In some embodiments, the nucleolar localization signal may also be a nuclear localization signal. In some embodiments, the nucleolar localization signal may overlap with a nuclear localization signal. In some embodiments, the nucleolar localization signal may comprise a stretch of basic residues. In some embodiments, the nucleolar localization signal may be rich in arginine and lysine residues. In some embodiments, the nucleolar localization signal may be derived from a protein that is enriched in the nucleolus. In some embodiments, the nucleolar localization signal may be derived from a protein enriched at ribosomal RNA loci. In some embodiments, the nucleolar localization signal may be derived from a protein that binds rRNA. In some embodiments, the nucleolar localization signal may be derived from MSP58. In some embodiments, the nucleolar localization signal may be a monopartite motif. In some embodiments, the nucleolar localization signal may be a bipartite motif. In some embodiments, the nucleolar localization signal may consist of a multiple monopartite or bipartite motifs. In some embodiments, the nucleolar localization signal may consist of a mix of monopartite and bipartite motifs. In some embodiments, the nucleolar localization signal may be a dual bipartite motif. In some embodiments, the nucleolar localization motif may be a KRASSQALGTIPKRRSSSRFIKRKK (SEQ ID NO: 5017). In some embodiments, the nucleolar localization signal may be derived from nuclear factor-KB-inducing kinase. In some embodiments, the nucleolar localization signal may be an RKKRKKK motif (SEQ ID NO: 5018) (described in Birbach et al., Journal of Cell Science, 117 (3615-3624), 2004).


Evolved Variants of Gene Modifying Polypeptides and Systems


In some embodiments, the invention provides evolved variants of gene modifying polypeptides as described herein. Evolved variants can, in some embodiments, be produced by mutagenizing a reference gene modifying polypeptide, or one of the fragments or domains comprised therein. In some embodiments, one or more of the domains (e.g., the reverse transcriptase domain) is evolved. One or more of such evolved variant domains can, in some embodiments, be evolved alone or together with other domains. An evolved variant domain or domains may, in some embodiments, be combined with unevolved cognate component(s) or evolved variants of the cognate component(s), e.g., which may have been evolved in either a parallel or serial manner.


In some embodiments, the process of mutagenizing a reference gene modifying polypeptide, or fragment or domain thereof, comprises mutagenizing the reference gene modifying polypeptide or fragment or domain thereof. In embodiments, the mutagenesis comprises a continuous evolution method (e.g., PACE) or non-continuous evolution method (e.g., PANCE), e.g., as described herein. In some embodiments, the evolved gene modifying polypeptide, or a fragment or domain thereof, comprises one or more amino acid variations introduced into its amino acid sequence relative to the amino acid sequence of the reference gene modifying polypeptide, or fragment or domain thereof. In embodiments, amino acid sequence variations may include one or more mutated residues (e.g., conservative substitutions, non-conservative substitutions, or a combination thereof) within the amino acid sequence of a reference gene modifying polypeptide, e.g., as a result of a change in the nucleotide sequence encoding the gene modifying polypeptide that results in, e.g., a change in the codon at any particular position in the coding sequence, the deletion of one or more amino acids (e.g., a truncated protein), the insertion of one or more amino acids, or any combination of the foregoing. The evolved variant gene modifying polypeptide may include variants in one or more components or domains of the gene modifying polypeptide (e.g., variants introduced into a reverse transcriptase domain).


In some aspects, the disclosure provides gene modifying polypeptides, systems, kits, and methods using or comprising an evolved variant of a gene modifying polypeptide, e.g., employs an evolved variant of a gene modifying polypeptide or a gene modifying polypeptide produced or producible by PACE or PANCE. In embodiments, the unevolved reference gene modifying polypeptide is a gene modifying polypeptide as disclosed herein.


The term “phage-assisted continuous evolution (PACE),”as used herein, generally refers to continuous evolution that employs phage as viral vectors. Examples of PACE technology have been described, for example, in International PCT Application No. PCT/US 2009/056194, filed Sep. 8, 2009, published as WO 2010/028347 on Mar. 11, 2010; International PCT Application, PCT/US2011/066747, filed Dec. 22, 2011, published as WO 2012/088381 on Jun. 28, 2012; U.S. Pat. No. 9,023,594, issued May 5, 2015; U.S. Pat. No. 9,771,574, issued Sep. 26, 2017; U.S. Pat. No. 9,394,537, issued Jul. 19, 2016; International PCT Application, PCT/US2015/012022, filed Jan. 20, 2015, published as WO 2015/134121 on Sep. 11, 2015; U.S. Pat. No. 10,179,911, issued Jan. 15, 2019; and International PCT Application, PCT/US2016/027795, filed Apr. 15, 2016, published as WO 2016/168631 on Oct. 20, 2016, the entire contents of each of which are incorporated herein by reference.


The term “phage-assisted non-continuous evolution (PANCE),” as used herein, generally refers to non-continuous evolution that employs phage as viral vectors. Examples of PANCE technology have been described, for example, in Suzuki T. et al, Crystal structures reveal an elusive functional domain of pyrrolysyl-tRNA synthetase, Nat Chem Biol. 13(12): 1261-1266 (2017), incorporated herein by reference in its entirety. Briefly, PANCE is a technique for rapid in vivo directed evolution using serial flask transfers of evolving selection phage (SP), which contain a gene of interest to be evolved, across fresh host cells (e.g., E. coli cells). Genes inside the host cell may be held constant while genes contained in the SP continuously evolve. Following phage growth, an aliquot of infected cells may be used to transfect a subsequent flask containing host E. coli. This process can be repeated and/or continued until the desired phenotype is evolved, e.g., for as many transfers as desired.


Methods of applying PACE and PANCE to gene modifying polypeptides may be readily appreciated by the skilled artisan by reference to, inter alia, the foregoing references. Additional exemplary methods for directing continuous evolution of genome-modifying proteins or systems, e.g., in a population of host cells, e.g., using phage particles, can be applied to generate evolved variants of gene modifying polypeptides, or fragments or subdomains thereof. Non-limiting examples of such methods are described in International PCT Application, PCT/US2009/056194, filed Sep. 8, 2009, published as WO 2010/028347 on Mar. 11, 2010; International PCT Application, PCT/US2011/066747, filed Dec. 22, 2011, published as WO 2012/088381 on Jun. 28, 2012; U.S. Pat. No. 9,023,594, issued May 5, 2015; U.S. Pat. No. 9,771,574, issued Sep. 26, 2017; U.S. Pat. No. 9,394,537, issued Jul. 19, 2016; International PCT Application, PCT/US2015/012022, filed Jan. 20, 2015, published as WO 2015/134121 on Sep. 11, 2015; U.S. Pat. No. 10,179,911, issued Jan. 15, 2019; International Application No. PCT/US2019/37216, filed Jun. 14, 2019, International Patent Publication WO 2019/023680, published Jan. 31, 2019, International PCT Application, PCT/US2016/027795, filed Apr. 15, 2016, published as WO 2016/168631 on Oct. 20, 2016, and International Patent Publication No. PCT/US2019/47996, filed Aug. 23, 2019, each of which is incorporated herein by reference in its entirety.


In some non-limiting illustrative embodiments, a method of evolution of a evolved variant gene modifying polypeptide, of a fragment or domain thereof, comprises: (a) contacting a population of host cells with a population of viral vectors comprising the gene of interest (the starting gene modifying polypeptide or fragment or domain thereof), wherein: (1) the host cell is amenable to infection by the viral vector; (2) the host cell expresses viral genes required for the generation of viral particles; (3) the expression of at least one viral gene required for the production of an infectious viral particle is dependent on a function of the gene of interest; and/or (4) the viral vector allows for expression of the protein in the host cell, and can be replicated and packaged into a viral particle by the host cell. In some embodiments, the method comprises (b) contacting the host cells with a mutagen, using host cells with mutations that elevate mutation rate (e.g., either by carrying a mutation plasmid or some genome modification—e.g., proofing-impaired DNA polymerase, SOS genes, such as UmuC, UmuD′, and/or RecA, which mutations, if plasmid-bound, may be under control of an inducible promoter), or a combination thereof. In some embodiments, the method comprises (c) incubating the population of host cells under conditions allowing for viral replication and the production of viral particles, wherein host cells are removed from the host cell population, and fresh, uninfected host cells are introduced into the population of host cells, thus replenishing the population of host cells and creating a flow of host cells. In some embodiments, the cells are incubated under conditions allowing for the gene of interest to acquire a mutation. In some embodiments, the method further comprises (d) isolating a mutated version of the viral vector, encoding an evolved gene product (e.g., an evolved variant gene modifying polypeptide, or fragment or domain thereof), from the population of host cells.


The skilled artisan will appreciate a variety of features employable within the above-described framework. For example, in some embodiments, the viral vector or the phage is a filamentous phage, for example, an M13 phage, e.g., an M13 selection phage. In certain embodiments, the gene required for the production of infectious viral particles is the M13 gene III (gIII) In embodiments, the phage may lack a functional gIII, but otherwise comprise gI, gII, gIV, gV, gVI, gVII, gVIII, gIX, and a gX. In some embodiments, the generation of infectious VSV particles involves the envelope protein VSV-G. Various embodiments can use different retroviral vectors, for example, Murine Leukemia Virus vectors, or Lentiviral vectors. In embodiments, the retroviral vectors can efficiently be packaged with VSV-G envelope protein, e.g., as a substitute for the native envelope protein of the virus.


In some embodiments, host cells are incubated according to a suitable number of viral life cycles, e.g., at least 10, at least 20, at least 30, at least 40, at least 50, at least 100, at least 200, at least 300, at least 400, at least, 500, at least 600, at least 700, at least 800, at least 900, at least 1000, at least 1250, at least 1500, at least 1750, at least 2000, at least 2500, at least 3000, at least 4000, at least 5000, at least 7500, at least 10000, or more consecutive viral life cycles, which in on illustrative and non-limiting examples of M13 phage is 10-20 minutes per virus life cycle. Similarly, conditions can be modulated to adjust the time a host cell remains in a population of host cells, e.g., about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 20, about 21, about 22, about 23, about 24, about 25, about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 70, about 80, about 90, about 100, about 120, about 150, or about 180 minutes. Host cell populations can be controlled in part by density of the host cells, or, in some embodiments, the host cell density in an inflow, e.g., 103 cells/ml, about 104 cells/ml, about 105 cells/ml, about 5-105 cells/ml, about 106 cells/ml, about 5-106 cells/ml, about 107 cells/ml, about 5-107 cells/ml, about 108 cells/ml, about 5-108 cells/ml, about 109 cells/ml, about 5. 109 cells/ml, about 1010 cells/ml, or about 5·1010 cells/ml.


Inteins


In some embodiments, as described in more detail below, an intein-N(intN) domain may be fused to the N-terminal portion of a first domain of a gene modifying polypeptide described herein, and an intein-C(intC) domain may be fused to the C-terminal portion of a second domain of a gene modifying polypeptide described herein for the joining of the N-terminal portion to the C-terminal portion, thereby joining the first and second domains. In some embodiments, the first and second domains are each independently chosen from a DNA binding domain, an RNA binding domain, an RT domain, and an endonuclease domain.


Inteins can occur as self-splicing protein intron (e.g., peptide), e.g., which ligates flanking N-terminal and C-terminal exteins (e.g., fragments to be joined). An intein may, in some instances, comprise a fragment of a protein that is able to excise itself and join the remaining fragments (the exteins) with a peptide bond in a process known as protein splicing. Inteins are also referred to as “protein introns.” The process of an intein excising itself and joining the remaining portions of the protein is herein termed “protein splicing” or “intein-mediated protein splicing.”


In some embodiments, an intein of a precursor protein (an intein containing protein prior to intein-mediated protein splicing) comes from two genes. Such intein is referred to herein as a split intein (e.g., split intein-N and split intein-C). Accordingly, an intein-based approach may be used to join a first polypeptide sequence and a second polypeptide sequence together. For example, in cyanobacteria, DnaE, the catalytic subunit a of DNA polymerase III, is encoded by two separate genes, dnaE-n and dnaE-c. An intein-N domain, such as that encoded by the dnaE-n gene, when situated as part of a first polypeptide sequence, may join the first polypeptide sequence with a second polypeptide sequence, wherein the second polypeptide sequence comprises an intein-C domain, such as that encoded by the dnaE-c gene. Accordingly, in some embodiments, a protein can be made by providing nucleic acid encoding the first and second polypeptide sequences (e.g., wherein a first nucleic acid molecule encodes the first polypeptide sequence and a second nucleic acid molecule encodes the second polypeptide sequence), and the nucleic acid is introduced into the cell under conditions that allow for production of the first and second polypeptide sequences, and for joining of the first to the second polypeptide sequence via an intein-based mechanism.


Use of inteins for joining heterologous protein fragments is described, for example, in Wood et al., J. Biol. Chem.289(21); 14512-9 (2014) (incorporated herein by reference in its entirety). For example, when fused to separate protein fragments, the inteins IntN and IntC may recognize each other, splice themselves out, and/or simultaneously ligate the flanking N- and C-terminal exteins of the protein fragments to which they were fused, thereby reconstituting a full-length protein from the two protein fragments.


In some embodiments, a synthetic intein based on the dnaE intein, the Cfa-N(e.g., split intein-N) and Cfa-C(e.g., split intein-C) intein pair, is used. Examples of such inteins have been described, e.g., in Stevens et al., J Am Chem Soc. 2016 Feb. 24; 138(7):2162-5 (incorporated herein by reference in its entirety). Non-limiting examples of intein pairs that may be used in accordance with the present disclosure include: Cfa DnaE intein, Ssp GyrB intein, Ssp DnaX intein, Ter DnaE3 intein, Ter ThyX intein, Rma DnaB intein and Cne Prp8 intein (e.g., as described in U.S. Pat. No. 8,394,604, incorporated herein by reference.


In some embodiments involving a split Cas9, an intein-N domain and an intein-C domain may be fused to the N-terminal portion of the split Cas9 and the C-terminal portion of a split Cas9, respectively, for the joining of the N-terminal portion of the split Cas9 and the C-terminal portion of the split Cas9. For example, in some embodiments, an intein-N is fused to the C-terminus of the N-terminal portion of the split Cas9, i.e., to form a structure of N—[N-terminal portion of the split Cas9]-[intein-N]˜C. In some embodiments, an intein-C is fused to the N-terminus of the C-terminal portion of the split Cas9, i.e., to form a structure of N-[intein-C]˜[C-terminal portion of the split Cas9]-C. The mechanism of intein-mediated protein splicing for joining the proteins the inteins are fused to (e.g., split Cas9) is described in Shah et al., Chem Sci. 2014; 5(1):446-461, incorporated herein by reference. Methods for designing and using inteins are known in the art and described, for example by WO2020051561, WO2014004336, WO2017132580, US20150344549, and US20180127780, each of which is incorporated herein by reference in their entirety.


In some embodiments, a split refers to a division into two or more fragments. In some embodiments, a split Cas9 protein or split Cas9 comprises a Cas9 protein that is provided as an N-terminal fragment and a C-terminal fragment encoded by two separate nucleotide sequences. The polypeptides corresponding to the N-terminal portion and the C-terminal portion of the Cas9 protein may be spliced to form a reconstituted Cas9 protein. In embodiments, the Cas9 protein is divided into two fragments within a disordered region of the protein, e.g., as described in Nishimasu et al., Cell, Volume 156, Issue 5, pp. 935-949, 2014, or as described in Jiang et al. (2016) Science 351: 867-871 and PDB file: 5F9R (each of which is incorporated herein by reference in its entirety). A disordered region may be determined by one or more protein structure determination techniques known in the art, including, without limitation, X-ray crystallography, NMR spectroscopy, electron microscopy (e.g., cryoEM), and/or in silico protein modeling. In some embodiments, the protein is divided into two fragments at any C, T, A, or S, e.g., within a region of SpCas9 between amino acids A292-G364, F445-K483, or E565-T637, or at corresponding positions in any other Cas9, Cas9 variant (e.g., nCas9, dCas9), or other napDNAbp. In some embodiments, protein is divided into two fragments at SpCas9 T310, T313, A456, S469, or C574. In some embodiments, the process of dividing the protein into two fragments is referred to as splitting the protein.


In some embodiments, a protein fragment ranges from about 2-1000 amino acids (e.g., between 2-10, 10-50, 50-100, 100-200, 200-300, 300-400, 400-500, 500-600, 600-700, 700-800, 800-900, or 900-1000 amino acids) in length. In some embodiments, a protein fragment ranges from about 5-500 amino acids (e.g., between 5-10, 10-50, 50-100, 100-200, 200-300, 300-400, or 400-500 amino acids) in length. In some embodiments, a protein fragment ranges from about 20-200 amino acids (e.g., between 20-30, 30-40, 40-50, 50-100, or 100-200 amino acids) in length.


In some embodiments, a portion or fragment of a gene modifying polypeptide is fused to an intein. The nuclease can be fused to the N-terminus or the C-terminus of the intein. In some embodiments, a portion or fragment of a fusion protein is fused to an intein and fused to an AAV capsid protein. The intein, nuclease and capsid protein can be fused together in any arrangement (e.g., nuclease-intein-capsid, intein-nuclease-capsid, capsid-intein-nuclease, etc.). In some embodiments, the N-terminus of an intein is fused to the C-terminus of a fusion protein and the C-terminus of the intein is fused to the N-terminus of an AAV capsid protein.


In some embodiments, an endonuclease domain (e.g., a nickase Cas9 domain) is fused to intein-N and a polypeptide comprising an RT domain is fused to an intein-C.


Exemplary nucleotide and amino acid sequences of intein-N domains and compatible intein-C domains are provided below:









DnaE Intein-N DNA:


(SEQ ID NO: 5029)


TGCCTGTCATACGAAACCGAGATACTGACAGTAGAATATGGCCTTCTGCC





AATCGGGAAGATTGTGGAGAAACGGATAGAATGCACAGTTTACTCTGTCG





ATAACAATGGTAACATTTATACTCAGCCAGTTGCCCAGTGGCACGACCGG





GGAGAGCAGGAAGTATTCGAATACTGTCTGGAGGATGGAAGTCTCATTAG





GGCCACTAAGGACCACAAATTTATGACAGTCGATGGCCAGATGCTGCCTA





TAGACGAAATCTTTGAGCGAGAGTTGGACCTCATGCGAGTTGACAACCTT





CCTAAT





DnaE Intein-N Protein:


(SEQ ID NO: 5030)


CLSYETEILTVEYGLLPIGKIVEKRIECTVYSVDNNGNIYTQPVAQWHDR





GEQEVFEYCLEDGSLIRATKDHKFMTVDGQMLPIDEIFERELDLMRVDNL





PN





DnaE Intein-C DNA:


(SEQ ID NO: 5031)


ATGATCAAGATAGCTACAAGGAAGTATCTTGGCAAACAAAACGTTTATGA





TATTGGAGTCGAAAGAGATCACAACTTTGCTCTGAAGAACGGATTCATAG





CTTCTAAT





DnaE Intein-C Protein:


(SEQ ID NO: 5032)


MIKIATRKYLGKQNVYDIGVERDHNFALKNGFIASN





Cfa-N DNA:


(SEQ ID NO: 5033)


TGCCTGTCTTATGATACCGAGATACTTACCGTTGAATATGGCTTCTTGCC





TATTGGAAAGATTGTCGAAGAGAGAATTGAATGCACAGTATATACTGTAG





ACAAGAATGGTTTCGTTTACACACAGCCCATTGCTCAATGGCACAATCGC





GGCGAACAAGAAGTATTTGAGTACTGTCTCGAGGATGGAAGCATCATACG





AGCAACTAAAGATCATAAATTCATGACCACTGACGGGCAGATGTTGCCAA





TAGATGAGATATTCGAGCGGGGCTTGGATCTCAAACAAGTGGATGGATTG





CCA





Cfa-N Protein:


(SEQ ID NO: 5034)


CLSYDTEILTVEYGFLPIGKIVEERIECTVYTVDKNGFVYTQPIAQWHNR





GEQEVFEYCLEDGSIIRATKDHKFMTTDGQMLPIDEIFERGLDLKQVDGL





P





Cfa-C DNA:


(SEQ ID NO: 5035)


ATGAAGAGGACTGCCGATGGATCAGAGTTTGAATCTCCCAAGAAGAAGAG





GAAAGTAAAGATAATATCTCGAAAAAGTCTTGGTACCCAAAATGTCTATG





ATATTGGAGTGGAGAAAGATCACAACTTCCTTCTCAAGAACGGTCTCGTA





GCCAGCAAC





Cfa-C Protein:


(SEQ ID NO: 5036)


MKRTADGSEFESPKKKRKVKIISRKSLGTQNVYDIGVEKDHNFLLKNGLV





ASN






Additional Domains


The gene modifying polypeptide can bind a target DNA sequence and template nucleic acid (e.g., template RNA), nick the target site, and write (e.g., reverse transcribe) the template into DNA, resulting in a modification of the target site. In some embodiments, additional domains may be added to the polypeptide to enhance the efficiency of the process. In some embodiments, the gene modifying polypeptide may contain an additional DNA ligation domain to join reverse transcribed DNA to the DNA of the target site. In some embodiments, the polypeptide may comprise a heterologous RNA-binding domain. In some embodiments, the polypeptide may comprise a domain having 5′ to 3′ exonuclease activity (e.g., wherein the 5′ to 3′ exonuclease activity increases repair of the alteration of the target site, e.g., in favor of alteration over the original genomic sequence). In some embodiments, the polypeptide may comprise a domain having 3′ to 5′ exonuclease activity, e.g., proof-reading activity. In some embodiments, the writing domain, e.g., RT domain, has 3′ to 5′ exonuclease activity, e.g., proof-reading activity.


Template Nucleic Acids

The gene modifying systems described herein can modify a host target DNA site using a template nucleic acid sequence. In some embodiments, the gene modifying systems described herein transcribe an RNA sequence template into host target DNA sites by target-primed reverse transcription (TPRT). By modifying DNA sequence(s) via reverse transcription of the RNA sequence template directly into the host genome, the gene modifying system can insert an object sequence into a target genome without the need for exogenous DNA sequences to be introduced into the host cell (unlike, for example, CRISPR systems), as well as eliminate an exogenous DNA insertion step. The gene modifying system can also delete a sequence from the target genome or introduce a substitution using an object sequence. Therefore, the gene modifying system provides a platform for the use of customized RNA sequence templates containing object sequences, e.g., sequences comprising heterologous gene coding and/or function information.


In some embodiments, the template nucleic acid comprises one or more sequence (e.g., 2 sequences) that binds the gene modifying polypeptide.


In some embodiments a system or method described herein comprises a single template nucleic acid (e.g., template RNA). In some embodiments a system or method described herein comprises a plurality of template nucleic acids (e.g., template RNAs). For example, a system described herein comprises a first RNA comprising (e.g., from 5′ to 3′) a sequence that binds the gene modifying polypeptide (e.g., the DNA-binding domain and/or the endonuclease domain, e.g., a gRNA) and a sequence that binds a target site (e.g., a second strand of a site in a target genome), and a second RNA (e.g., a template RNA) comprising (e.g., from 5′ to 3′) optionally a sequence that binds the gene modifying polypeptide (e.g., that specifically binds the RT domain), a heterologous object sequence, and a PBS sequence. In some embodiments, when the system comprises a plurality of nucleic acids, each nucleic acid comprises a conjugating domain. In some embodiments, a conjugating domain enables association of nucleic acid molecules, e.g., by hybridization of complementary sequences. For example, in some embodiments a first RNA comprises a first conjugating domain and a second RNA comprises a second conjugating domain, and the first and second conjugating domains are capable of hybridizing to one another, e.g., under stringent conditions. In some embodiments, the stringent conditions for hybridization include hybridization in 4× sodium chloride/sodium citrate (SSC), at about 65 C, followed by a wash in 1×SSC, at about 65 C.


In some embodiments, the template nucleic acid comprises RNA. In some embodiments, the template nucleic acid comprises DNA (e.g., single stranded or double stranded DNA).


In some embodiments, the template nucleic acid comprises one or more (e.g., 2) homology domains that have homology to the target sequence. In some embodiments, the homology domains are about 10-20, 20-50, or 50-100 nucleotides in length.


In some embodiments, a template RNA can comprise a gRNA sequence, e.g., to direct the gene modifying polypeptide to a target site of interest. In some embodiments, a template RNA comprises (e.g., from 5′ to 3′) (i) optionally a gRNA spacer that binds a target site (e.g., a second strand of a site in a target genome), (ii) optionally a gRNA scaffold that binds a polypeptide described herein (e.g., a gene modifying polypeptide or a Cas polypeptide), (iii) a heterologous object sequence comprising a mutation region (optionally the heterologous object sequence comprises, from 5′ to 3′, a first homology region, a mutation region, and a second homology region), and (iv) a primer binding site (PBS) sequence comprising a 3′ target homology domain.


The template nucleic acid (e.g., template RNA) component of a genome editing system described herein typically is able to bind the gene modifying polypeptide of the system. In some embodiments the template nucleic acid (e.g., template RNA) has a 3′ region that is capable of binding a gene modifying polypeptide. The binding region, e.g., 3′ region, may be a structured RNA region, e.g., having at least 1, 2 or 3 hairpin loops, capable of binding the gene modifying polypeptide of the system. The binding region may associate the template nucleic acid (e.g., template RNA) with any of the polypeptide modules. In some embodiments, the binding region of the template nucleic acid (e.g., template RNA) may associate with an RNA-binding domain in the polypeptide. In some embodiments, the binding region of the template nucleic acid (e.g., template RNA) may associate with the reverse transcription domain of the gene modifying polypeptide (e.g., specifically bind to the RT domain). In some embodiments, the template nucleic acid (e.g., template RNA) may associate with the DNA binding domain of the polypeptide, e.g., a gRNA associating with a Cas9-derived DNA binding domain. In some embodiments, the binding region may also provide DNA target recognition, e.g., a gRNA hybridizing to the target DNA sequence and binding the polypeptide, e.g., a Cas9 domain. In some embodiments, the template nucleic acid (e.g., template RNA) may associate with multiple components of the polypeptide, e.g., DNA binding domain and reverse transcription domain.


In some embodiments the template RNA has a poly-A tail at the 3′ end. In some embodiments the template RNA does not have a poly-A tail at the 3′ end.


In some embodiments, the template nucleic acid is a template RNA. In some embodiments, the template RNA comprises one or more modified nucleotides. For example, in some embodiments, the template RNA comprises one or more deoxyribonucleotides. In some embodiments, regions of the template RNA are replaced by DNA nucleotides, e.g., to enhance stability of the molecule. For example, the 3′ end of the template may comprise DNA nucleotides, while the rest of the template comprises RNA nucleotides that can be reverse transcribed. For instance, in some embodiments, the heterologous object sequence is primarily or wholly made up of RNA nucleotides (e.g., at least 90%, 95%, 98%, or 99% RNA nucleotides). In some embodiments, the PBS sequence is primarily or wholly made up of DNA nucleotides (e.g., at least 90%, 95%, 98%, or 99% DNA nucleotides). In other embodiments, the heterologous object sequence for writing into the genome may comprise DNA nucleotides. In some embodiments, the DNA nucleotides in the template are copied into the genome by a domain capable of DNA-dependent DNA polymerase activity. In some embodiments, the DNA-dependent DNA polymerase activity is provided by a DNA polymerase domain in the polypeptide. In some embodiments, the DNA-dependent DNA polymerase activity is provided by a reverse transcriptase domain that is also capable of DNA-dependent DNA polymerization, e.g., second strand synthesis. In some embodiments, the template molecule is composed of only DNA nucleotides.


In some embodiments, a system described herein comprises two nucleic acids which together comprise the sequences of a template RNA described herein. In some embodiments, the two nucleic acids are associated with each other non-covalently, e.g., directly associated with each other (e.g., via base pairing), or indirectly associated as part of a complex comprising one or more additional molecule.


A template RNA described herein may comprise, from 5′ to 3′: (1) a gRNA spacer; (2) a gRNA scaffold; (3) heterologous object sequence (4) a primer binding site (PBS) sequence. Each of these components is now described in more detail.


gRNA spacer and gRNA scaffold


A template RNA described herein may comprise a gRNA spacer that directs the gene modifying system to a target nucleic acid, and a gRNA scaffold that promotes association of the template RNA with the Cas domain of the gene modifying polypeptide. The systems described herein can also comprise a gRNA that is not part of a template nucleic acid. For example, a gRNA that comprises a gRNA spacer and gRNA scaffold, but not a heterologous object sequence or a PBS sequence, can be used, e.g., to induce second strand nicking, e.g., as described in the section herein entitled “Second Strand Nicking”.


In some embodiments, the gRNA is a short synthetic RNA composed of a scaffold sequence that participates in CRISPR-associated protein binding and a user-defined −20 nucleotide targeting sequence for a genomic target. The structure of a complete gRNA was described by Nishimasu et al. Cell 156, P935-949 (2014). The gRNA (also referred to as sgRNA for single-guide RNA) consists of crRNA- and tracrRNA-derived sequences connected by an artificial tetraloop. The crRNA sequence can be divided into guide (20 nt) and repeat (12 nt) regions, whereas the tracrRNA sequence can be divided into anti-repeat (14 nt) and three tracrRNA stem loops (Nishimasu et al. Cell 156, P935-949 (2014)). In practice, guide RNA sequences are generally designed to have a length of between 17-24 nucleotides (e.g., 19, 20, or 21 nucleotides) and be complementary to a targeted nucleic acid sequence. Custom gRNA generators and algorithms are available commercially for use in the design of effective guide RNAs. In some embodiments, the gRNA comprises two RNA components from the native CRISPR system, e.g. crRNA and tracrRNA. As is well known in the art, the gRNA may also comprise a chimeric, single guide RNA (sgRNA) containing sequence from both a tracrRNA (for binding the nuclease) and at least one crRNA (to guide the nuclease to the sequence targeted for editing/binding). Chemically modified sgRNAs have also been demonstrated to be effective for use with CRISPR-associated proteins; see, for example, Hendel et al. (2015) Nature Biotechnol., 985-991. In some embodiments, a gRNA spacer comprises a nucleic acid sequence that is complementary to a DNA sequence associated with a target gene.


In some embodiments, the region of the template nucleic acid, e.g., template RNA, comprising the gRNA adopts an underwound ribbon-like structure of gRNA bound to target DNA (e.g., as described in Mulepati et al. Science 19 Sep. 2014:Vol. 345, Issue 6203, pp. 1479-1484). Without wishing to be bound by theory, this non-canonical structure is thought to be facilitated by rotation of every sixth nucleotide out of the RNA-DNA hybrid. Thus, in some embodiments, the region of the template nucleic acid, e.g., template RNA, comprising the gRNA may tolerate increased mismatching with the target site at some interval, e.g., every sixth base. In some embodiments, the region of the template nucleic acid, e.g., template RNA, comprising the gRNA comprising homology to the target site may possess wobble positions at a regular interval, e.g., every sixth base, that do not need to base pair with the target site.


In some embodiments, a Cas9 derivative with enhanced activity may be used in the gene modification polypeptide. In some embodiments, a Cas9 derivative may comprise mutations that improve activity of the HNH endonuclease domain, e.g., SpyCas9 R221K, N394K, or mutations that improve R-loop formation, e.g., SpyCas9 L1245V, or comprise a combination of such mutations, e.g., SpyCas9 R221K/N394K, SpyCas9 N394K/L1245V, SpyCas9 R221K/L1245V, or SpyCas9 R221K/N394K/L1245V (see, e.g., Spencer and Zhang Sci Rep 7:16836 (2017), the Cas9 derivatives and comprising mutations of which are incorporated herein by reference). In some embodiments, a Cas9 derivative may comprise one or more types of mutations described herein, e.g., PAM-modifying mutations, protein stabilizing mutations, activity enhancing mutations, and/or mutations partially or fully inactivating one or two endonuclease domains relative to the parental enzyme (e.g., one or more mutations to abolish endonuclease activity towards one or both strands of a target DNA, e.g., a nickase or catalytically dead enzyme). In some embodiments, a Cas9 enzyme used in a system described herein may comprise mutations that confer nickase activity toward the enzyme (e.g., SpyCas9 N863A or H840A) in addition to mutations improving catalytic efficiency (e.g., SpyCas9 R221K, N394K, and/or L1245V). In some embodiments, a Cas9 enzyme used in a system described herein is a SpyCas9 enzyme or derivative that further comprises an N863A mutation to confer nickase activity in addition to R221K and N394K mutations to improve catalytic efficiency.


In some embodiments, the template nucleic acid (e.g., template RNA) has at least 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 bases of at least 80%, 85%, 90%, 95%, 99%, or 100% homology to the target site, e.g., at the 5′ end, e.g., comprising a gRNA spacer sequence of length appropriate to the Cas9 domain of the gene modifying polypeptide (Table 8).


Table 12 provides parameters to define components for designing gRNA and/or Template RNAs to apply Cas variants listed in Table 8 for gene modifying. The cut site indicates the validated or predicted protospacer adjacent motif (PAM) requirements, validated or predicted location of cut site (relative to the most upstream base of the PAM site). The gRNA for a given enzyme can be assembled by concatenating the crRNA, Tetraloop, and tracrRNA sequences, and further adding a 5′ spacer of a length within Spacer (min) and Spacer (max) that matches a protospacer at a target site. Further, the predicted location of the ssDNA nick at the target is important for designing a PBS sequence of a Template RNA that can anneal to the sequence immediately 5′ of the nick in order to initiate target primed reverse transcription. In some embodiments, a gRNA scaffold described herein comprises a nucleic acid sequence comprising, in the 5′ to 3′ direction, a crRNA of Table 12, a tetraloop from the same row of Table 12, and a tracrRNA from the same row of Table 12, or a sequence having at least 70%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the gRNA or template RNA comprising the scaffold further comprises a gRNA spacer having a length within the Spacer (min) and Spacer (max) indicated in the same row of Table 12. In some embodiments, the gRNA or template RNA having a sequence according to Table 12 is comprised by a system that further comprises a gene modifying polypeptide, wherein the gene modifying polypeptide comprises a Cas domain described in the same row of Table 12.









TABLE 12







Parameters to define components for designing gRNA and/or Template RNAs


to apply Cas variants listed in Table 8 in gene modifying systems.





















Spacer
Spacer

SEQ ID


SEQ ID


Variant
PAM(s)
Cut
Tier
(min)
(max)
crRNA
NO:
Tetraloop
tracrRNA
NO:





Nme2Cas9
NNNNCC
-3
1
22
24
GTTGTAGC
10,051
GAAA
CGAAATGAGAACCGTTGCTACAATAAGGC
10,151








TCCCTTTCT


CGTCTGAAAAGATGTGCCGCAACGCTCTG









CATTTCG


CCCCTTAAAGCTTCTGCTTTAAGGGGCATC












GTTTA






PpnCas9
NNNNRTT

1
21
24
GTTGTAGC
10,052
GAAA
GCGAAATGAAAAACGTTGTTACAATAAGA
10,152








TCCCTTTTT


GATGAATTTCTCGCAAAGCTCTGCCTCTTG









CATTTCGC


AAATTTCGGTTTCAAGAGGCATC






SauCas9
NNGRR;
-3
1
21
23
GTTTTAGT
10,053
GAAA
CAGAATCTACTAAAACAAGGCAAAATGCC
10,153



NNGRRT




ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA






SauCas9-
NNNRR;
-3
1
21
21
GTTTTAGT
10,054
GAAA
CAGAATCTACTAAAACAAGGCAAAATGCC
10,154


KKH
NNNRRT




ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA






SauriCas9
NNGG
-3
1
21
21
GTTTTAGT
10,055
GAAA
CAGAATCTACTAAAACAAGGCAAAATGCC
10,155








ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA






SauriCas9-
NNRG
-3
1
21
21
GTTTTAGT
10,056
GAAA
CAGAATCTACTAAAACAAGGCAAAATGCC
10,156


KKH





ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA






ScaCas9-
NNG
-3
1
20
20
GTTTTAGA
10,057
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,157


Sc++





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9
NGG
-3
1
20
20
GTTTTAGA
10,058
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,158








GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9_
NGG
-3
1
20
20
GTTTTAGA
10,058
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,193


i_v1





GCTA


TCAACTTGGACTTCGGTCCAAGTGGCACC












GAGTCGGTGC






SpyCas9_
NGG
-3
1
20
20
GTTTTAGA
10,058
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,194


i_v2





GCTA


TCAACTTGGAGCTTGCTCCAAGTGGCACC












GAGTCGGTGC






SpyCas9_
NGG
-3
1
20
20
GTTTTAGA
10,058
GAAA
GTTTTAGAGCTAGAAATAGCAAGTTAAAA
10,195


i_v3





GCTA


TAAGGCTAGTCCGTTATCGACTTGAAAAA












GTCGCACCGAGTCGGTGC






SpyCas9-
NG
-3
1
20
20
GTTTTAGA
10,059
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,159


NG
(NGG =




GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT




NGA =







GC




NGT >












NGC)














SpyCas9-
NRN >
-3
1
20
20
GTTTTAGA
10,060
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,160


SpRY
NYN




GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






St1Cas9
NNAGAAW >
-3
1
20
20
GTCTTTGTA
10,061
GTAC
CAGAAGCTACAAAGATAAGGCTTCATGCC
10,161



NNAGGAW =




CTCTG


GAAATCAACACCCTGTCATTTTATGGCAG




NNGGAAW







GGTGTTTT






BlatCas9
NNNNCNAA >
-3
1
19
23
GCTATAGT
10,062
GAAA
GGTAAGTTGCTATAGTAAGGGCAACAGAC
10,162



NNNNCNDD >




TCCTTACT


CCGAGGCGTTGGGGATCGCCTAGCCCGTG




NNNNC







TTTACGGGCTCTCCCCATATTCAAAATAAT












GACAGACGAGCACCTTGGAGCATTTATCT












CCGAGGTGCT






cCas9-v16
NNVACT;
-3
2
21
21
GTCTTAGT
10,063
GAAA
CAGAATCTACTAAGACAAGGCAAAATGCC
10,163



NNVATGM;




ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA




NNVATT;












NNVGCT;












NNVGTG;












NNVGTT














cCas9-v17
NNVRRN
-3
2
21
21
GTCTTAGT
10,064
GAAA
CAGAATCTACTAAGACAAGGCAAAATGCC
10,164








ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA






cCas9-v21
NNVACT;
-3
2
21
21
GTCTTAGT
10,065
GAAA
CAGAATCTACTAAGACAAGGCAAAATGCC
10,165



NNVATGM;




ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA




NNVATT;












NNVGCT;












NNVGTG;












NNVGTT














cCas9-v42
NNVRRN
-3
2
21
21
GTCTTAGT
10,066
GAAA
CAGAATCTACTAAGACAAGGCAAAATGCC
10,166








ACTCTG


GTGTTTATCTCGTCAACTTGTTGGCGAGA






CdiCas9
NNRHHHY;

2
22
22
ACTGGGGT
10,067
GAAA
CTGAACCTCAGTAAGCATTGGCTCGTTTCC
10,167



NNRAAAY




TCAG


AATGTTGATTGCTCCGCCGGTGCTCCTTAT












TTTTAAGGGCGCCGGC






CjeCas9
NNNNRYAC
-3
2
21
23
GTTTTAGTC
10,068
GAAA
AGGGACTAAAATAAAGAGTTTGCGGGACT
10,168








CCT


CTGCGGGGTTACAATCCCCTAAAACCGC






GeoCas9
NNNNCRAA

2
21
23
GTCATAGT
10,069
GAAA
TCAGGGTTACTATGATAAGGGCTTTCTGCC
10,169








TCCCCTGA


TAAGGCAGACTGACCCGCGGCGTTGGGG












ATCGCCTGTCGCCCGCTTTTGGCGGGCATT












CCCCATCCTT






iSpy 
NAAN
-3
2
19
21
GTTTTAGA
10,070
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,170


MacCas9





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






NmeCas9
NNNNGAYT;
-3
2
20
24
GTTGTAGC
10,071
GAAA
CGAAATGAGAACCGTTGCTACAATAAGGC
10,171



NNNNGYTT;




TCCCTTTCT


CGTCTGAAAAGATGTGCCGCAACGCTCTG




NNNNGAYA;




CATTTCG


CCCCTTAAAGCTTCTGCTTTAAGGGGCATC




NNNNGTCT







GTTTA






ScaCas9
NNG
-3
2
20
20
GTTTTAGA
10,072
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,172








GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






ScaCas9-
NNG
-3
2
20
20
GTTTTAGA
10,073
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,173


HiFi-Sc++





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9-
NRRH
-3
2
20
20
GTTTAAGA
10,074
GAAA
CAGCATAGCAAGTTTAAATAAGGCTAGTC
10,174


3var-NRRH





GCTATGCT


CGTTATCAACTTGAAAAAGTGGCACCGAG









G


TCGGTGC






SpyCas9-
NRTH
-3
2
20
20
GTTTAAGA
10,075
GAAA
CAGCATAGCAAGTTTAAATAAGGCTAGTC
10,175


3var-NRTH





GCTATGCT


CGTTATCAACTTGAAAAAGTGGCACCGAG









G


TCGGTGC






SpyCas9-
NRCH
-3
2
20
20
GTTTAAGA
10,076
GAAA
CAGCATAGCAAGTTTAAATAAGGCTAGTC
10,176


3var-NRCH





GCTATGCT


CGTTATCAACTTGAAAAAGTGGCACCGAG









G


TCGGTGC






SpyCas9-
NGG
-3
2
20
20
GTTTTAGA
10,077
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,177


HF1





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9-
NAAG
-3
2
20
20
GTTTTAGA
10,078
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,178


QQR1





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9-
NGN
-3
2
20
20
GTTTTAGA
10,079
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,179


SpG





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9-
NGAN
-3
2
20
20
GTTTTAGA
10,080
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,180


VQR





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9-
NGCG
-3
2
20
20
GTTTTAGA
10,081
GAAA
TAGCAAGTTAAAATAAGGCTAGTCCGTTA
10,181


VRER





GCTA


TCAACTTGAAAAAGTGGCACCGAGTCGGT












GC






SpyCas9-
NG;GAA;
-3
2
20
20
GTTTAAGA
10,082
GAAA
CAGCATAGCAAGTTTAAATAAGGCTAGTC
10,182


xCas
GAT




GCTATGCT


CGTTATCAACTTGAAAAAGTGGCACCGAG









G


TCGGTGC






SpyCas9-
NG
-3
2
20
20
GTTTAAGA
10,083
GAAA
CAGCATAGCAAGTTTAAATAAGGCTAGTC
10,183


xCas-NG





GCTATGCT


CGTTATCAACTTGAAAAAGTGGCACCGAG









G


TCGGTGC






St1Cas9-
NNACAA
-3
2
20
20
GTCTTTGTA
10,084
GTAC
CAGAAGCTACAAAGATAAGGCTTCATGCC
10,184


CNRZ1066





CTCTG


GAAATCAACACCCTGTCATTTTATGGCAG












GGTGTTTT






St1Cas9-
NNGCAA
-3
2
20
20
GTCTTTGTA
10,085
GTAC
CAGAAGCTACAAAGATAAGGCTTCATGCC
10,185


LMG1831





CTCTG


GAAATCAACACCCTGTCATTTTATGGCAG












GGTGTTTT






St1Cas9-
NNAAAA
-3
2
20
20
GTCTTTGTA
10,086
GTAC
CAGAAGCTACAAAGATAAGGCTTCATGCC
10,186


MTH17CL396





CTCTG


GAAATCAACACCCTGTCATTTTATGGCAG












GGTGTTTT






St1Cas9-
NNGAAA
-3
2
20
20
GTCTTTGTA
10,087
GTAC
CAGAAGCTACAAAGATAAGGCTTCATGCC
10,187


TH1477





CTCTG


GAAATCAACACCCTGTCATTTTATGGCAG












GGTGTTTT






SRGN3.1
NNGG

1
21
23
GTTTTAGT
10,088
GAAA
CAGAATCTACTGAAACAAGACAATATGTC
10,188








ACTCTG


GTGTTTATCCCATCAATTTATTGGTGGGAT












TTT






sRGN3.3
NNGG

1
21
23
GTTTTAGT
10,089
GAAA
CAGAATCTACTGAAACAAGACAATATGTC
10,189








ACTCTG


GTGTTTATCCCATCAATTTATTGGTGGGAT












TTT









Herein, when an RNA sequence (e.g., a template RNA sequence) is said to comprise a particular sequence (e.g., a sequence of Table 12 or a portion thereof) that comprises thymine (T), it is of course understood that the RNA sequence may (and frequently does) comprise uracil (U) in place of T. For instance, the RNA sequence may comprise U at every position shown as T in the sequence in Table 12. More specifically, the present disclosure provides an RNA sequence according to every gRNA scaffold sequence of Table 12, wherein the RNA sequence has a U in place of each T in the sequence in Table 12. Additionally, it is understood that terminal Us and Ts may optionally be added or removed from tracrRNA sequences and may be modified or unmodified when provided as RNA. Without wishing to be bound by example, versions of gRNA scaffold sequences alternative to those exemplified in Table 12 may also function with the different Cas9 enzymes or derivatives thereof exemplified in Table 8, e.g., alternate gRNA scaffold sequences with nucleotide additions, substitutions, or deletions, e.g., sequences with stem-loop structures added or removed. It is contemplated herein that the gRNA scaffold sequences represent a component of gene modifying systems that can be similarly optimized for a given system, Cas-RT fusion polypeptide, indication, target mutation, template RNA, or delivery vehicle.


Heterologous Object Sequence

A template RNA described herein may comprise a heterologous object sequence that the gene modifying polypeptide can use as a template for reverse transcription, to write a desired sequence into the target nucleic acid. In some embodiments, the heterologous object sequence comprises, from 5′ to 3′, a post-edit homology region, the mutation region, and a pre-edit homology region. Without wishing to be bound by theory, an RT performing reverse transcription on the template RNA first reverse transcribes the pre-edit homology region, then the mutation region, and then the post-edit homology region, thereby creating a DNA strand comprising the desired mutation with a homology region on either side.


In some embodiments, the heterologous object sequence is at least 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 120, 140, 160, 180, 200, 500, or 1,000 nucleotides (nts) in length, or at least 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, or 10 kilobases in length. In some embodiments, the heterologous object sequence is no more than 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 120, 140, 160, 180, 200, 500, 1,000, or 2000 nucleotides (nts) in length, or no more than 20, 15, 10, 9, 8, 7, 6, 5, 4, or 3 kilobases in length. In some embodiments, the heterologous object sequence is 30-1000, 40-1000, 50-1000, 60-1000, 70-1000, 74-1000, 75-1000, 76-1000, 77-1000, 78-1000, 79-1000, 80-1000, 85-1000, 90-1000, 100-1000, 120-1000, 140-1000, 160-1000, 180-1000, 200-1000, 500-1000, 30-500, 40-500, 50-500, 60-500, 70-500, 74-500, 75-500, 76-500, 77-500, 78-500, 79-500, 80-500, 85-500, 90-500, 100-500, 120-500, 140-500, 160-500, 180-500, 200-500, 30-200, 40-200, 50-200, 60-200, 70-200, 74-200, 75-200, 76-200, 77-200, 78-200, 79-200, 80-200, 85-200, 90-200, 100-200, 120-200, 140-200, 160-200, 180-200, 30-100, 40-100, 50-100, 60-100, 70-100, 74-100, 75-100, 76-100, 77-100, 78-100, 79-100, 80-100, 85-100, or 90-100 nucleotides (nts) in length, or 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, 1-2, 2-20, 2-15, 2-10, 2-9, 2-8, 2-7, 2-6, 2-5, 2-4, 2-3, 3-20, 3-15, 3-10, 3-9, 3-8, 3-7, 3-6, 3-5, 3-4, 4-20, 4-15, 4-10, 4-9, 4-8, 4-7, 4-6, 4-5, 5-20, 5-15, 5-10, 5-9, 5-8, 5-7, 5-6, 6-20, 6-15, 6-10, 6-9, 6-8, 6-7, 7-20, 7-15, 7-10, 7-9, 7-8, 8-20, 8-15, 8-10, 8-9, 9-20, 9-15, 9-10, 10-15, 10-20, or 15-20 kilobases in length. In some embodiments, the heterologous object sequence is 10-100, 10-90, 10-80, 10-70, 10-60, 10-50, 10-40, 10-30, or 10-20 nt in length, e.g., 10-80, 10-50, or 10-20 nt in length, e.g., about 10-20 nt in length. In some embodiments, the heterologous object sequence is 8-30, 9-25, 10-20, 11-16, or 12-15 nucleotides in length, e.g., is 11-16 nt in length. Without wishing to be bound by theory, in some embodiments, a larger insertion size, larger region of editing (e.g., the distance between a first edit/substitution and a second edit/substitution in the target region), and/or greater number of desired edits (e.g., mismatches of the heterologous object sequence to the target genome), may result in a longer optimal heterologous object sequence.


In certain embodiments, the template nucleic acid comprises a customized RNA sequence template which can be identified, designed, engineered and constructed to contain sequences altering or specifying host genome function, for example by introducing a heterologous coding region into a genome; affecting or causing exon structure/alternative splicing, e.g., leading to exon skipping of one or more exons; causing disruption of an endogenous gene, e.g., creating a genetic knockout; causing transcriptional activation of an endogenous gene; causing epigenetic regulation of an endogenous DNA; causing up-regulation of one or more operably linked genes, e.g., leading to gene activation or overexpression; causing down-regulation of one or more operably linked genes, e.g., creating a genetic knock-down; etc. In certain embodiments, a customized RNA sequence template can be engineered to contain sequences coding for exons and/or transgenes, provide binding sites for transcription factor activators, repressors, enhancers, etc., and combinations thereof. In some embodiments, a customized template can be engineered to encode a nucleic acid or peptide tag to be expressed in an endogenous RNA transcript or endogenous protein operably linked to the target site. In other embodiments, the coding sequence can be further customized with splice donor sites, splice acceptor sites, or poly-A tails.


The template nucleic acid (e.g., template RNA) of the system typically comprises an object sequence (e.g., a heterologous object sequence) for writing a desired sequence into a target DNA. The object sequence may be coding or non-coding. The template nucleic acid (e.g., template RNA) can be designed to result in insertions, mutations, or deletions at the target DNA locus. In some embodiments, the template nucleic acid (e.g., template RNA) may be designed to cause an insertion in the target DNA. For example, the template nucleic acid (e.g., template RNA) may contain a heterologous sequence, wherein the reverse transcription will result in insertion of the heterologous sequence into the target DNA. In other embodiments, the RNA template may be designed to introduce a deletion into the target DNA. For example, the template nucleic acid (e.g., template RNA) may match the target DNA upstream and downstream of the desired deletion, wherein the reverse transcription will result in the copying of the upstream and downstream sequences from the template nucleic acid (e.g., template RNA) without the intervening sequence, e.g., causing deletion of the intervening sequence. In other embodiments, the template nucleic acid (e.g., template RNA) may be designed to introduce an edit into the target DNA. For example, the template RNA may match the target DNA sequence with the exception of one or more nucleotides, wherein the reverse transcription will result in the copying of these edits into the target DNA, e.g., resulting in mutations, e.g., transition or transversion mutations.


In some embodiments, writing of an object sequence into a target site results in the substitution of nucleotides, e.g., where the full length of the object sequence corresponds to a matching length of the target site with one or more mismatched bases. In some embodiments, a heterologous object sequence may be designed such that a combination of sequence alterations may occur, e.g., a simultaneous addition and deletion, addition and substitution, or deletion and substitution.


In some embodiments, the heterologous object sequence may contain an open reading frame or a fragment of an open reading frame. In some embodiments the heterologous object sequence has a Kozak sequence. In some embodiments the heterologous object sequence has an internal ribosome entry site. In some embodiments the heterologous object sequence has a self-cleaving peptide such as a T2A or P2A site. In some embodiments the heterologous object sequence has a start codon. In some embodiments the template RNA has a splice acceptor site. In some embodiments the template RNA has a splice donor site. Exemplary splice acceptor and splice donor sites are described in WO2016044416, incorporated herein by reference in its entirety. Exemplary splice acceptor site sequences are known to those of skill in the art. In some embodiments the template RNA has a microRNA binding site downstream of the stop codon. In some embodiments the template RNA has a polyA tail downstream of the stop codon of an open reading frame. In some embodiments the template RNA comprises one or more exons. In some embodiments the template RNA comprises one or more introns. In some embodiments the template RNA comprises a eukaryotic transcriptional terminator. In some embodiments the template RNA comprises an enhanced translation element or a translation enhancing element. In some embodiments the RNA comprises the human T-cell leukemia virus (HTLV-1) R region. In some embodiments the RNA comprises a posttranscriptional regulatory element that enhances nuclear export, such as that of Hepatitis B Virus (HPRE) or Woodchuck Hepatitis Virus (WPRE).


In some embodiments, the heterologous object sequence may contain a non-coding sequence. For example, the template nucleic acid (e.g., template RNA) may comprise a regulatory element, e.g., a promoter or enhancer sequence or miRNA binding site. In some embodiments, integration of the object sequence at a target site will result in upregulation of an endogenous gene. In some embodiments, integration of the object sequence at a target site will result in downregulation of an endogenous gene. In some embodiments the template nucleic acid (e.g., template RNA) comprises a tissue specific promoter or enhancer, each of which may be unidirectional or bidirectional. In some embodiments the promoter is an RNA polymerase I promoter, RNA polymerase II promoter, or RNA polymerase III promoter. In some embodiments the promoter comprises a TATA element. In some embodiments the promoter comprises a B recognition element. In some embodiments the promoter has one or more binding sites for transcription factors.


In some embodiments, the template nucleic acid (e.g., template RNA) comprises a site that coordinates epigenetic modification. In some embodiments, the template nucleic acid (e.g., template RNA) comprises a chromatin insulator. For example, the template nucleic acid (e.g., template RNA) comprises a CTCF site or a site targeted for DNA methylation.


In some embodiments, the template nucleic acid (e.g., template RNA) comprises a gene expression unit composed of at least one regulatory region operably linked to an effector sequence. The effector sequence may be a sequence that is transcribed into RNA (e.g., a coding sequence or a non-coding sequence such as a sequence encoding a micro RNA).


In some embodiments, the heterologous object sequence of the template nucleic acid (e.g., template RNA) is inserted into a target genome in an endogenous intron. In some embodiments, the heterologous object sequence of the template nucleic acid (e.g., template RNA) is inserted into a target genome and thereby acts as a new exon. In some embodiments, the insertion of the heterologous object sequence into the target genome results in replacement of a natural exon or the skipping of a natural exon.


The template nucleic acid (e.g., template RNA) can be designed to result in insertions, mutations, or deletions at the target DNA locus. In some embodiments, the template nucleic acid (e.g., template RNA) may be designed to cause an insertion in the target DNA. For example, the template nucleic acid (e.g., template RNA) may contain a heterologous object sequence, wherein the reverse transcription will result in insertion of the heterologous object sequence into the target DNA. In other embodiments, the RNA template may be designed to write a deletion into the target DNA. For example, the template nucleic acid (e.g., template RNA) may match the target DNA upstream and downstream of the desired deletion, wherein the reverse transcription will result in the copying of the upstream and downstream sequences from the template nucleic acid (e.g., template RNA) without the intervening sequence, e.g., causing deletion of the intervening sequence. In other embodiments, the template nucleic acid (e.g., template RNA) may be designed to write an edit into the target DNA. For example, the template RNA may match the target DNA sequence with the exception of one or more nucleotides, wherein the reverse transcription will result in the copying of these edits into the target DNA, e.g., resulting in mutations, e.g., transition or transversion mutations.


In some embodiments, the pre-edit homology domain comprises a nucleic acid sequence having 100% sequence identity with a nucleic acid sequence comprised in a target nucleic acid molecule.


In some embodiments, the post-edit homology domain comprises a nucleic acid sequence having 100% sequence identity with a nucleic acid sequence comprised in a target nucleic acid molecule.


PBS Sequence

In some embodiments, a template nucleic acid (e.g., template RNA) comprises a PBS sequence. In some embodiments, a PBS sequence is disposed 3′ of the heterologous object sequence and is complementary to a sequence adjacent to a site to be modified by a system described herein, or comprises no more than 1, 2, 3, 4, or 5 mismatches to a sequence complementary to the sequence adjacent to a site to be modified by the system/gene modifying polypeptide. In some embodiments, the PBS sequence binds within 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides of a nick site in the target nucleic acid molecule. In some embodiments, binding of the PBS sequence to the target nucleic acid molecule permits initiation of target-primed reverse transcription (TPRT), e.g., with the 3′ homology domain acting as a primer for TPRT. In some embodiments, the PBS sequence is 3-5, 5-10, 10-30, 10-25, 10-20, 10-19, 10-18, 10-17, 10-16, 10-15, 10-14, 10-13, 10-12, 10-11, 11-30, 11-25, 11-20, 11-19, 11-18, 11-17, 11-16, 11-15, 11-14, 11-13, 11-12, 12-30, 12-25, 12-20, 12-19, 12-18, 12-17, 12-16, 12-15, 12-14, 12-13, 13-30, 13-25, 13-20, 13-19, 13-18, 13-17, 13-16, 13-15, 13-14, 14-30, 14-25, 14-20, 14-19, 14-18, 14-17, 14-16, 14-15, 15-30, 15-25, 15-20, 15-19, 15-18, 15-17, 15-16, 16-30, 16-25, 16-20, 16-19, 16-18, 16-17, 17-30, 17-25, 17-20, 17-19, 17-18, 18-30, 18-25, 18-20, 18-19, 19-30, 19-25, 19-20, 20-30, 20-25, or 25-30 nucleotides in length, e.g., 10-17, 12-16, or 12-14 nucleotides in length. In some embodiments, the PBS sequence is 5-20, 8-16, 8-14, 8-13, 9-13, 9-12, or 10-12 nucleotides in length, e.g., 9-12 nucleotides in length.


The template nucleic acid (e.g., template RNA) may have some homology to the target DNA. In some embodiments, the template nucleic acid (e.g., template RNA) PBS sequence domain may serve as an annealing region to the target DNA, such that the target DNA is positioned to prime the reverse transcription of the template nucleic acid (e.g., template RNA). In some embodiments the template nucleic acid (e.g., template RNA) has at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 175, 200 or more bases of exact homology to the target DNA at the 3′ end of the RNA. In some embodiments the template nucleic acid (e.g., template RNA) has at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 175, 200 or more bases of at least 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% homology to the target DNA, e.g., at the 5′ end of the template nucleic acid (e.g., template RNA).


Exemplary Template Sequences

In some embodiments of the systems and methods herein, the template RNA comprises a gRNA spacer comprising the core nucleotides of a gRNA spacer sequence of Table 1. In some embodiments, the gRNA spacer additionally comprises one or more (e.g., 2, 3, or all) consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the gRNA spacer. In some embodiments, the template RNA comprising a sequence of Table 1 is comprised by a system that further comprises a gene modifying polypeptide having an RT domain listed in the same line of Table 1. RT domain amino acid sequences can be found, e.g., in Table 6 herein.









TABLE 1







Exemplary gRNA spacer Cas pairs


Table 1 provides a gRNA database for correcting the pathogenic E342K


mutation in SERPINA1. List of spacers, PAMs, and Cas variants for


generating a nick at an appropriate position to enable installation


of a desired genomic edit with a gene modifying system. The spacers


in this table are designed to be used with a gene modifying


polypeptide comprising a nickase variant of the Cas species


indicated in the table. Tables 2, 3, and 4 detail the other


components of the system and are organized such that the ID number


shown here in Column 1 (“ID”) is meant to correspond to the same ID


number in the subsequent tables.














Pam

SEQ


overlaps


ID
sequence
gRNA spacer
ID NO
Cas species
distance
mutation





   1
AAAGG
GCTGTGCTGACCATCGACAAG
17085
SauCas9KKH
  0
0





   2
AAAG
GCTGTGCTGACCATCGACAAG
17086
SauriCas9-
  0
0






KKH







   3
AAAG
CTGTGCTGACCATCGACAAG
17087
SpyCas9-
  0
0






QQR1







   4
AAAG
gcTGTGCTGACCATCGACAAG
17088
iSpyMacCas9
  0
0





   5
AAA
CTGTGCTGACCATCGACAAG
17089
SpyCas9-
  0
0






SpRY







   6
AAAGGG
GCTGTGCTGACCATCGACAAG
17090
cCas9-v17
  0
0





   7
AAAGGG
GCTGTGCTGACCATCGACAAG
17091
cCas9-v42
  0
0





   8
GAAAG
GGCTGTGCTGACCATCGACAA
17092
SauCas9KKH
  1
0





   9
TCG
CAGCTTCAGTCCCTTTCTTG
17093
ScaCas9
  1
0





  10
TCG
CAGCTTCAGTCCCTTTCTTG
17094
ScaCas9-HiFi-
  1
0






Sc++







  11
TCG
CAGCTTCAGTCCCTTTCTTG
17095
ScaCas9-
  1
0






Sc++







  12
TCG
CAGCTTCAGTCCCTTTCTTG
17096
SpyCas9-
  1
0






SpRY







  13
GAA
GCTGTGCTGACCATCGACAA
17097
SpyCas9-
  1
0






SpRY







  14
GAA
GCTGTGCTGACCATCGACAA
17098
SpyCas9-
  1
0






xCas







  15
GAAAGG
GGCTGTGCTGACCATCGACAA
17099
cCas9-v17
  1
0





  16
GAAAGG
GGCTGTGCTGACCATCGACAA
17100
cCas9-v42
  1
0





  17
GAAA
GCTGTGCTGACCATCGACAA
17101
SpyCas9-
  1
0






3var-NRRH







  18
GAAA
ggCTGTGCTGACCATCGACAA
17102
iSpyMacCas9
  1
0





  19
AGAAA
AGGCTGTGCTGACCATCGACA
17103
SauCas9KKH
  2
0





  20
GTCGA
AGCAGCTTCAGTCCCTTTCTT
17104
SauCas9KKH
  2
0





  21
GTCGAT
AGCAGCTTCAGTCCCTTTCTT
17105
SauCas9KKH
  2
0





  22
GTCGAT
AGCAGCTTCAGTCCCTTTCTT
17106
cCas9-v17
  2
0





  23
GTCGAT
AGCAGCTTCAGTCCCTTTCTT
17107
cCas9-v42
  2
0





  24
AG
GGCTGTGCTGACCATCGACA
17108
SpyCas9-NG
  2
0





  25
AG
GGCTGTGCTGACCATCGACA
17109
SpyCas9-
  2
0






xCas







  26
AG
GGCTGTGCTGACCATCGACA
17110
SpyCas9-
  2
0






xCas-NG







  27
AGA
GGCTGTGCTGACCATCGACA
17111
SpyCas9-SpG
  2
0





  28
AGA
GGCTGTGCTGACCATCGACA
17112
SpyCas9-
  2
0






SpRY







  29
GTC
GCAGCTTCAGTCCCTTTCTT
17113
SpyCas9-
  2
0






SpRY







  30
AGAAAG
AGGCTGTGCTGACCATCGACA
17114
cCas9-v17
  2
0





  31
AGAAAG
AGGCTGTGCTGACCATCGACA
17115
cCas9-v42
  2
0





  32
AGAA
GGCTGTGCTGACCATCGACA
17116
SpyCas9-
  2
0






3var-NRRH







  33
AGAA
GGCTGTGCTGACCATCGACA
17117
SpyCas9-VQR
  2
0





  34
aAGAA
atAAGGCTGTGCTGACCATCGAC
17118
SauCas9
  3
1





  35
aAGAA
AAGGCTGTGCTGACCATCGAC
17119
SauCas9KKH
  3
1





  36
aAG
AGGCTGTGCTGACCATCGAC
17120
ScaCas9
  3
1





  37
aAG
AGGCTGTGCTGACCATCGAC
17121
ScaCas9-HiFi-
  3
1






Sc++







  38
aAG
AGGCTGTGCTGACCATCGAC
17122
ScaCas9-
  3
1






Sc++







  39
aAG
AGGCTGTGCTGACCATCGAC
17123
SpyCas9-
  3
1






SpRY







  40
tG
AGCAGCTTCAGTCCCTTTCT
17124
SpyCas9-NG
  3
1





  41
tG
AGCAGCTTCAGTCCCTTTCT
17125
SpyCas9-
  3
1






xCas







  42
tG
AGCAGCTTCAGTCCCTTTCT
17126
SpyCas9-
  3
1






xCas-NG







  43
tGT
AGCAGCTTCAGTCCCTTTCT
17127
SpyCas9-SpG
  3
1





  44
tGT
AGCAGCTTCAGTCCCTTTCT
17128
SpyCas9-
  3
1






SpRY







  45
aAGAAA
AGGCTGTGCTGACCATCGAC
17129
St1Cas9-
  3
1






TH1477







  46
aAGAAA
AAGGCTGTGCTGACCATCGAC
17130
cCas9-v17
  3
1





  47
aAGAAA
AAGGCTGTGCTGACCATCGAC
17131
cCas9-v42
  3
1





  48
aAGA
AGGCTGTGCTGACCATCGAC
17132
SpyCas9-
  3
1






3var-NRRH







  49
tGTC
AGCAGCTTCAGTCCCTTTCT
17133
SpyCas9-
  3
1






3var-NRTH







  50
CaAGA
TAAGGCTGTGCTGACCATCGA
17134
SauCas9KKH
  4
1





  51
CaAG
TAAGGCTGTGCTGACCATCGA
17135
SauriCas9-
  4
1






KKH







  52
CaAG
AAGGCTGTGCTGACCATCGA
17136
SpyCas9-
  4
1






QQR1







  53
CaAG
taAGGCTGTGCTGACCATCGA
17137
iSpyMacCas9
  4
1





  54
TtG
CAGCAGCTTCAGTCCCTTTC
17138
ScaCas9
  4
1





  55
TtG
CAGCAGCTTCAGTCCCTTTC
17139
ScaCas9-HiFi-
  4
1






Sc++







  56
TtG
CAGCAGCTTCAGTCCCTTTC
17140
ScaCas9-
  4
1






Sc++







  57
TtG
CAGCAGCTTCAGTCCCTTTC
17141
SpyCas9-
  4
1






SpRY







  58
CaA
AAGGCTGTGCTGACCATCGA
17142
SpyCas9-
  4
1






SpRY







  59
CaAGAAA
AAGGCTGTGCTGACCATCGA
17143
St1Cas9
  4
1





  60
TtGTCGAT
ccccAGCAGCTTCAGTCCCTTTC
17144
BlatCas9
  4
1





  61
TtGTC
ccccAGCAGCTTCAGTCCCTTTC
17145
BlatCas9
  4
1





  62
CaAGAA
TAAGGCTGTGCTGACCATCGA
17146
cCas9-v17
  4
1





  63
CaAGAA
TAAGGCTGTGCTGACCATCGA
17147
cCas9-v42
  4
1





  64
ACaAG
ATAAGGCTGTGCTGACCATCG
17148
SauCas9KKH
  5
1





  65
CTt
CCAGCAGCTTCAGTCCCTTT
17149
SpyCas9-
  5
1






SpRY







  66
ACa
TAAGGCTGTGCTGACCATCG
17150
SpyCas9-
  5
1






SpRY







  67
ACaAGA
ATAAGGCTGTGCTGACCATCG
17151
cCas9-v17
  5
1





  68
ACaAGA
ATAAGGCTGTGCTGACCATCG
17152
cCas9-v42
  5
1





  69
GACaA
CATAAGGCTGTGCTGACCATC
17153
SauCas9KKH
  6
1





  70
GAC
ATAAGGCTGTGCTGACCATC
17154
SpyCas9-
  6
0






SpRY







  71
TCT
CCCAGCAGCTTCAGTCCCTT
17155
SpyCas9-
  6
0






SpRY







  72
GACaAG
CATAAGGCTGTGCTGACCATC
17156
cCas9-v17
  6
1





  73
GACaAG
CATAAGGCTGTGCTGACCATC
17157
cCas9-v42
  6
1





  74
GACa
ATAAGGCTGTGCTGACCATC
17158
SpyCas9-
  6
1






3var-NRCH







  75
CG
CATAAGGCTGTGCTGACCAT
17159
SpyCas9-NG
  7
0





  76
CG
CATAAGGCTGTGCTGACCAT
17160
SpyCas9-
  7
0






xCas







  77
CG
CATAAGGCTGTGCTGACCAT
17161
SpyCas9-
  7
0






xCas-NG







  78
CGA
CATAAGGCTGTGCTGACCAT
17162
SpyCas9-SpG
  7
0





  79
CGA
CATAAGGCTGTGCTGACCAT
17163
SpyCas9-
  7
0






SpRY







  80
TTC
CCCCAGCAGCTTCAGTCCCT
17164
SpyCas9-
  7
0






SpRY







  81
CGAC
CATAAGGCTGTGCTGACCAT
17165
SpyCas9-
  7
0






3var-NRRH







  82
CGAC
CATAAGGCTGTGCTGACCAT
17166
SpyCas9-VQR
  7
0





  83
CGACaA
CATAAGGCTGTGCTGACCAT
17167
St1Cas9-
  7
1






CNRZ1066







  84
TCG
GCATAAGGCTGTGCTGACCA
17168
ScaCas9
  8
0





  85
TCG
GCATAAGGCTGTGCTGACCA
17169
ScaCas9-HiFi-
  8
0






Sc++







  86
TCG
GCATAAGGCTGTGCTGACCA
17170
ScaCas9-
  8
0






Sc++







  87
TCG
GCATAAGGCTGTGCTGACCA
17171
SpyCas9-
  8
0






SpRY







  88
TTT
GCCCCAGCAGCTTCAGTCCC
17172
SpyCas9-
  8
0






SpRY







  89
TCGACaAG
cgtgCATAAGGCTGTGCTGACCA
17173
BlatCas9
  8
1





  90
TCGAC
cgtgCATAAGGCTGTGCTGACCA
17174
BlatCas9
  8
0





  91
ATCGA
GTGCATAAGGCTGTGCTGACC
17175
SauCas9KKH
  9
0





  92
CTT
GGCCCCAGCAGCTTCAGTCC
17176
SpyCas9-
  9
0






SpRY







  93
ATC
TGCATAAGGCTGTGCTGACC
17177
SpyCas9-
  9
0






SpRY







  94
CTTTCTtG
catgGCCCCAGCAGCTTCAGTCC
17178
BlatCas9
  9
1





  95
CTTTC
catgGCCCCAGCAGCTTCAGTCC
17179
BlatCas9
  9
0





  96
ATCGAC
GTGCATAAGGCTGTGCTGACC
17180
cCas9-v17
  9
0





  97
ATCGAC
GTGCATAAGGCTGTGCTGACC
17181
cCas9-v42
  9
0





  98
CAT
GTGCATAAGGCTGTGCTGAC
17182
SpyCas9-
 10
0






SpRY







  99
CCT
TGGCCCCAGCAGCTTCAGTC
17183
SpyCas9-
 10
0






SpRY







 100
CATCGACa
ggccGTGCATAAGGCTGTGCTGAC
17184
NmeCas9
 10
1





 101
CATC
GTGCATAAGGCTGTGCTGAC
17185
SpyCas9-
 10
0






3var-NRTH







 102
CCC
ATGGCCCCAGCAGCTTCAGT
17186
SpyCas9-
 11
0






SpRY







 103
CCA
CGTGCATAAGGCTGTGCTGA
17187
SpyCas9-
 11
0






SpRY







 104
CCATC
ggccGTGCATAAGGCTGTGCTGA
17188
BlatCas9
 11
0





 105
TCC
CATGGCCCCAGCAGCTTCAG
17189
SpyCas9-
 12
0






SpRY







 106
ACC
CCGTGCATAAGGCTGTGCTG
17190
SpyCas9-
 12
0






SpRY







 107
GAC
GCCGTGCATAAGGCTGTGCT
17191
SpyCas9-
 13
0






SpRY







 108
GTC
ACATGGCCCCAGCAGCTTCA
17192
SpyCas9-
 13
0






SpRY







 109
GTCCCTTT
aaaaCATGGCCCCAGCAGCTTCA
17193
BlatCas9
 13
0





 110
GTCCC
aaaaCATGGCCCCAGCAGCTTCA
17194
BlatCas9
 13
0





 111
GACC
GCCGTGCATAAGGCTGTGCT
17195
SpyCas9-
 13
0






3var-NRCH







 112
AGTCCC
taAAAACATGGCCCCAGCAGCTTC
17196
Nme2Cas9
 14
0





 113
AG
AACATGGCCCCAGCAGCTTC
17197
SpyCas9-NG
 14
0





 114
AG
AACATGGCCCCAGCAGCTTC
17198
SpyCas9-
 14
0






xCas







 115
AG
AACATGGCCCCAGCAGCTTC
17199
SpyCas9-
 14
0






xCas-NG







 116
TG
GGCCGTGCATAAGGCTGTGC
17200
SpyCas9-NG
 14
0





 117
TG
GGCCGTGCATAAGGCTGTGC
17201
SpyCas9-
 14
0






xCas







 118
TG
GGCCGTGCATAAGGCTGTGC
17202
SpyCas9-
 14
0






xCas-NG







 119
AGT
AACATGGCCCCAGCAGCTTC
17203
SpyCas9-SpG
 14
0





 120
AGT
AACATGGCCCCAGCAGCTTC
17204
SpyCas9-
 14
0






SpRY







 121
TGA
GGCCGTGCATAAGGCTGTGC
17205
SpyCas9-SpG
 14
0





 122
TGA
GGCCGTGCATAAGGCTGTGC
17206
SpyCas9-
 14
0






SpRY







 123
AGTCCCTT
aaaaACATGGCCCCAGCAGCTTC
17207
BlatCas9
 14
0





 124
AGTCC
aaaaACATGGCCCCAGCAGCTTC
17208
BlatCas9
 14
0





 125
TGACC
ccagGCCGTGCATAAGGCTGTGC
17209
BlatCas9
 14
0





 126
TGACCAT
CAGGCCGTGCATAAGGCTGTGC
17210
CdiCas9
 14
0





 127
TGAC
GGCCGTGCATAAGGCTGTGC
17211
SpyCas9-
 14
0






3var-NRRH







 128
TGAC
GGCCGTGCATAAGGCTGTGC
17212
SpyCas9-VQR
 14
0





 129
AGTC
AACATGGCCCCAGCAGCTTC
17213
SpyCas9-
 14
0






3var-NRTH







 130
CAGTCC
ctAAAAACATGGCCCCAGCAGCTT
17214
Nme2Cas9
 15
0





 131
CTGACC
ctCCAGGCCGTGCATAAGGCTGTG
17215
Nme2Cas9
 15
0





 132
CAG
AAACATGGCCCCAGCAGCTT
17216
ScaCas9
 15
0





 133
CAG
AAACATGGCCCCAGCAGCTT
17217
ScaCas9-HiFi-
 15
0






Sc++







 134
CAG
AAACATGGCCCCAGCAGCTT
17218
ScaCas9-
 15
0






Sc++







 135
CAG
AAACATGGCCCCAGCAGCTT
17219
SpyCas9-
 15
0






SpRY







 136
CTG
AGGCCGTGCATAAGGCTGTG
17220
ScaCas9
 15
0





 137
CTG
AGGCCGTGCATAAGGCTGTG
17221
ScaCas9-HiFi-
 15
0






Sc++







 138
CTG
AGGCCGTGCATAAGGCTGTG
17222
ScaCas9-
 15
0






Sc++







 139
CTG
AGGCCGTGCATAAGGCTGTG
17223
SpyCas9-
 15
0






SpRY







 140
CTGACCAT
tccaGGCCGTGCATAAGGCTGTG
17224
BlatCas9
 15
0





 141
CAGTC
taaaAACATGGCCCCAGCAGCTT
17225
BlatCas9
 15
0





 142
CTGAC
tccaGGCCGTGCATAAGGCTGTG
17226
BlatCas9
 15
0





 143
CAGTCCC
AAAAACATGGCCCCAGCAGCTT
17227
CdiCas9
 15
0





 144
CAGT
AAACATGGCCCCAGCAGCTT
17228
SpyCas9-
 15
0






3var-NRRH







 145
GCTGA
CCAGGCCGTGCATAAGGCTGT
17229
SauCas9KKH
 16
0





 146
TCAG
AAAAACATGGCCCCAGCAGCT
17230
SauriCas9-
 16
0






KKH







 147
TCA
AAAACATGGCCCCAGCAGCT
17231
SpyCas9-
 16
0






SpRY







 148
GCT
CAGGCCGTGCATAAGGCTGT
17232
SpyCas9-
 16
0






SpRY







 149
TTCAG
TAAAAACATGGCCCCAGCAGC
17233
SauCas9KKH
 17
0





 150
TTCAGT
TAAAAACATGGCCCCAGCAGC
17234
SauCas9KKH
 17
0





 151
TTCAGT
TAAAAACATGGCCCCAGCAGC
17235
cCas9-v17
 17
0





 152
TTCAGT
TAAAAACATGGCCCCAGCAGC
17236
cCas9-v42
 17
0





 153
TG
CCAGGCCGTGCATAAGGCTG
17237
SpyCas9-NG
 17
0





 154
TG
CCAGGCCGTGCATAAGGCTG
17238
SpyCas9-
 17
0






xCas







 155
TG
CCAGGCCGTGCATAAGGCTG
17239
SpyCas9-
 17
0






xCas-NG







 156
TGC
CCAGGCCGTGCATAAGGCTG
17240
SpyCas9-SpG
 17
0





 157
TGC
CCAGGCCGTGCATAAGGCTG
17241
SpyCas9-
 17
0






SpRY







 158
TTC
AAAAACATGGCCCCAGCAGC
17242
SpyCas9-
 17
0






SpRY







 159
TGCT
CCAGGCCGTGCATAAGGCTG
17243
SpyCas9-
 17
0






3var-NRCH







 160
GTG
TCCAGGCCGTGCATAAGGCT
17244
ScaCas9
 18
0





 161
GTG
TCCAGGCCGTGCATAAGGCT
17245
ScaCas9-HiFi-
 18
0






Sc++







 162
GTG
TCCAGGCCGTGCATAAGGCT
17246
ScaCas9-
 18
0






Sc++







 163
GTG
TCCAGGCCGTGCATAAGGCT
17247
SpyCas9-
 18
0






SpRY







 164
CTT
TAAAAACATGGCCCCAGCAG
17248
SpyCas9-
 18
0






SpRY







 165
TG
CTCCAGGCCGTGCATAAGGC
17249
SpyCas9-NG
 19
0





 166
TG
CTCCAGGCCGTGCATAAGGC
17250
SpyCas9-
 19
0






xCas







 167
TG
CTCCAGGCCGTGCATAAGGC
17251
SpyCas9-
 19
0






xCas-NG







 168
TGT
CTCCAGGCCGTGCATAAGGC
17252
SpyCas9-SpG
 19
0





 169
TGT
CTCCAGGCCGTGCATAAGGC
17253
SpyCas9-
 19
0






SpRY







 170
GCT
CTAAAAACATGGCCCCAGCA
17254
SpyCas9-
 19
0






SpRY







 171
GCTTCAGT
cctcTAAAAACATGGCCCCAGCA
17255
BlatCas9
 19
0





 172
TGTGCTGA
ccccTCCAGGCCGTGCATAAGGC
17256
BlatCas9
 19
0





 173
GCTTC
cctcTAAAAACATGGCCCCAGCA
17257
BlatCas9
 19
0





 174
TGTGC
ccccTCCAGGCCGTGCATAAGGC
17258
BlatCas9
 19
0





 175
CTG
CCTCCAGGCCGTGCATAAGG
17259
ScaCas9
 20
0





 176
CTG
CCTCCAGGCCGTGCATAAGG
17260
ScaCas9-HiFi-
 20
0






Sc++







 177
CTG
CCTCCAGGCCGTGCATAAGG
17261
ScaCas9
 20
0






Sc++







 178
CTG
CCTCCAGGCCGTGCATAAGG
17262
SpyCas9-
 20
0






SpRY







 179
AG
TCTAAAAACATGGCCCCAGC
17263
SpyCas9-NG
 20
0





 180
AG
TCTAAAAACATGGCCCCAGC
17264
SpyCas9-
 20
0






xCas







 181
AG
TCTAAAAACATGGCCCCAGC
17265
SpyCas9-
 20
0






xCas-NG







 182
AGC
TCTAAAAACATGGCCCCAGC
17266
SpyCas9-SpG
 20
0





 183
AGC
TCTAAAAACATGGCCCCAGC
17267
SpyCas9-
 20
0






SpRY







 184
AGCT
TCTAAAAACATGGCCCCAGC
17268
SpyCas9-
 20
0






3var-NRCH







 185
CAG
CTCTAAAAACATGGCCCCAG
17269
ScaCas9
 21
0





 186
CAG
CTCTAAAAACATGGCCCCAG
17270
ScaCas9-HiFi-
 21
0






Sc++







 187
CAG
CTCTAAAAACATGGCCCCAG
17271
ScaCas9-
 21
0






Sc++







 188
CAG
CTCTAAAAACATGGCCCCAG
17272
SpyCas9-
 21
0






SpRY







 189
GCT
CCCTCCAGGCCGTGCATAAG
17273
SpyCas9-
 21
0






SpRY







 190
CAGCTTC
GCCTCTAAAAACATGGCCCCAG
17274
CdiCas9
 21
0





 191
CAGC
CTCTAAAAACATGGCCCCAG
17275
SpyCas9-
 21
0






3var-NRRH







 192
GCAG
GCCTCTAAAAACATGGCCCCA
17276
SauriCas9-
 22
0






KKH







 193
GG
CCCCTCCAGGCCGTGCATAA
17277
SpyCas9-NG
 22
0





 194
GG
CCCCTCCAGGCCGTGCATAA
17278
SpyCas9-
 22
0






xCas







 195
GG
CCCCTCCAGGCCGTGCATAA
17279
SpyCas9-
 22
0






xCas-NG







 196
GGC
CCCCTCCAGGCCGTGCATAA
17280
SpyCas9-SpG
 22
0





 197
GGC
CCCCTCCAGGCCGTGCATAA
17281
SpyCas9-
 22
0






SpRY







 198
GCA
CCTCTAAAAACATGGCCCCA
17282
SpyCas9-
 22
0






SpRY







 199
GCAGC
tggcCTCTAAAAACATGGCCCCA
17283
BlatCas9
 22
0





 200
GCAGCT
GCCTCTAAAAACATGGCCCCA
17284
cCas9-v16
 22
0





 201
GCAGCT
GCCTCTAAAAACATGGCCCCA
17285
cCas9-v21
 22
0





 202
GGCT
CCCCTCCAGGCCGTGCATAA
17286
SpyCas9-
 22
0






3var-NRCH







 203
AGCAG
GGCCTCTAAAAACATGGCCCC
17287
SauCas9KKH
 23
0





 204
AGG
TCCCCTCCAGGCCGTGCATA
17288
ScaCas9
 23
0





 205
AGG
TCCCCTCCAGGCCGTGCATA
17289
ScaCas9-HiFi-
 23
0






Sc++







 206
AGG
TCCCCTCCAGGCCGTGCATA
17290
ScaCas9-
 23
0






Sc++







 207
AGG
TCCCCTCCAGGCCGTGCATA
17291
SpyCas9
 23
0





 208
AGG
TCCCCTCCAGGCCGTGCATA
17292
SpyCas9-HF1
 23
0





 209
AGG
TCCCCTCCAGGCCGTGCATA
17293
SpyCas9-SpG
 23
0





 210
AGG
TCCCCTCCAGGCCGTGCATA
17294
SpyCas9-
 23
0






SpRY







 211
AG
GCCTCTAAAAACATGGCCCC
17295
SpyCas9-NG
 23
0





 212
AG
GCCTCTAAAAACATGGCCCC
17296
SpyCas9-
 23
0






xCas







 213
AG
GCCTCTAAAAACATGGCCCC
17297
SpyCas9-
 23
0






xCas-NG







 214
AG
TCCCCTCCAGGCCGTGCATA
17298
SpyCas9-NG
 23
0





 215
AG
TCCCCTCCAGGCCGTGCATA
17299
SpyCas9-
 23
0






xCas







 216
AG
TCCCCTCCAGGCCGTGCATA
17300
SpyCas9-
 23
0






xCas-NG







 217
AGC
GCCTCTAAAAACATGGCCCC
17301
SpyCas9-SpG
 23
0





 218
AGC
GCCTCTAAAAACATGGCCCC
17302
SpyCas9-
 23
0






SpRY







 219
AGCAGC
GGCCTCTAAAAACATGGCCCC
17303
cCas9-v17
 23
0





 220
AGCAGC
GGCCTCTAAAAACATGGCCCC
17304
cCas9-v42
 23
0





 221
AGCAGCTT
tatgGCCTCTAAAAACATGGCCCC
17305
NmeCas9
 23
0





 222
AGGC
TCCCCTCCAGGCCGTGCATA
17306
SpyCas9-
 23
0






3var-NRRH







 223
AGCA
GCCTCTAAAAACATGGCCCC
17307
SpyCas9-
 23
0






3var-NRCH







 224
AAGG
TCTCCCCTCCAGGCCGTGCAT
17308
SauriCas9
 24
0





 225
AAGG
TCTCCCCTCCAGGCCGTGCAT
17309
SauriCas9-
 24
0






KKH







 226
CAG
GGCCTCTAAAAACATGGCCC
17310
ScaCas9
 24
0





 227
CAG
GGCCTCTAAAAACATGGCCC
17311
ScaCas9-HiFi-
 24
0






Sc++







 228
CAG
GGCCTCTAAAAACATGGCCC
17312
ScaCas9-
 24
0






Sc++







 229
CAG
GGCCTCTAAAAACATGGCCC
17313
SpyCas9-
 24
0






SpRY







 230
AAG
CTCCCCTCCAGGCCGTGCAT
17314
ScaCas9
 24
0





 231
AAG
CTCCCCTCCAGGCCGTGCAT
17315
ScaCas9-HiFi-
 24
0






Sc++







 232
AAG
CTCCCCTCCAGGCCGTGCAT
17316
ScaCas9-
 24
0






Sc++







 233
AAG
CTCCCCTCCAGGCCGTGCAT
17317
SpyCas9-
 24
0






SpRY







 234
AAGGCTGT
tctcTCCCCTCCAGGCCGTGCAT
17318
BlatCas9
 24
0





 235
AAGGC
tctcTCCCCTCCAGGCCGTGCAT
17319
BlatCas9
 24
0





 236
AAGGCT
TCTCCCCTCCAGGCCGTGCAT
17320
cCas9-v16
 24
0





 237
AAGGCT
TCTCCCCTCCAGGCCGTGCAT
17321
cCas9-v21
 24
0





 238
CAGC
GGCCTCTAAAAACATGGCCC
17322
SpyCas9-
 24
0






3var-NRRH







 239
TAAGG
CTCTCCCCTCCAGGCCGTGCA
17323
SauCas9KKH
 25
0





 240
CCAG
ATGGCCTCTAAAAACATGGCC
17324
SauriCas9-
 25
0






KKH







 241
TAAG
CTCTCCCCTCCAGGCCGTGCA
17325
SauriCas9-
 25
0






KKH







 242
TAAG
TCTCCCCTCCAGGCCGTGCA
17326
SpyCas9-
 25
0






QQR1







 243
TAAG
ctCTCCCCTCCAGGCCGTGCA
17327
iSpyMacCas9
 25
0





 244
TAA
TCTCCCCTCCAGGCCGTGCA
17328
SpyCas9-
 25
0






SpRY







 245
CCA
TGGCCTCTAAAAACATGGCC
17329
SpyCas9-
 25
0






SpRY







 246
CCAGC
gtatGGCCTCTAAAAACATGGCC
17330
BlatCas9
 25
0





 247
TAAGGC
CTCTCCCCTCCAGGCCGTGCA
17331
cCas9-v17
 25
0





 248
TAAGGC
CTCTCCCCTCCAGGCCGTGCA
17332
cCas9-v42
 25
0





 249
CCCAG
TATGGCCTCTAAAAACATGGC
17333
SauCas9KKH
 26
0





 250
ATAAG
TCTCTCCCCTCCAGGCCGTGC
17334
SauCas9KKH
 26
0





 251
CCC
ATGGCCTCTAAAAACATGGC
17335
SpyCas9-
 26
0






SpRY







 252
ATA
CTCTCCCCTCCAGGCCGTGC
17336
SpyCas9-
 26
0






SpRY







 253
CCCAGC
TATGGCCTCTAAAAACATGGC
17337
cCas9-v17
 26
0





 254
CCCAGC
TATGGCCTCTAAAAACATGGC
17338
cCas9-v42
 26
0





 255
ATAAGG
TCTCTCCCCTCCAGGCCGTGC
17339
cCas9-v17
 26
0





 256
ATAAGG
TCTCTCCCCTCCAGGCCGTGC
17340
cCas9-v42
 26
0





 257
CATAA
TTCTCTCCCCTCCAGGCCGTG
17341
SauCas9KKH
 27
0





 258
CAT
TCTCTCCCCTCCAGGCCGTG
17342
SpyCas9-
 27
0






SpRY







 259
CCC
TATGGCCTCTAAAAACATGG
17343
SpyCas9-
 27
0






SpRY







 260
CATA
TCTCTCCCCTCCAGGCCGTG
17344
SpyCas9-
 27
0






3var-NRTH







 261
GCC
GTATGGCCTCTAAAAACATG
17345
SpyCas9-
 28
0






SpRY







 262
GCA
TTCTCTCCCCTCCAGGCCGT
17346
SpyCas9-
 28
0






SpRY







 263
GCCCC
tgggTATGGCCTCTAAAAACATG
17347
BlatCas9
 28
0





 264
GGCCCC
caTGGGTATGGCCTCTAAAAACAT
17348
Nme2Cas9
 29
0





 265
GG
GGTATGGCCTCTAAAAACAT
17349
SpyCas9-NG
 29
0





 266
GG
GGTATGGCCTCTAAAAACAT
17350
SpyCas9-
 29
0






xCas







 267
GG
GGTATGGCCTCTAAAAACAT
17351
SpyCas9-
 29
0






xCas-NG







 268
TG
CTTCTCTCCCCTCCAGGCCG
17352
SpyCas9-NG
 29
0





 269
TG
CTTCTCTCCCCTCCAGGCCG
17353
SpyCas9-
 29
0






xCas







 270
TG
CTTCTCTCCCCTCCAGGCCG
17354
SpyCas9-
 29
0






xCas-NG







 271
GGC
GGTATGGCCTCTAAAAACAT
17355
SpyCas9-SpG
 29
0





 272
GGC
GGTATGGCCTCTAAAAACAT
17356
SpyCas9-
 29
0






SpRY







 273
TGC
CTTCTCTCCCCTCCAGGCCG
17357
SpyCas9-SpG
 29
0





 274
TGC
CTTCTCTCCCCTCCAGGCCG
17358
SpyCas9-
 29
0






SpRY







 275
GGCCCCAG
atggGTATGGCCTCTAAAAACAT
17359
BlatCas9
 29
0





 276
GGCCC
atggGTATGGCCTCTAAAAACAT
17360
BlatCas9
 29
0





 277
GGCC
GGTATGGCCTCTAAAAACAT
17361
SpyCas9-
 29
0






3var-NRCH







 278
TGCA
CTTCTCTCCCCTCCAGGCCG
17362
SpyCas9-
 29
0






3var-NRCH







 279
TGGCCC
acATGGGTATGGCCTCTAAAAACA
17363
Nme2Cas9
 30
0





 280
TGG
GGGTATGGCCTCTAAAAACA
17364
ScaCas9
 30
0





 281
TGG
GGGTATGGCCTCTAAAAACA
17365
ScaCas9-HiFi-
 30
0






Sc++







 282
TGG
GGGTATGGCCTCTAAAAACA
17366
ScaCas9-
 30
0






Sc++







 283
TGG
GGGTATGGCCTCTAAAAACA
17367
SpyCas9
 30
0





 284
TGG
GGGTATGGCCTCTAAAAACA
17368
SpyCas9-HF1
 30
0





 285
TGG
GGGTATGGCCTCTAAAAACA
17369
SpyCas9-SpG
 30
0





 286
TGG
GGGTATGGCCTCTAAAAACA
17370
SpyCas9-
 30
0






SpRY







 287
GTG
GCTTCTCTCCCCTCCAGGCC
17371
ScaCas9
 30
0





 288
GTG
GCTTCTCTCCCCTCCAGGCC
17372
ScaCas9-HiFi-
 30
0






Sc++







 289
GTG
GCTTCTCTCCCCTCCAGGCC
17373
ScaCas9-
 30
0






Sc++







 290
GTG
GCTTCTCTCCCCTCCAGGCC
17374
SpyCas9-
 30
0






SpRY







 291
TG
GGGTATGGCCTCTAAAAACA
17375
SpyCas9-NG
 30
0





 292
TG
GGGTATGGCCTCTAAAAACA
17376
SpyCas9-
 30
0






xCas







 293
TG
GGGTATGGCCTCTAAAAACA
17377
SpyCas9-
 30
0






xCas-NG







 294
TGGCC
catgGGTATGGCCTCTAAAAACA
17378
BlatCas9
 30
0





 295
TGGCCCC
ATGGGTATGGCCTCTAAAAACA
17379
CdiCas9
 30
0





 296
TGGC
GGGTATGGCCTCTAAAAACA
17380
SpyCas9-
 30
0






3var-NRRH







 297
ATGGCC
gaCATGGGTATGGCCTCTAAAAAC
17381
Nme2Cas9
 31
0





 298
ATGG
ATGGGTATGGCCTCTAAAAAC
17382
SauriCas9
 31
0





 299
ATGG
ATGGGTATGGCCTCTAAAAAC
17383
SauriCas9-
 31
0






KKH







 300
ATG
TGGGTATGGCCTCTAAAAAC
17384
ScaCas9
 31
0





 301
ATG
TGGGTATGGCCTCTAAAAAC
17385
ScaCas9-HiFi-
 31
0






Sc++







 302
ATG
TGGGTATGGCCTCTAAAAAC
17386
ScaCas9-
 31
0






Sc++







 303
ATG
TGGGTATGGCCTCTAAAAAC
17387
SpyCas9-
 31
0






SpRY







 304
CG
TGCTTCTCTCCCCTCCAGGC
17388
SpyCas9-NG
 31
0





 305
CG
TGCTTCTCTCCCCTCCAGGC
17389
SpyCas9-
 31
0






xCas







 306
CG
TGCTTCTCTCCCCTCCAGGC
17390
SpyCas9-
 31
0






xCas-NG







 307
CGT
TGCTTCTCTCCCCTCCAGGC
17391
SpyCas9-SpG
 31
0





 308
CGT
TGCTTCTCTCCCCTCCAGGC
17392
SpyCas9-
 31
0






SpRY







 309
CGTGCATA
ctctGCTTCTCTCCCCTCCAGGC
17393
BlatCas9
 31
0





 310
ATGGC
acatGGGTATGGCCTCTAAAAAC
17394
BlatCas9
 31
0





 311
CGTGC
ctctGCTTCTCTCCCCTCCAGGC
17395
BlatCas9
 31
0





 312
CATGG
CATGGGTATGGCCTCTAAAAA
17396
SauCas9KKH
 32
0





 313
CCG
CTGCTTCTCTCCCCTCCAGG
17397
ScaCas9
 32
0





 314
CCG
CTGCTTCTCTCCCCTCCAGG
17398
ScaCas9-HiFi-
 32
0






Sc++







 315
CCG
CTGCTTCTCTCCCCTCCAGG
17399
ScaCas9-
 32
0






Sc++







 316
CCG
CTGCTTCTCTCCCCTCCAGG
17400
SpyCas9-
 32
0






SpRY







 317
CAT
ATGGGTATGGCCTCTAAAAA
17401
SpyCas9-
 32
0






SpRY







 318
ACA
CATGGGTATGGCCTCTAAAA
17402
SpyCas9-
 33
0






SpRY







 319
GCC
TCTGCTTCTCTCCCCTCCAG
17403
SpyCas9-
 33
0






SpRY







 320
GCCGTG
CTCTGCTTCTCTCCCCTCCAG
17404
cCas9-v16
 33
0





 321
GCCGTG
CTCTGCTTCTCTCCCCTCCAG
17405
cCas9-v21
 33
0





 322
GG
CTCTGCTTCTCTCCCCTCCA
17406
SpyCas9-NG
 34
0





 323
GG
CTCTGCTTCTCTCCCCTCCA
17407
SpyCas9-
 34
0






xCas







 324
GG
CTCTGCTTCTCTCCCCTCCA
17408
SpyCas9-
 34
0






xCas-NG







 325
AAC
ACATGGGTATGGCCTCTAAA
17409
SpyCas9-
 34
0






SpRY







 326
GGC
CTCTGCTTCTCTCCCCTCCA
17410
SpyCas9-SpG
 34
0





 327
GGC
CTCTGCTTCTCTCCCCTCCA
17411
SpyCas9-
 34
0






SpRY







 328
AACA
ACATGGGTATGGCCTCTAAA
17412
SpyCas9-
 34
0






3var-NRCH







 329
GGCC
CTCTGCTTCTCTCCCCTCCA
17413
SpyCas9-
 34
0






3var-NRCH







 330
AGG
TCTCTGCTTCTCTCCCCTCC
17414
ScaCas9
 35
0





 331
AGG
TCTCTGCTTCTCTCCCCTCC
17415
ScaCas9-HiFi-
 35
0






Sc++







 332
AGG
TCTCTGCTTCTCTCCCCTCC
17416
ScaCas9-
 35
0






Sc++







 333
AGG
TCTCTGCTTCTCTCCCCTCC
17417
SpyCas9
 35
0





 334
AGG
TCTCTGCTTCTCTCCCCTCC
17418
SpyCas9-HF1
 35
0





 335
AGG
TCTCTGCTTCTCTCCCCTCC
17419
SpyCas9-SpG
 35
0





 336
AGG
TCTCTGCTTCTCTCCCCTCC
17420
SpyCas9-
 35
0






SpRY







 337
AG
TCTCTGCTTCTCTCCCCTCC
17421
SpyCas9-NG
 35
0





 338
AG
TCTCTGCTTCTCTCCCCTCC
17422
SpyCas9-
 35
0






xCas







 339
AG
TCTCTGCTTCTCTCCCCTCC
17423
SpyCas9-
 35
0






xCas-NG







 340
AAA
GACATGGGTATGGCCTCTAA
17424
SpyCas9-
 35
0






SpRY







 341
AGGCCGTG
gtgtCTCTGCTTCTCTCCCCTCC
17425
BlatCas9
 35
0





 342
AGGCC
gtgtCTCTGCTTCTCTCCCCTCC
17426
BlatCas9
 35
0





 343
AAAC
GACATGGGTATGGCCTCTAA
17427
SpyCas9-
 35
0






3var-NRRH







 344
AAAC
agACATGGGTATGGCCTCTAA
17428
iSpyMacCas9
 35
0





 345
AGGC
TCTCTGCTTCTCTCCCCTCC
17429
SpyCas9-
 35
0






3var-NRRH







 346
CAGGCC
acGTGTCTCTGCTTCTCTCCCCTC
17430
Nme2Cas9
 36
0





 347
CAGG
TGTCTCTGCTTCTCTCCCCTC
17431
SauriCas9
 36
0





 348
CAGG
TGTCTCTGCTTCTCTCCCCTC
17432
SauriCas9-
 36
0






KKH







 349
CAG
GTCTCTGCTTCTCTCCCCTC
17433
ScaCas9
 36
0





 350
CAG
GTCTCTGCTTCTCTCCCCTC
17434
ScaCas9-HiFi-
 36
0






Sc++







 351
CAG
GTCTCTGCTTCTCTCCCCTC
17435
ScaCas9-
 36
0






Sc++







 352
CAG
GTCTCTGCTTCTCTCCCCTC
17436
SpyCas9-
 36
0






SpRY







 353
AAA
AGACATGGGTATGGCCTCTA
17437
SpyCas9-
 36
0






SpRY







 354
AAAACATG
gataGACATGGGTATGGCCTCTA
17438
BlatCas9
 36
0





 355
CAGGCCGT
cgtgTCTCTGCTTCTCTCCCCTC
17439
BlatCas9
 36
0





 356
AAAAC
gataGACATGGGTATGGCCTCTA
17440
BlatCas9
 36
0





 357
CAGGC
cgtgTCTCTGCTTCTCTCCCCTC
17441
BlatCas9
 36
0





 358
AAAACAT
ATAGACATGGGTATGGCCTCTA
17442
CdiCas9
 36
0





 359
AAAA
AGACATGGGTATGGCCTCTA
17443
SpyCas9-
 36
0






3var-NRRH







 360
AAAA
taGACATGGGTATGGCCTCTA
17444
iSpyMacCas9
 36
0





 361
AAAAA
ATAGACATGGGTATGGCCTCT
17445
SauCas9KKH
 37
0





 362
CCAGG
GTGTCTCTGCTTCTCTCCCCT
17446
SauCas9KKH
 37
0





 363
CCAG
GTGTCTCTGCTTCTCTCCCCT
17447
SauriCas9-
 37
0






KKH







 364
AAA
TAGACATGGGTATGGCCTCT
17448
SpyCas9-
 37
0






SpRY







 365
CCA
TGTCTCTGCTTCTCTCCCCT
17449
SpyCas9-
 37
0






SpRY







 366
AAAAAC
ATAGACATGGGTATGGCCTCT
17450
cCas9-v17
 37
0





 367
AAAAAC
ATAGACATGGGTATGGCCTCT
17451
cCas9-v42
 37
0





 368
CCAGGC
GTGTCTCTGCTTCTCTCCCCT
17452
cCas9-v17
 37
0





 369
CCAGGC
GTGTCTCTGCTTCTCTCCCCT
17453
cCas9-v42
 37
0





 370
AAAA
TAGACATGGGTATGGCCTCT
17454
SpyCas9-
 37
0






3var-NRRH 







 371
AAAA
atAGACATGGGTATGGCCTCT
17455
iSpyMacCas9
 37
0





 372
TAAAA
GATAGACATGGGTATGGCCTC
17456
SauCas9KKH
 38
0





 373
TCCAG
CGTGTCTCTGCTTCTCTCCCC
17457
SauCas9KKH
 38
0





 374
TAA
ATAGACATGGGTATGGCCTC
17458
SpyCas9-
 38
0






SpRY







 375
TCC
GTGTCTCTGCTTCTCTCCCC
17459
SpyCas9-
 38
0






SpRY







 376
TAAAAA
ATAGACATGGGTATGGCCTC
17460
St1Cas9-
 38
0






MTH17CL396







 377
TAAAAA
GATAGACATGGGTATGGCCTC
17461
cCas9-v17
 38
0





 378
TAAAAA
GATAGACATGGGTATGGCCTC
17462
cCas9-v42
 38
0





 379
TCCAGG
CGTGTCTCTGCTTCTCTCCCC
17463
cCas9-v17
 38
0





 380
TCCAGG
CGTGTCTCTGCTTCTCTCCCC
17464
cCas9-v42
 38
0





 381
TAAAAAC
GGATAGACATGGGTATGGCCTC
17465
CdiCas9
 38
0





 382
TAAAAAC
GGATAGACATGGGTATGGCCTC
17466
CdiCas9
 38
0





 383
TAAA
ATAGACATGGGTATGGCCTC
17467
SpyCas9-
 38
0






3var-NRRH







 384
TAAA
gaTAGACATGGGTATGGCCTC
17468
iSpyMacCas9
 38
0





 385
CTAAA
GGATAGACATGGGTATGGCCT
17469
SauCas9KKH
 39
0





 386
CTA
GATAGACATGGGTATGGCCT
17470
SpyCas9-
 39
0






SpRY







 387
CTC
CGTGTCTCTGCTTCTCTCCC
17471
SpyCas9-
 39
0






SpRY







 388
CTAAAA
GATAGACATGGGTATGGCCT
17472
St1Cas9-
 39
0






MTH17CL396







 389
CTAAAA
GGATAGACATGGGTATGGCCT
17473
cCas9-v17
 39
0





 390
CTAAAA
GGATAGACATGGGTATGGCCT
17474
cCas9-v42
 39
0





 391
TCTAA
GGGATAGACATGGGTATGGCC
17475
SauCas9KKH
 40
0





 392
TCT
GGATAGACATGGGTATGGCC
17476
SpyCas9-
 40
0






SpRY







 393
CCT
ACGTGTCTCTGCTTCTCTCC
17477
SpyCas9-
 40
0






SpRY







 394
CCTCCAGG
acaaCGTGTCTCTGCTTCTCTCC
17478
BlatCas9
 40
0





 395
CCTCC
acaaCGTGTCTCTGCTTCTCTCC
17479
BlatCas9
 40
0





 396
CCCTCC
ttACAACGTGTCTCTGCTTCTCTC
17480
Nme2Cas9
 41
0





 397
CTC
GGGATAGACATGGGTATGGC
17481
SpyCas9-
 41
0






SpRY







 398
CCC
AACGTGTCTCTGCTTCTCTC
17482
SpyCas9-
 41
0






SpRY







 399
CCCTCCAG
tacaACGTGTCTCTGCTTCTCTC
17483
BlatCas9
 41
0





 400
CCCTC
tacaACGTGTCTCTGCTTCTCTC
17484
BlatCas9
 41
0





 401
CCT
GGGGATAGACATGGGTATGG
17485
SpyCas9-
 42
0






SpRY







 402
CCC
CAACGTGTCTCTGCTTCTCT
17486
SpyCas9-
 42
0






SpRY







 403
GCC
GGGGGATAGACATGGGTATG
17487
SpyCas9-
 43
0






SpRY







 404
TCC
ACAACGTGTCTCTGCTTCTC
17488
SpyCas9-
 43
0






SpRY







 405
GCCTCTAA
cgggGGGGATAGACATGGGTATG
17489
BlatCas9
 43
0





 406
GCCTCTAA
cgggGGGGATAGACATGGGTATG
17490
BlatCas9
 43
0





 407
GCCTC
cgggGGGGATAGACATGGGTATG
17491
BlatCas9
 43
0





 408
TCCCC
cttaCAACGTGTCTCTGCTTCTC
17492
BlatCas9
 43
0





 409
CTCCCC
gcCTTACAACGTGTCTCTGCTTCT
17493
Nme2Cas9
 44
0





 410
GG
GGGGGGATAGACATGGGTAT
17494
SpyCas9-NG
 44
0





 411
GG
GGGGGGATAGACATGGGTAT
17495
SpyCas9-
 44
0






xCas







 412
GG
GGGGGGATAGACATGGGTAT
17496
SpyCas9-
 44
0






xCas-NG







 413
GGC
GGGGGGATAGACATGGGTAT
17497
SpyCas9-SpG
 44
0





 414
GGC
GGGGGGATAGACATGGGTAT
17498
SpyCas9-
 44
0






SpRY







 415
CTC
TACAACGTGTCTCTGCTTCT
17499
SpyCas9-
 44
0






SpRY







 416
CTCCC
ccttACAACGTGTCTCTGCTTCT
17500
BlatCas9
 44
0





 417
GGCC
GGGGGGATAGACATGGGTAT
17501
SpyCas9-
 44
0






3var-NRCH







 418
TCTCCC
agCCTTACAACGTGTCTCTGCTTC
17502
Nme2Cas9
 45
0





 419
TGG
GGGGGGGATAGACATGGGTA
17503
ScaCas9
 45
0





 420
TGG
GGGGGGGATAGACATGGGTA
17504
ScaCas9-HiFi-
 45
0






Sc++







 421
TGG
GGGGGGGATAGACATGGGTA
17505
ScaCas9-
 45
0






Sc++







 422
TGG
GGGGGGGATAGACATGGGTA
17506
SpyCas9
 45
0





 423
TGG
GGGGGGGATAGACATGGGTA
17507
SpyCas9-HF1
 45
0





 424
TGG
GGGGGGGATAGACATGGGTA
17508
SpyCas9-SpG
 45
0





 425
TGG
GGGGGGGATAGACATGGGTA
17509
SpyCas9-
 45
0






SpRY







 426
TG
GGGGGGGATAGACATGGGTA
17510
SpyCas9-NG
 45
0





 427
TG
GGGGGGGATAGACATGGGTA
17511
SpyCas9-
 45
0






xCas







 428
TG
GGGGGGGATAGACATGGGTA
17512
SpyCas9-
 45
0






xCas-NG







 429
TCT
TTACAACGTGTCTCTGCTTC
17513
SpyCas9-
 45
0






SpRY







 430
TGGCC
ctcgGGGGGGATAGACATGGGTA
17514
BlatCas9
 45
0





 431
TCTCC
gcctTACAACGTGTCTCTGCTTC
17515
BlatCas9
 45
0





 432
TGGCCTC
TCGGGGGGGATAGACATGGGTA
17516
CdiCas9
 45
0





 433
TGGC
GGGGGGGATAGACATGGGTA
17517
SpyCas9-
 45
0






3var-NRRH







 434
ATGGCC
acCTCGGGGGGGATAGACATGGGT
17518
Nme2Cas9
 46
0





 435
CTCTCC
caGCCTTACAACGTGTCTCTGCTT
17519
Nme2Cas9
 46
0





 436
ATGG
TCGGGGGGGATAGACATGGGT
17520
SauriCas9
 46
0





 437
ATGG
TCGGGGGGGATAGACATGGGT
17521
SauriCas9-
 46
0






KKH







 438
ATG
CGGGGGGGATAGACATGGGT
17522
ScaCas9
 46
0





 439
ATG
CGGGGGGGATAGACATGGGT
17523
ScaCas9-HiFi-
 46
0






Sc++







 440
ATG
CGGGGGGGATAGACATGGGT
17524
ScaCas9-
 46
0






Sc++







 441
ATG
CGGGGGGGATAGACATGGGT
17525
SpyCas9-
 46
0






SpRY







 442
CTC
CTTACAACGTGTCTCTGCTT
17526
SpyCas9-
 46
0






SpRY







 443
ATGGC
cctcGGGGGGGATAGACATGGGT
17527
BlatCas9
 46
0





 444
CTCTC
agccTTACAACGTGTCTCTGCTT
17528
BlatCas9
 46
0





 445
TATGG
CTCGGGGGGGATAGACATGGG
17529
SauCas9KKH
 47
0





 446
TAT
TCGGGGGGGATAGACATGGG
17530
SpyCas9-
 47
0






SpRY







 447
TCT
CCTTACAACGTGTCTCTGCT
17531
SpyCas9-
 47
0






SpRY







 448
GTA
CTCGGGGGGGATAGACATGG
17532
SpyCas9-
 48
0






SpRY







 449
TTC
GCCTTACAACGTGTCTCTGC
17533
SpyCas9-
 48
0






SpRY







 450
TTCTC
tcagCCTTACAACGTGTCTCTGC
17534
BlatCas9
 48
0





 451
GG
CCTCGGGGGGGATAGACATG
17535
SpyCas9-NG
 49
0





 452
GG
CCTCGGGGGGGATAGACATG
17536
SpyCas9-
 49
0






xCas







 453
GG
CCTCGGGGGGGATAGACATG
17537
SpyCas9-
 49
0






xCas-NG







 454
GGT
CCTCGGGGGGGATAGACATG
17538
SpyCas9-SpG
 49
0





 455
GGT
CCTCGGGGGGGATAGACATG
17539
SpyCas9-
 49
0






SpRY







 456
CTT
AGCCTTACAACGTGTCTCTG
17540
SpyCas9-
 49
0






SpRY







 457
GGTA
CCTCGGGGGGGATAGACATG
17541
SpyCas9-
 49
0






3var-NRTH







 458
GGG
ACCTCGGGGGGGATAGACAT
17542
ScaCas9
 50
0





 459
GGG
ACCTCGGGGGGGATAGACAT
17543
ScaCas9-HiFi-
 50
0






Sc++







 460
GGG
ACCTCGGGGGGGATAGACAT
17544
ScaCas9-
 50
0






Sc++







 461
GGG
ACCTCGGGGGGGATAGACAT
17545
SpyCas9
 50
0





 462
GGG
ACCTCGGGGGGGATAGACAT
17546
SpyCas9-HF1
 50
0





 463
GGG
ACCTCGGGGGGGATAGACAT
17547
SpyCas9-SpG
 50
0





 464
GGG
ACCTCGGGGGGGATAGACAT
17548
SpyCas9-
 50
0






SpRY







 465
GG
ACCTCGGGGGGGATAGACAT
17549
SpyCas9-NG
 50
0





 466
GG
ACCTCGGGGGGGATAGACAT
17550
SpyCas9-
 50
0






xCas







 467
GG
ACCTCGGGGGGGATAGACAT
17551
SpyCas9-
 50
0






xCas-NG







 468
GCT
CAGCCTTACAACGTGTCTCT
17552
SpyCas9-
 50
0






SpRY







 469
GCTTC
gatcAGCCTTACAACGTGTCTCT
17553
BlatCas9
 50
0





 470
GGGT
ACCTCGGGGGGGATAGACAT
17554
SpyCas9-
 50
0






3var-NRRH







 471
TGGG
TGACCTCGGGGGGGATAGACA
17555
SauriCas9
 51
0





 472
TGGG
TGACCTCGGGGGGGATAGACA
17556
SauriCas9-
 51
0






KKH







 473
TGG
GACCTCGGGGGGGATAGACA
17557
ScaCas9
 51
0





 474
TGG
GACCTCGGGGGGGATAGACA
17558
ScaCas9-HiFi-
 51
0






Sc++







 475
TGG
GACCTCGGGGGGGATAGACA
17559
ScaCas9-
 51
0






Sc++







 476
TGG
GACCTCGGGGGGGATAGACA
17560
SpyCas9
 51
0





 477
TGG
GACCTCGGGGGGGATAGACA
17561
SpyCas9-HF1
 51
0





 478
TGG
GACCTCGGGGGGGATAGACA
17562
SpyCas9-SpG
 51
0





 479
TGG
GACCTCGGGGGGGATAGACA
17563
SpyCas9-
 51
0






SpRY







 480
TG
GACCTCGGGGGGGATAGACA
17564
SpyCas9-NG
 51
0





 481
TG
GACCTCGGGGGGGATAGACA
17565
SpyCas9-
 51
0






xCas







 482
TG
GACCTCGGGGGGGATAGACA
17566
SpyCas9-
 51
0






xCas-NG







 483
TG
TCAGCCTTACAACGTGTCTC
17567
SpyCas9-NG
 51
0





 484
TG
TCAGCCTTACAACGTGTCTC
17568
SpyCas9-
 51
0






xCas







 485
TG
TCAGCCTTACAACGTGTCTC
17569
SpyCas9-
 51
0






xCas-NG







 486
TGC
TCAGCCTTACAACGTGTCTC
17570
SpyCas9-SpG
 51
0





 487
TGC
TCAGCCTTACAACGTGTCTC
17571
SpyCas9-
 51
0






SpRY







 488
TGCT
TCAGCCTTACAACGTGTCTC
17572
SpyCas9-
 51
0






3var-NRCH







 489
ATGGG
acTTGACCTCGGGGGGGATAGAC
17573
SauCas9
 52
0





 490
ATGGG
TTGACCTCGGGGGGGATAGAC
17574
SauCas9KKH
 52
0





 491
ATGGGT
acTTGACCTCGGGGGGGATAGAC
17575
SauCas9
 52
0





 492
ATGGGT
TTGACCTCGGGGGGGATAGAC
17576
SauCas9KKH
 52
0





 493
ATGGGT
TTGACCTCGGGGGGGATAGAC
17577
cCas9-v17
 52
0





 494
ATGGGT
TTGACCTCGGGGGGGATAGAC
17578
cCas9-v42
 52
0





 495
ATGG
TTGACCTCGGGGGGGATAGAC
17579
SauriCas9
 52
0





 496
ATGG
TTGACCTCGGGGGGGATAGAC
17580
SauriCas9-
 52
0






KKH







 497
ATG
TGACCTCGGGGGGGATAGAC
17581
ScaCas9
 52
0





 498
ATG
TGACCTCGGGGGGGATAGAC
17582
ScaCas9-HiFi-
 52
0






Sc++







 499
ATG
TGACCTCGGGGGGGATAGAC
17583
ScaCas9-
 52
0






Sc++







 500
ATG
TGACCTCGGGGGGGATAGAC
17584
SpyCas9-
 52
0






SpRY







 501
CTG
ATCAGCCTTACAACGTGTCT
17585
ScaCas9
 52
0





 502
CTG
ATCAGCCTTACAACGTGTCT
17586
ScaCas9-HiFi-
 52
0






Sc++







 503
CTG
ATCAGCCTTACAACGTGTCT
17587
ScaCas9-
 52
0






Sc++







 504
CTG
ATCAGCCTTACAACGTGTCT
17588
SpyCas9-
 52
0






SpRY







 505
CTGCTTC
GGATCAGCCTTACAACGTGTCT
17589
CdiCas9
 52
0





 506
CATGG
CTTGACCTCGGGGGGGATAGA
17590
SauCas9KKH
 53
0





 507
CAT
TTGACCTCGGGGGGGATAGA
17591
SpyCas9-
 53
0






SpRY







 508
TCT
GATCAGCCTTACAACGTGTC
17592
SpyCas9-
 53
0






SpRY







 509
TCTGC
tgggATCAGCCTTACAACGTGTC
17593
BlatCas9
 53
0





 510
ACA
CTTGACCTCGGGGGGGATAG
17594
SpyCas9-
 54
0






SpRY







 511
CTC
GGATCAGCCTTACAACGTGT
17595
SpyCas9-
 54
0






SpRY







 512
CTCTGCTT
cctgGGATCAGCCTTACAACGTGT
17596
NmeCas9
 54
0





 513
GAC
ACTTGACCTCGGGGGGGATA
17597
SpyCas9-
 55
0






SpRY







 514
TCT
GGGATCAGCCTTACAACGTG
17598
SpyCas9-
 55
0






SpRY







 515
GACA
ACTTGACCTCGGGGGGGATA
17599
SpyCas9-
 55
0






3var-NRCH







 516
AG
AACTTGACCTCGGGGGGGAT
17600
SpyCas9-NG
 56
0





 517
AG
AACTTGACCTCGGGGGGGAT
17601
SpyCas9-
 56
0






xCas







 518
AG
AACTTGACCTCGGGGGGGAT
17602
SpyCas9-
 56
0






xCas-NG







 519
AGA
AACTTGACCTCGGGGGGGAT
17603
SpyCas9-SpG
 56
0





 520
AGA
AACTTGACCTCGGGGGGGAT
17604
SpyCas9-
 56
0






SpRY







 521
GTC
TGGGATCAGCCTTACAACGT
17605
SpyCas9-
 56
0






SpRY







 522
GTCTC
gcctGGGATCAGCCTTACAACGT
17606
BlatCas9
 56
0





 523
AGAC
AACTTGACCTCGGGGGGGAT
17607
SpyCas9-
 56
0






3var-NRRH







 524
AGAC
AACTTGACCTCGGGGGGGAT
17608
SpyCas9-VQR
 56
0





 525
TAG
GAACTTGACCTCGGGGGGGA
17609
ScaCas9
 57
0





 526
TAG
GAACTTGACCTCGGGGGGGA
17610
ScaCas9-HiFi-
 57
0






Sc++







 527
TAG
GAACTTGACCTCGGGGGGGA
17611
ScaCas9-
 57
0






Sc++







 528
TAG
GAACTTGACCTCGGGGGGGA
17612
SpyCas9-
 57
0






SpRY







 529
TG
CTGGGATCAGCCTTACAACG
17613
SpyCas9-NG
 57
0





 530
TG
CTGGGATCAGCCTTACAACG
17614
SpyCas9-
 57
0






xCas







 531
TG
CTGGGATCAGCCTTACAACG
17615
SpyCas9-
 57
0






xCas-NG







 532
TGT
CTGGGATCAGCCTTACAACG
17616
SpyCas9-SpG
 57
0





 533
TGT
CTGGGATCAGCCTTACAACG
17617
SpyCas9-
 57
0






SpRY







 534
TAGACATG
gttgAACTTGACCTCGGGGGGGA
17618
BlatCas9
 57
0





 535
TAGAC
gttgAACTTGACCTCGGGGGGGA
17619
BlatCas9
 57
0





 536
TAGACAT
TTGAACTTGACCTCGGGGGGGA
17620
CdiCas9
 57
0





 537
TAGA
GAACTTGACCTCGGGGGGGA
17621
SpyCas9-
 57
0






3var-NRRH







 538
TGTC
CTGGGATCAGCCTTACAACG
17622
SpyCas9-
 57
0






3var-NRTH







 539
ATAGA
TTGAACTTGACCTCGGGGGGG
17623
SauCas9KKH
 58
0





 540
ATAG
TTGAACTTGACCTCGGGGGGG
17624
SauriCas9-
 58
0






KKH







 541
GTG
CCTGGGATCAGCCTTACAAC
17625
ScaCas9
 58
0





 542
GTG
CCTGGGATCAGCCTTACAAC
17626
ScaCas9-HiFi-
 58
0






Sc++







 543
GTG
CCTGGGATCAGCCTTACAAC
17627
ScaCas9-
 58
0






Sc++







 544
GTG
CCTGGGATCAGCCTTACAAC
17628
SpyCas9-
 58
0






SpRY







 545
ATA
TGAACTTGACCTCGGGGGGG
17629
SpyCas9-
 58
0






SpRY







 546
GTGTC
aggcCTGGGATCAGCCTTACAAC
17630
BlatCas9
 58
0





 547
ATAGAC
TTGAACTTGACCTCGGGGGGG
17631
cCas9-v17
 58
0





 548
ATAGAC
TTGAACTTGACCTCGGGGGGG
17632
cCas9-v42
 58
0





 549
GTGTCTC
GGCCTGGGATCAGCCTTACAAC
17633
CdiCas9
 58
0





 550
GATAG
GTTGAACTTGACCTCGGGGGG
17634
SauCas9KKH
 59
0





 551
CG
GCCTGGGATCAGCCTTACAA
17635
SpyCas9-NG
 59
0





 552
CG
GCCTGGGATCAGCCTTACAA
17636
SpyCas9-
 59
0






xCas







 553
CG
GCCTGGGATCAGCCTTACAA
17637
SpyCas9-
 59
0






xCas-NG







 554
GAT
TTGAACTTGACCTCGGGGGG
17638
SpyCas9-
 59
0






SpRY







 555
GAT
TTGAACTTGACCTCGGGGGG
17639
SpyCas9-
 59
0






xCas







 556
CGT
GCCTGGGATCAGCCTTACAA
17640
SpyCas9-SpG
 59
0





 557
CGT
GCCTGGGATCAGCCTTACAA
17641
SpyCas9-
 59
0






SpRY







 558
GATAGACA
tttgTTGAACTTGACCTCGGGGGG
17642
NmeCas9
 59
0





 559
GATA
TTGAACTTGACCTCGGGGGG
17643
SpyCas9-
 59
0






3var-NRTH







 560
ACG
GGCCTGGGATCAGCCTTACA
17644
ScaCas9
 60
0





 561
ACG
GGCCTGGGATCAGCCTTACA
17645
ScaCas9-HiFi-
 60
0






Sc++







 562
ACG
GGCCTGGGATCAGCCTTACA
17646
ScaCas9-
 60
0






Sc++







 563
ACG
GGCCTGGGATCAGCCTTACA
17647
SpyCas9-
 60
0






SpRY







 564
GG
GTTGAACTTGACCTCGGGGG
17648
SpyCas9-NG
 60
0





 565
GG
GTTGAACTTGACCTCGGGGG
17649
SpyCas9-
 60
0






xCas







 566
GG
GTTGAACTTGACCTCGGGGG
17650
SpyCas9-
 60
0






xCas-NG







 567
GGA
GTTGAACTTGACCTCGGGGG
17651
SpyCas9-SpG
 60
0





 568
GGA
GTTGAACTTGACCTCGGGGG
17652
SpyCas9-
 60
0






SpRY







 569
ACGTGTCT
tcgaGGCCTGGGATCAGCCTTACA
17653
NmeCas9
 60
0





 570
GGAT
GTTGAACTTGACCTCGGGGG
17654
SpyCas9-
 60
0






3var-NRRH







 571
GGAT
GTTGAACTTGACCTCGGGGG
17655
SpyCas9-VQR
 60
0





 572
GGG
TGTTGAACTTGACCTCGGGG
17656
ScaCas9
 61
0





 573
GGG
TGTTGAACTTGACCTCGGGG
17657
ScaCas9-HiFi-
 61
0






Sc++







 574
GGG
TGTTGAACTTGACCTCGGGG
17658
ScaCas9-
 61
0






Sc++







 575
GGG
TGTTGAACTTGACCTCGGGG
17659
SpyCas9
 61
0





 576
GGG
TGTTGAACTTGACCTCGGGG
17660
SpyCas9-HF1
 61
0





 577
GGG
TGTTGAACTTGACCTCGGGG
17661
SpyCas9-SpG
 61
0





 578
GGG
TGTTGAACTTGACCTCGGGG
17662
SpyCas9-
 61
0






SpRY







 579
GG
TGTTGAACTTGACCTCGGGG
17663
SpyCas9-NG
 61
0





 580
GG
TGTTGAACTTGACCTCGGGG
17664
SpyCas9-
 61
0






xCas







 581
GG
TGTTGAACTTGACCTCGGGG
17665
SpyCas9-
 61
0






xCas-NG







 582
AAC
AGGCCTGGGATCAGCCTTAC
17666
SpyCas9-
 61
0






SpRY







 583
AACGTG
GAGGCCTGGGATCAGCCTTAC
17667
cCas9-v16
 61
0





 584
AACGTG
GAGGCCTGGGATCAGCCTTAC
17668
cCas9-v21
 61
0





 585
GGGA
TGTTGAACTTGACCTCGGGG
17669
SpyCas9-
 61
0






3var-NRRH







 586
GGGGA
ggTTTGTTGAACTTGACCTCGGG
17670
SauCas9
 62
0





 587
GGGGA
TTTGTTGAACTTGACCTCGGG
17671
SauCas9KKH
 62
0





 588
GGGGAT
ggTTTGTTGAACTTGACCTCGGG
17672
SauCas9
 62
0





 589
GGGGAT
TTTGTTGAACTTGACCTCGGG
17673
SauCas9KKH
 62
0





 590
GGGGAT
TTTGTTGAACTTGACCTCGGG
17674
cCas9-v17
 62
0





 591
GGGGAT
TTTGTTGAACTTGACCTCGGG
17675
cCas9-v42
 62
0





 592
GGGG
TTTGTTGAACTTGACCTCGGG
17676
SauriCas9
 62
0





 593
GGGG
TTTGTTGAACTTGACCTCGGG
17677
SauriCas9-
 62
0






KKH







 594
GGG
TTGTTGAACTTGACCTCGGG
17678
ScaCas9
 62
0





 595
GGG
TTGTTGAACTTGACCTCGGG
17679
ScaCas9-HiFi-
 62
0






Sc++







 596
GGG
TTGTTGAACTTGACCTCGGG
17680
ScaCas9-
 62
0






Sc++







 597
GGG
TTGTTGAACTTGACCTCGGG
17681
SpyCas9
 62
0





 598
GGG
TTGTTGAACTTGACCTCGGG
17682
SpyCas9-HF1
 62
0





 599
GGG
TTGTTGAACTTGACCTCGGG
17683
SpyCas9-SpG
 62
0





 600
GGG
TTGTTGAACTTGACCTCGGG
17684
SpyCas9-
 62
0






SpRY







 601
GG
TTGTTGAACTTGACCTCGGG
17685
SpyCas9-NG
 62
0





 602
GG
TTGTTGAACTTGACCTCGGG
17686
SpyCas9-
 62
0






xCas







 603
GG
TTGTTGAACTTGACCTCGGG
17687
SpyCas9-
 62
0






xCas-NG







 604
CAA
GAGGCCTGGGATCAGCCTTA
17688
SpyCas9-
 62
0






SpRY







 605
CAAC
GAGGCCTGGGATCAGCCTTA
17689
SpyCas9-
 62
0






3var-NRRH







 606
CAAC
cgAGGCCTGGGATCAGCCTTA
17690
iSpyMacCas9
 62
0





 607
GGGGG
ggGTTTGTTGAACTTGACCTCGG
17691
SauCas9
 63
0





 608
GGGGG
GTTTGTTGAACTTGACCTCGG
17692
SauCas9KKH
 63
0





 609
GGGG
GTTTGTTGAACTTGACCTCGG
17693
SauriCas9
 63
0





 610
GGGG
GTTTGTTGAACTTGACCTCGG
17694
SauriCas9-
 63
0






KKH







 611
GGG
TTTGTTGAACTTGACCTCGG
17695
ScaCas9
 63
0





 612
GGG
TTTGTTGAACTTGACCTCGG
17696
ScaCas9-HiFi-
 63
0






Sc++







 613
GGG
TTTGTTGAACTTGACCTCGG
17697
ScaCas9-
 63
0






Sc++







 614
GGG
TTTGTTGAACTTGACCTCGG
17698
SpyCas9
 63
0





 615
GGG
TTTGTTGAACTTGACCTCGG
17699
SpyCas9-HF1
 63
0





 616
GGG
TTTGTTGAACTTGACCTCGG
17700
SpyCas9-SpG
 63
0





 617
GGG
TTTGTTGAACTTGACCTCGG
17701
SpyCas9-
 63
0






SpRY







 618
GG
TTTGTTGAACTTGACCTCGG
17702
SpyCas9-NG
 63
0





 619
GG
TTTGTTGAACTTGACCTCGG
17703
SpyCas9-
 63
0






xCas







 620
GG
TTTGTTGAACTTGACCTCGG
17704
SpyCas9-
 63
0






xCas-NG







 621
ACA
CGAGGCCTGGGATCAGCCTT
17705
SpyCas9-
 63
0






SpRY







 622
ACAACGTG
gctcGAGGCCTGGGATCAGCCTT
17706
BlatCas9
 63
0





 623
ACAAC
gctcGAGGCCTGGGATCAGCCTT
17707
BlatCas9
 63
0





 624
GGGGGA
GTTTGTTGAACTTGACCTCGG
17708
cCas9-v17
 63
0





 625
GGGGGA
GTTTGTTGAACTTGACCTCGG
17709
cCas9-v42
 63
0





 626
GGGGGATA
agggTTTGTTGAACTTGACCTCGG
17710
NmeCas9
 63
0





 627
GGGGG
agGGTTTGTTGAACTTGACCTCG
17711
SauCas9
 64
0





 628
GGGGG
GGTTTGTTGAACTTGACCTCG
17712
SauCas9KKH
 64
0





 629
TACAA
CTCGAGGCCTGGGATCAGCCT
17713
SauCas9KKH
 64
0





 630
GGGG
GGTTTGTTGAACTTGACCTCG
17714
SauriCas9
 64
0





 631
GGGG
GGTTTGTTGAACTTGACCTCG
17715
SauriCas9-
 64
0






KKH







 632
GGG
GTTTGTTGAACTTGACCTCG
17716
ScaCas9
 64
0





 633
GGG
GTTTGTTGAACTTGACCTCG
17717
ScaCas9-HiFi-
 64
0






Sc++







 634
GGG
GTTTGTTGAACTTGACCTCG
17718
ScaCas9-
 64
0






Sc++







 635
GGG
GTTTGTTGAACTTGACCTCG
17719
SpyCas9
 64
0





 636
GGG
GTTTGTTGAACTTGACCTCG
17720
SpyCas9-HF1
 64
0





 637
GGG
GTTTGTTGAACTTGACCTCG
17721
SpyCas9-SpG
 64
0





 638
GGG
GTTTGTTGAACTTGACCTCG
17722
SpyCas9-
 64
0






SpRY







 639
GG
GTTTGTTGAACTTGACCTCG
17723
SpyCas9-NG
 64
0





 640
GG
GTTTGTTGAACTTGACCTCG
17724
SpyCas9-
 64
0






xCas







 641
GG
GTTTGTTGAACTTGACCTCG
17725
SpyCas9-
 64
0






xCas-NG







 642
TAC
TCGAGGCCTGGGATCAGCCT
17726
SpyCas9-
 64
0






SpRY







 643
GGGGGG
GGTTTGTTGAACTTGACCTCG
17727
cCas9-v17
 64
0





 644
GGGGGG
GGTTTGTTGAACTTGACCTCG
17728
cCas9-v42
 64
0





 645
TACAAC
CTCGAGGCCTGGGATCAGCCT
17729
cCas9-v17
 64
0





 646
TACAAC
CTCGAGGCCTGGGATCAGCCT
17730
cCas9-v42
 64
0





 647
TACA
TCGAGGCCTGGGATCAGCCT
17731
SpyCas9-
 64
0






3var-NRCH







 648
GGGGG
aaGGGTTTGTTGAACTTGACCTC
17732
SauCas9
 65
0





 649
GGGGG
GGGTTTGTTGAACTTGACCTC
17733
SauCas9KKH
 65
0





 650
GGGG
GGGTTTGTTGAACTTGACCTC
17734
SauriCas9
 65
0





 651
GGGG
GGGTTTGTTGAACTTGACCTC
17735
SauriCas9-
 65
0






KKH







 652
GGG
GGTTTGTTGAACTTGACCTC
17736
ScaCas9
 65
0





 653
GGG
GGTTTGTTGAACTTGACCTC
17737
ScaCas9-HiFi-
 65
0






Sc++







 654
GGG
GGTTTGTTGAACTTGACCTC
17738
ScaCas9-
 65
0






Sc++







 655
GGG
GGTTTGTTGAACTTGACCTC
17739
SpyCas9
 65
0





 656
GGG
GGTTTGTTGAACTTGACCTC
17740
SpyCas9-HF1
 65
0





 657
GGG
GGTTTGTTGAACTTGACCTC
17741
SpyCas9-SpG
 65
0





 658
GGG
GGTTTGTTGAACTTGACCTC
17742
SpyCas9-
 65
0






SpRY







 659
GG
GGTTTGTTGAACTTGACCTC
17743
SpyCas9-NG
 65
0





 660
GG
GGTTTGTTGAACTTGACCTC
17744
SpyCas9-
 65
0






xCas







 661
GG
GGTTTGTTGAACTTGACCTC
17745
SpyCas9-
 65
0






xCas-NG







 662
TTA
CTCGAGGCCTGGGATCAGCC
17746
SpyCas9-
 65
0






SpRY







 663
GGGGGG
GGGTTTGTTGAACTTGACCTC
17747
cCas9-v17
 65
0





 664
GGGGGG
GGGTTTGTTGAACTTGACCTC
17748
cCas9-v42
 65
0





 665
TTACAAC
TGCTCGAGGCCTGGGATCAGCC
17749
CdiCas9
 65
0





 666
TTACAA
CTCGAGGCCTGGGATCAGCC
17750
St1Cas9-
 65
0






CNRZ1066







 667
CGGGG
aaAGGGTTTGTTGAACTTGACCT
17751
SauCas9
 66
0





 668
CGGGG
AGGGTTTGTTGAACTTGACCT
17752
SauCas9KKH
 66
0





 669
CGGG
AGGGTTTGTTGAACTTGACCT
17753
SauriCas9
 66
0





 670
CGGG
AGGGTTTGTTGAACTTGACCT
17754
SauriCas9-
 66
0






KKH







 671
CGG
GGGTTTGTTGAACTTGACCT
17755
ScaCas9
 66
0





 672
CGG
GGGTTTGTTGAACTTGACCT
17756
ScaCas9-HiFi-
 66
0






Sc++







 673
CGG
GGGTTTGTTGAACTTGACCT
17757
ScaCas9-
 66
0






Sc++







 674
CGG
GGGTTTGTTGAACTTGACCT
17758
SpyCas9
 66
0





 675
CGG
GGGTTTGTTGAACTTGACCT
17759
SpyCas9-HF1
 66
0





 676
CGG
GGGTTTGTTGAACTTGACCT
17760
SpyCas9-SpG
 66
0





 677
CGG
GGGTTTGTTGAACTTGACCT
17761
SpyCas9-
 66
0






SpRY







 678
CG
GGGTTTGTTGAACTTGACCT
17762
SpyCas9-NG
 66
0





 679
CG
GGGTTTGTTGAACTTGACCT
17763
SpyCas9-
 66
0






xCas







 680
CG
GGGTTTGTTGAACTTGACCT
17764
SpyCas9-
 66
0






xCas-NG







 681
CTT
GCTCGAGGCCTGGGATCAGC
17765
SpyCas9-
 66
0






SpRY







 682
CTTAC
cttgCTCGAGGCCTGGGATCAGC
17766
BlatCas9
 66
0





 683
CGGGGG
AGGGTTTGTTGAACTTGACCT
17767
cCas9-v17
 66
0





 684
CGGGGG
AGGGTTTGTTGAACTTGACCT
17768
cCas9-v42
 66
0





 685
TCGGG
caAAGGGTTTGTTGAACTTGACC
17769
SauCas9
 67
0





 686
TCGGG
AAGGGTTTGTTGAACTTGACC
17770
SauCas9KKH
 67
0





 687
TCGG
AAGGGTTTGTTGAACTTGACC
17771
SauriCas9
 67
0





 688
TCGG
AAGGGTTTGTTGAACTTGACC
17772
SauriCas9-
 67
0






KKH







 689
TCG
AGGGTTTGTTGAACTTGACC
17773
ScaCas9
 67
0





 690
TCG
AGGGTTTGTTGAACTTGACC
17774
ScaCas9-HiFi-
 67
0






Sc++







 691
TCG
AGGGTTTGTTGAACTTGACC
17775
ScaCas9-
 67
0






Sc++







 692
TCG
AGGGTTTGTTGAACTTGACC
17776
SpyCas9-
 67
0






SpRY







 693
CCT
TGCTCGAGGCCTGGGATCAG
17777
SpyCas9-
 67
0






SpRY







 694
TCGGGG
AAGGGTTTGTTGAACTTGACC
17778
cCas9-v17
 67
0





 695
TCGGGG
AAGGGTTTGTTGAACTTGACC
17779
cCas9-v42
 67
0





 696
CTCGG
AAAGGGTTTGTTGAACTTGAC
17780
SauCas9KKH
 68
0





 697
CTC
AAGGGTTTGTTGAACTTGAC
17781
SpyCas9-
 68
0






SpRY







 698
GCC
TTGCTCGAGGCCTGGGATCA
17782
SpyCas9-
 68
0






SpRY







 699
CTCGGG
AAAGGGTTTGTTGAACTTGAC
17783
cCas9-v17
 68
0





 700
CTCGGG
AAAGGGTTTGTTGAACTTGAC
17784
cCas9-v42
 68
0





 701
AG
CTTGCTCGAGGCCTGGGATC
17785
SpyCas9-NG
 69
0





 702
AG
CTTGCTCGAGGCCTGGGATC
17786
SpyCas9-
 69
0






xCas







 703
AG
CTTGCTCGAGGCCTGGGATC
17787
SpyCas9-
 69
0






xCas-NG







 704
AGC
CTTGCTCGAGGCCTGGGATC
17788
SpyCas9-SpG
 69
0





 705
AGC
CTTGCTCGAGGCCTGGGATC
17789
SpyCas9-
 69
0






SpRY







 706
CCT
AAAGGGTTTGTTGAACTTGA
17790
SpyCas9-
 69
0






SpRY







 707
AGCC
CTTGCTCGAGGCCTGGGATC
17791
SpyCas9-
 69
0






3var-NRCH







 708
CAG
CCTTGCTCGAGGCCTGGGAT
17792
ScaCas9
 70
0





 709
CAG
CCTTGCTCGAGGCCTGGGAT
17793
ScaCas9-HiFi-
 70
0






Sc++







 710
CAG
CCTTGCTCGAGGCCTGGGAT
17794
ScaCas9-
 70
0






Sc++







 711
CAG
CCTTGCTCGAGGCCTGGGAT
17795
SpyCas9-
 70
0






SpRY







 712
ACC
CAAAGGGTTTGTTGAACTTG
17796
SpyCas9-
 70
0






SpRY







 713
ACCTCGGG
agacAAAGGGTTTGTTGAACTTG
17797
BlatCas9
 70
0





 714
CAGCCTTA
gagcCTTGCTCGAGGCCTGGGAT
17798
BlatCas9
 70
0





 715
ACCTC
agacAAAGGGTTTGTTGAACTTG
17799
BlatCas9
 70
0





 716
CAGCC
gagcCTTGCTCGAGGCCTGGGAT
17800
BlatCas9
 70
0





 717
CAGCCTT
AGCCTTGCTCGAGGCCTGGGAT
17801
CdiCas9
 70
0





 718
CAGC
CCTTGCTCGAGGCCTGGGAT
17802
SpyCas9-
 70
0






3var-NRRH







 719
TCAGCC
gtGAGCCTTGCTCGAGGCCTGGGA
17803
Nme2Cas9
 71
0





 720
TCAG
AGCCTTGCTCGAGGCCTGGGA
17804
SauriCas9-
 71
0






KKH







 721
GAC
ACAAAGGGTTTGTTGAACTT
17805
SpyCas9-
 71
0






SpRY







 722
TCA
GCCTTGCTCGAGGCCTGGGA
17806
SpyCas9-
 71
0






SpRY







 723
TCAGCCTT
tgagCCTTGCTCGAGGCCTGGGA
17807
BlatCas9
 71
0





 724
TCAGC
tgagCCTTGCTCGAGGCCTGGGA
17808
BlatCas9
 71
0





 725
GACC
ACAAAGGGTTTGTTGAACTT
17809
SpyCas9-
 71
0






3var-NRCH







 726
ATCAG
GAGCCTTGCTCGAGGCCTGGG
17810
SauCas9KKH
 72
0





 727
TG
GACAAAGGGTTTGTTGAACT
17811
SpyCas9-NG
 72
0





 728
TG
GACAAAGGGTTTGTTGAACT
17812
SpyCas9-
 72
0






xCas







 729
TG
GACAAAGGGTTTGTTGAACT
17813
SpyCas9-
 72
0






xCas-NG







 730
TGA
GACAAAGGGTTTGTTGAACT
17814
SpyCas9-SpG
 72
0





 731
TGA
GACAAAGGGTTTGTTGAACT
17815
SpyCas9-
 72
0






SpRY







 732
ATC
AGCCTTGCTCGAGGCCTGGG
17816
SpyCas9-
 72
0






SpRY







 733
TGACC
gaagACAAAGGGTTTGTTGAACT
17817
BlatCas9
 72
0





 734
ATCAGC
GAGCCTTGCTCGAGGCCTGGG
17818
cCas9-v17
 72
0





 735
ATCAGC
GAGCCTTGCTCGAGGCCTGGG
17819
cCas9-v42
 72
0





 736
TGACCTC
AAGACAAAGGGTTTGTTGAACT
17820
CdiCas9
 72
0





 737
TGAC
GACAAAGGGTTTGTTGAACT
17821
SpyCas9-
 72
0






3var-NRRH







 738
TGAC
GACAAAGGGTTTGTTGAACT
17822
SpyCas9-VQR
 72
0





 739
TTGACC
aaGAAGACAAAGGGTTTGTTGAAC
17823
Nme2Cas9
 73
0





 740
TTG
AGACAAAGGGTTTGTTGAAC
17824
ScaCas9
 73
0





 741
TTG
AGACAAAGGGTTTGTTGAAC
17825
ScaCas9-HiFi-
 73
0






Sc++







 742
TTG
AGACAAAGGGTTTGTTGAAC
17826
ScaCas9-
 73
0






Sc++







 743
TTG
AGACAAAGGGTTTGTTGAAC
17827
SpyCas9-
 73
0






SpRY







 744
GAT
GAGCCTTGCTCGAGGCCTGG
17828
SpyCas9-
 73
0






SpRY







 745
GAT
GAGCCTTGCTCGAGGCCTGG
17829
SpyCas9-
 73
0






xCas







 746
TTGAC
agaaGACAAAGGGTTTGTTGAAC
17830
BlatCas9
 73
0





 747
TTGACCT
GAAGACAAAGGGTTTGTTGAAC
17831
CdiCas9
 73
0





 748
GATC
GAGCCTTGCTCGAGGCCTGG
17832
SpyCas9-
 73
0






3var-NRTH







 749
CTTGA
GAAGACAAAGGGTTTGTTGAA
17833
SauCas9KKH
 74
0





 750
GG
TGAGCCTTGCTCGAGGCCTG
17834
SpyCas9-NG
 74
0





 751
GG
TGAGCCTTGCTCGAGGCCTG
17835
SpyCas9-
 74
0






xCas







 752
GG
TGAGCCTTGCTCGAGGCCTG
17836
SpyCas9-
 74
0






xCas-NG







 753
GGA
TGAGCCTTGCTCGAGGCCTG
17837
SpyCas9-SpG
 74
0





 754
GGA
TGAGCCTTGCTCGAGGCCTG
17838
SpyCas9-
 74
0






SpRY







 755
CTT
AAGACAAAGGGTTTGTTGAA
17839
SpyCas9-
 74
0






SpRY







 756
GGATC
acgtGAGCCTTGCTCGAGGCCTG
17840
BlatCas9
 74
0





 757
GGAT
TGAGCCTTGCTCGAGGCCTG
17841
SpyCas9-
 74
0






3var-NRRH







 758
GGAT
TGAGCCTTGCTCGAGGCCTG
17842
SpyCas9-VQR
 74
0





 759
GGG
GTGAGCCTTGCTCGAGGCCT
17843
ScaCas9
 75
0





 760
GGG
GTGAGCCTTGCTCGAGGCCT
17844
ScaCas9-HiFi-
 75
0






Sc++







 761
GGG
GTGAGCCTTGCTCGAGGCCT
17845
ScaCas9-
 75
0






Sc++







 762
GGG
GTGAGCCTTGCTCGAGGCCT
17846
SpyCas9
 75
0





 763
GGG
GTGAGCCTTGCTCGAGGCCT
17847
SpyCas9-HF1
 75
0





 764
GGG
GTGAGCCTTGCTCGAGGCCT
17848
SpyCas9-SpG
 75
0





 765
GGG
GTGAGCCTTGCTCGAGGCCT
17849
SpyCas9-
 75
0






SpRY







 766
GG
GTGAGCCTTGCTCGAGGCCT
17850
SpyCas9-NG
 75
0





 767
GG
GTGAGCCTTGCTCGAGGCCT
17851
SpyCas9-
 75
0






xCas







 768
GG
GTGAGCCTTGCTCGAGGCCT
17852
SpyCas9-
 75
0






xCas-NG







 769
ACT
GAAGACAAAGGGTTTGTTGA
17853
SpyCas9-
 75
0






SpRY







 770
GGGA
GTGAGCCTTGCTCGAGGCCT
17854
SpyCas9-
 75
0






3var-NRRH







 771
TGGGA
ccACGTGAGCCTTGCTCGAGGCC
17855
SauCas9
 76
0





 772
TGGGA
ACGTGAGCCTTGCTCGAGGCC
17856
SauCas9KKH
 76
0





 773
TGGGAT
ccACGTGAGCCTTGCTCGAGGCC
17857
SauCas9
 76
0





 774
TGGGAT
ACGTGAGCCTTGCTCGAGGCC
17858
SauCas9KKH
 76
0





 775
TGGGAT
ACGTGAGCCTTGCTCGAGGCC
17859
cCas9-v17
 76
0





 776
TGGGAT
ACGTGAGCCTTGCTCGAGGCC
17860
cCas9-v42
 76
0





 777
TGGG
ACGTGAGCCTTGCTCGAGGCC
17861
SauriCas9
 76
0





 778
TGGG
ACGTGAGCCTTGCTCGAGGCC
17862
SauriCas9-
 76
0






KKH







 779
TGG
CGTGAGCCTTGCTCGAGGCC
17863
ScaCas9
 76
0





 780
TGG
CGTGAGCCTTGCTCGAGGCC
17864
ScaCas9-HiFi-
 76
0






Sc++







 781
TGG
CGTGAGCCTTGCTCGAGGCC
17865
ScaCas9-
 76
0






Sc++







 782
TGG
CGTGAGCCTTGCTCGAGGCC
17866
SpyCas9
 76
0





 783
TGG
CGTGAGCCTTGCTCGAGGCC
17867
SpyCas9-HF1
 76
0





 784
TGG
CGTGAGCCTTGCTCGAGGCC
17868
SpyCas9-SpG
 76
0





 785
TGG
CGTGAGCCTTGCTCGAGGCC
17869
SpyCas9-
 76
0






SpRY







 786
TG
CGTGAGCCTTGCTCGAGGCC
17870
SpyCas9-NG
 76
0





 787
TG
CGTGAGCCTTGCTCGAGGCC
17871
SpyCas9-
 76
0






xCas







 788
TG
CGTGAGCCTTGCTCGAGGCC
17872
SpyCas9-
 76
0






xCas-NG







 789
AAC
AGAAGACAAAGGGTTTGTTG
17873
SpyCas9-
 76
0






SpRY







 790
AACT
AGAAGACAAAGGGTTTGTTG
17874
SpyCas9-
 76
0






3var-NRCH







 791
CTGGG
tcCACGTGAGCCTTGCTCGAGGC
17875
SauCas9
 77
0





 792
CTGGG
CACGTGAGCCTTGCTCGAGGC
17876
SauCas9KKH
 77
0





 793
CTGG
CACGTGAGCCTTGCTCGAGGC
17877
SauriCas9
 77
0





 794
CTGG
CACGTGAGCCTTGCTCGAGGC
17878
SauriCas9-
 77
0






KKH







 795
CTG
ACGTGAGCCTTGCTCGAGGC
17879
ScaCas9
 77
0





 796
CTG
ACGTGAGCCTTGCTCGAGGC
17880
ScaCas9-HiFi-
 77
0






Sc++







 797
CTG
ACGTGAGCCTTGCTCGAGGC
17881
ScaCas9-
 77
0






Sc++







 798
CTG
ACGTGAGCCTTGCTCGAGGC
17882
SpyCas9-
 77
0






SpRY







 799
GAA
AAGAAGACAAAGGGTTTGTT
17883
SpyCas9-
 77
0






SpRY







 800
GAA
AAGAAGACAAAGGGTTTGTT
17884
SpyCas9-
 77
0






xCas







 801
CTGGGA
CACGTGAGCCTTGCTCGAGGC
17885
cCas9-v17
 77
0





 802
CTGGGA
CACGTGAGCCTTGCTCGAGGC
17886
cCas9-v42
 77
0





 803
GAAC
AAGAAGACAAAGGGTTTGTT
17887
SpyCas9-
 77
0






3var-NRRH







 804
GAAC
taAGAAGACAAAGGGTTTGTT
17888
iSpyMacCas9
 77
0





 805
CCTGG
CCACGTGAGCCTTGCTCGAGG
17889
SauCas9KKH
 78
0





 806
TG
TAAGAAGACAAAGGGTTTGT
17890
SpyCas9-NG
 78
0





 807
TG
TAAGAAGACAAAGGGTTTGT
17891
SpyCas9-
 78
0






xCas







 808
TG
TAAGAAGACAAAGGGTTTGT
17892
SpyCas9-
 78
0






xCas-NG







 809
TGA
TAAGAAGACAAAGGGTTTGT
17893
SpyCas9-SpG
 78
0





 810
TGA
TAAGAAGACAAAGGGTTTGT
17894
SpyCas9-
 78
0






SpRY







 811
CCT
CACGTGAGCCTTGCTCGAGG
17895
SpyCas9-
 78
0






SpRY







 812
TGAACTTG
cattAAGAAGACAAAGGGTTTGT
17896
BlatCas9
 78
0





 813
TGAAC
cattAAGAAGACAAAGGGTTTGT
17897
BlatCas9
 78
0





 814
TGAACT
TTAAGAAGACAAAGGGTTTGT
17898
cCas9-v16
 78
0





 815
TGAACT
TTAAGAAGACAAAGGGTTTGT
17899
cCas9-v21
 78
0





 816
TGAACTT
ATTAAGAAGACAAAGGGTTTGT
17900
CdiCas9
 78
0





 817
TGAA
TAAGAAGACAAAGGGTTTGT
17901
SpyCas9-
 78
0






3var-NRRH







 818
TGAA
TAAGAAGACAAAGGGTTTGT
17902
SpyCas9-VQR
 78
0





 819
TTGAA
tcATTAAGAAGACAAAGGGTTTG
17903
SauCas9
 79
0





 820
TTGAA
ATTAAGAAGACAAAGGGTTTG
17904
SauCas9KKH
 79
0





 821
TTG
TTAAGAAGACAAAGGGTTTG
17905
ScaCas9
 79
0





 822
TTG
TTAAGAAGACAAAGGGTTTG
17906
ScaCas9-HiFi-
 79
0






Sc++







 823
TTG
TTAAGAAGACAAAGGGTTTG
17907
ScaCas9-
 79
0






Sc++







 824
TTG
TTAAGAAGACAAAGGGTTTG
17908
SpyCas9-
 79
0






SpRY







 825
GCC
CCACGTGAGCCTTGCTCGAG
17909
SpyCas9-
 79
0






SpRY







 826
TTGAAC
ATTAAGAAGACAAAGGGTTTG
17910
cCas9-v17
 79
0





 827
TTGAAC
ATTAAGAAGACAAAGGGTTTG
17911
cCas9-v42
 79
0





 828
TTGAACT
CATTAAGAAGACAAAGGGTTTG
17912
CdiCas9
 79
0





 829
GTTGA
CATTAAGAAGACAAAGGGTTT
17913
SauCas9KKH
 80
0





 830
GG
TCCACGTGAGCCTTGCTCGA
17914
SpyCas9-NG
 80
0





 831
GG
TCCACGTGAGCCTTGCTCGA
17915
SpyCas9-
 80
0






xCas







 832
GG
TCCACGTGAGCCTTGCTCGA
17916
SpyCas9-
 80
0






xCas-NG







 833
GGC
TCCACGTGAGCCTTGCTCGA
17917
SpyCas9-SpG
 80
0





 834
GGC
TCCACGTGAGCCTTGCTCGA
17918
SpyCas9-
 80
0






SpRY







 835
GTT
ATTAAGAAGACAAAGGGTTT
17919
SpyCas9-
 80
0






SpRY







 836
GGCC
TCCACGTGAGCCTTGCTCGA
17920
SpyCas9-
 80
0






3var-NRCH







 837
AGG
GTCCACGTGAGCCTTGCTCG
17921
ScaCas9
 81
0





 838
AGG
GTCCACGTGAGCCTTGCTCG
17922
ScaCas9-HiFi-
 81
0






Sc++







 839
AGG
GTCCACGTGAGCCTTGCTCG
17923
ScaCas9-
 81
0






Sc++







 840
AGG
GTCCACGTGAGCCTTGCTCG
17924
SpyCas9
 81
0





 841
AGG
GTCCACGTGAGCCTTGCTCG
17925
SpyCas9-HF1
 81
0





 842
AGG
GTCCACGTGAGCCTTGCTCG
17926
SpyCas9-SpG
 81
0





 843
AGG
GTCCACGTGAGCCTTGCTCG
17927
SpyCas9-
 81
0






SpRY







 844
TG
CATTAAGAAGACAAAGGGTT
17928
SpyCas9-NG
 81
0





 845
TG
CATTAAGAAGACAAAGGGTT
17929
SpyCas9-
 81
0






xCas







 846
TG
CATTAAGAAGACAAAGGGTT
17930
SpyCas9-
 81
0






xCas-NG







 847
AG
GTCCACGTGAGCCTTGCTCG
17931
SpyCas9-NG
 81
0





 848
AG
GTCCACGTGAGCCTTGCTCG
17932
SpyCas9-
 81
0






xCas







 849
AG
GTCCACGTGAGCCTTGCTCG
17933
SpyCas9-
 81
0






xCas-NG







 850
TGT
CATTAAGAAGACAAAGGGTT
17934
SpyCas9-SpG
 81
0





 851
TGT
CATTAAGAAGACAAAGGGTT
17935
SpyCas9-
 81
0






SpRY







 852
AGGCCTGG
ggtgTCCACGTGAGCCTTGCTCG
17936
BlatCas9
 81
0





 853
AGGCC
ggtgTCCACGTGAGCCTTGCTCG
17937
BlatCas9
 81
0





 854
AGGC
GTCCACGTGAGCCTTGCTCG
17938
SpyCas9-
 81
0






3var-NRRH







 855
TGTT
CATTAAGAAGACAAAGGGTT
17939
SpyCas9-
 81
0






3var-NRTH







 856
GAGGCC
gaGGTGTCCACGTGAGCCTTGCTC
17940
Nme2Cas9
 82
0





 857
GAGG
GTGTCCACGTGAGCCTTGCTC
17941
SauriCas9
 82
0





 858
GAGG
GTGTCCACGTGAGCCTTGCTC
17942
SauriCas9-
 82
0






KKH







 859
TTG
TCATTAAGAAGACAAAGGGT
17943
ScaCas9
 82
0





 860
TTG
TCATTAAGAAGACAAAGGGT
17944
ScaCas9-HiFi-
 82
0






Sc++







 861
TTG
TCATTAAGAAGACAAAGGGT
17945
ScaCas9-
 82
0






Sc++







 862
TTG
TCATTAAGAAGACAAAGGGT
17946
SpyCas9-
 82
0






SpRY







 863
GAG
TGTCCACGTGAGCCTTGCTC
17947
ScaCas9
 82
0





 864
GAG
TGTCCACGTGAGCCTTGCTC
17948
ScaCas9-HiFi-
 82
0






Sc++







 865
GAG
TGTCCACGTGAGCCTTGCTC
17949
ScaCas9-
 82
0






Sc++







 866
GAG
TGTCCACGTGAGCCTTGCTC
17950
SpyCas9-
 82
0






SpRY







 867
GAGGCCTG
aggtGTCCACGTGAGCCTTGCTC
17951
BlatCas9
 82
0





 868
GAGGC
aggtGTCCACGTGAGCCTTGCTC
17952
BlatCas9
 82
0





 869
CGAGG
GGTGTCCACGTGAGCCTTGCT
17953
SauCas9KKH
 83
0





 870
CGAG
GGTGTCCACGTGAGCCTTGCT
17954
SauriCas9-
 83
0






KKH







 871
CGAG
GTGTCCACGTGAGCCTTGCT
17955
SpyCas9-VQR
 83
0





 872
CG
GTGTCCACGTGAGCCTTGCT
17956
SpyCas9-NG
 83
0





 873
CG
GTGTCCACGTGAGCCTTGCT
17957
SpyCas9-
 83
0






xCas







 874
CG
GTGTCCACGTGAGCCTTGCT
17958
SpyCas9-
 83
0






xCas-NG







 875
CGA
GTGTCCACGTGAGCCTTGCT
17959
SpyCas9-SpG
 83
0





 876
CGA
GTGTCCACGTGAGCCTTGCT
17960
SpyCas9-
 83
0






SpRY







 877
TTT
ATCATTAAGAAGACAAAGGG
17961
SpyCas9-
 83
0






SpRY







 878
CGAGGC
GGTGTCCACGTGAGCCTTGCT
17962
cCas9-v17
 83
0





 879
CGAGGC
GGTGTCCACGTGAGCCTTGCT
17963
cCas9-v42
 83
0





 880
GTTTGTT
gttCAATCATTAAGAAGACAAAGG
17964
PpnCas9
 84
0





 881
TCGAG
ggAGGTGTCCACGTGAGCCTTGC
17965
SauCas9
 84
0





 882
TCGAG
AGGTGTCCACGTGAGCCTTGC
17966
SauCas9KKH
 84
0





 883
TCG
GGTGTCCACGTGAGCCTTGC
17967
ScaCas9
 84
0





 884
TCG
GGTGTCCACGTGAGCCTTGC
17968
ScaCas9-HiFi-
 84
0






Sc++







 885
TCG
GGTGTCCACGTGAGCCTTGC
17969
ScaCas9-
 84
0






Sc++







 886
TCG
GGTGTCCACGTGAGCCTTGC
17970
SpyCas9-
 84
0






SpRY







 887
GTT
AATCATTAAGAAGACAAAGG
17971
SpyCas9-
 84
0






SpRY







 888
TCGAGG
AGGTGTCCACGTGAGCCTTGC
17972
cCas9-v17
 84
0





 889
TCGAGG
AGGTGTCCACGTGAGCCTTGC
17973
cCas9-v42
 84
0





 890
CTCGA
GAGGTGTCCACGTGAGCCTTG
17974
SauCas9KKH
 85
0





 891
GG
CAATCATTAAGAAGACAAAG
17975
SpyCas9-NG
 85
0





 892
GG
CAATCATTAAGAAGACAAAG
17976
SpyCas9-
 85
0






xCas







 893
GG
CAATCATTAAGAAGACAAAG
17977
SpyCas9-
 85
0






xCas-NG







 894
GGT
CAATCATTAAGAAGACAAAG
17978
SpyCas9-SpG
 85
0





 895
GGT
CAATCATTAAGAAGACAAAG
17979
SpyCas9-
 85
0






SpRY







 896
CTC
AGGTGTCCACGTGAGCCTTG
17980
SpyCas9-
 85
0






SpRY







 897
CTCGAG
GAGGTGTCCACGTGAGCCTTG
17981
cCas9-v17
 85
0





 898
CTCGAG
GAGGTGTCCACGTGAGCCTTG
17982
cCas9-v42
 85
0





 899
GGTT
CAATCATTAAGAAGACAAAG
17983
SpyCas9-
 85
0






3var-NRTH







 900
GGG
TCAATCATTAAGAAGACAAA
17984
ScaCas9
 86
0





 901
GGG
TCAATCATTAAGAAGACAAA
17985
ScaCas9-HiFi-
 86
0






Sc++







 902
GGG
TCAATCATTAAGAAGACAAA
17986
ScaCas9-
 86
0






Sc++







 903
GGG
TCAATCATTAAGAAGACAAA
17987
SpyCas9
 86
0





 904
GGG
TCAATCATTAAGAAGACAAA
17988
SpyCas9-HF1
 86
0





 905
GGG
TCAATCATTAAGAAGACAAA
17989
SpyCas9-SpG
 86
0





 906
GGG
TCAATCATTAAGAAGACAAA
17990
SpyCas9-
 86
0






SpRY







 907
GG
TCAATCATTAAGAAGACAAA
17991
SpyCas9-NG
 86
0





 908
GG
TCAATCATTAAGAAGACAAA
17992
SpyCas9-
 86
0






xCas







 909
GG
TCAATCATTAAGAAGACAAA
17993
SpyCas9-
 86
0






xCas-NG







 910
GCT
GAGGTGTCCACGTGAGCCTT
17994
SpyCas9-
 86
0






SpRY







 911
GGGT
TCAATCATTAAGAAGACAAA
17995
SpyCas9-
 86
0






3var-NRRH







 912
AGGG
GTTCAATCATTAAGAAGACAA
17996
SauriCas9
 87
0





 913
AGGG
GTTCAATCATTAAGAAGACAA
17997
SauriCas9-
 87
0






KKH







 914
AGG
TTCAATCATTAAGAAGACAA
17998
ScaCas9
 87
0





 915
AGG
TTCAATCATTAAGAAGACAA
17999
ScaCas9-HiFi-
 87
0






Sc++







 916
AGG
TTCAATCATTAAGAAGACAA
18000
ScaCas9-
 87
0






Sc++







 917
AGG
TTCAATCATTAAGAAGACAA
18001
SpyCas9
 87
0





 918
AGG
TTCAATCATTAAGAAGACAA
18002
SpyCas9-HF1
 87
0





 919
AGG
TTCAATCATTAAGAAGACAA
18003
SpyCas9-SpG
 87
0





 920
AGG
TTCAATCATTAAGAAGACAA
18004
SpyCas9-
 87
0






SpRY







 921
AG
TTCAATCATTAAGAAGACAA
18005
SpyCas9-NG
 87
0





 922
AG
TTCAATCATTAAGAAGACAA
18006
SpyCas9-
 87
0






xCas







 923
AG
TTCAATCATTAAGAAGACAA
18007
SpyCas9-
 87
0






xCas-NG







 924
TG
GGAGGTGTCCACGTGAGCCT
18008
SpyCas9-NG
 87
0





 925
TG
GGAGGTGTCCACGTGAGCCT
18009
SpyCas9-
 87
0






xCas







 926
TG
GGAGGTGTCCACGTGAGCCT
18010
SpyCas9-
 87
0






xCas-NG







 927
TGC
GGAGGTGTCCACGTGAGCCT
18011
SpyCas9-SpG
 87
0





 928
TGC
GGAGGTGTCCACGTGAGCCT
18012
SpyCas9-
 87
0






SpRY







 929
TGCTCGAG
ctggGAGGTGTCCACGTGAGCCT
18013
BlatCas9
 87
0





 930
TGCTC
ctggGAGGTGTCCACGTGAGCCT
18014
BlatCas9
 87
0





 931
AGGGTT
GTTCAATCATTAAGAAGACAA
18015
cCas9-v16
 87
0





 932
AGGGTT
GTTCAATCATTAAGAAGACAA
18016
cCas9-v21
 87
0





 933
TGCT
GGAGGTGTCCACGTGAGCCT
18017
SpyCas9-
 87
0






3var-NRCH







 934
AAGGGTT
tttTGTTCAATCATTAAGAAGACA
18018
PpnCas9
 88
0





 935
AAGGG
ttTGTTCAATCATTAAGAAGACA
18019
SauCas9
 88
0





 936
AAGGG
TGTTCAATCATTAAGAAGACA
18020
SauCas9KKH
 88
0





 937
AAGGGT
ttTGTTCAATCATTAAGAAGACA
18021
SauCas9
 88
0





 938
AAGGGT
TGTTCAATCATTAAGAAGACA
18022
SauCas9KKH
 88
0





 939
AAGGGT
TGTTCAATCATTAAGAAGACA
18023
cCas9-v17
 88
0





 940
AAGGGT
TGTTCAATCATTAAGAAGACA
18024
cCas9-v42
 88
0





 941
AAGG
TGTTCAATCATTAAGAAGACA
18025
SauriCas9
 88
0





 942
AAGG
TGTTCAATCATTAAGAAGACA
18026
SauriCas9-
 88
0






KKH







 943
AAG
GTTCAATCATTAAGAAGACA
18027
ScaCas9
 88
0





 944
AAG
GTTCAATCATTAAGAAGACA
18028
ScaCas9-HiFi-
 88
0






Sc++







 945
AAG
GTTCAATCATTAAGAAGACA
18029
ScaCas9-
 88
0






Sc++







 946
AAG
GTTCAATCATTAAGAAGACA
18030
SpyCas9-
 88
0






SpRY







 947
TTG
GGGAGGTGTCCACGTGAGCC
18031
ScaCas9
 88
0





 948
TTG
GGGAGGTGTCCACGTGAGCC
18032
ScaCas9-HiFi-
 88
0






Sc++







 949
TTG
GGGAGGTGTCCACGTGAGCC
18033
ScaCas9-
 88
0






Sc++







 950
TTG
GGGAGGTGTCCACGTGAGCC
18034
SpyCas9-
 88
0






SpRY







 951
AAGGGTTT
ttttGTTCAATCATTAAGAAGACA
18035
NmeCas9
 88
0





 952
AAAGG
TTGTTCAATCATTAAGAAGAC
18036
SauCas9KKH
 89
0





 953
AAAG
TTGTTCAATCATTAAGAAGAC
18037
SauriCas9-
 89
0






KKH







 954
AAAG
TGTTCAATCATTAAGAAGAC
18038
SpyCas9-
 89
0






QQR1







 955
AAAG
ttGTTCAATCATTAAGAAGAC
18039
iSpyMacCas9
 89
0





 956
AAA
TGTTCAATCATTAAGAAGAC
18040
SpyCas9-
 89
0






SpRY







 957
CTT
TGGGAGGTGTCCACGTGAGC
18041
SpyCas9-
 89
0






SpRY







 958
CTTGC
tcctGGGAGGTGTCCACGTGAGC
18042
BlatCas9
 89
0





 959
AAAGGG
TTGTTCAATCATTAAGAAGAC
18043
cCas9-v17
 89
0





 960
AAAGGG
TTGTTCAATCATTAAGAAGAC
18044
cCas9-v42
 89
0





 961
CAAAG
TTTGTTCAATCATTAAGAAGA
18045
SauCas9KKH
 90
0





 962
CAA
TTGTTCAATCATTAAGAAGA
18046
SpyCas9-
 90
0






SpRY







 963
CCT
CTGGGAGGTGTCCACGTGAG
18047
SpyCas9-
 90
0






SpRY







 964
CAAAGG
TTTGTTCAATCATTAAGAAGA
18048
cCas9-v17
 90
0





 965
CAAAGG
TTTGTTCAATCATTAAGAAGA
18049
cCas9-v42
 90
0





 966
CAAA
TTGTTCAATCATTAAGAAGA
18050
SpyCas9-
 90
0






3var-NRRH







 967
CAAA
ttTGTTCAATCATTAAGAAGA
18051
iSpyMacCas9
 90
0





 968
ACAAA
TTTTGTTCAATCATTAAGAAG
18052
SauCas9KKH
 91
0





 969
ACA
TTTGTTCAATCATTAAGAAG
18053
SpyCas9-
 91
0






SpRY







 970
GCC
CCTGGGAGGTGTCCACGTGA
18054
SpyCas9-
 91
0






SpRY







 971
ACAAAG
TTTTGTTCAATCATTAAGAAG
18055
cCas9-v17
 91
0





 972
ACAAAG
TTTTGTTCAATCATTAAGAAG
18056
cCas9-v42
 91
0





 973
GACAA
ATTTTGTTCAATCATTAAGAA
18057
SauCas9KKH
 92
0





 974
AG
TCCTGGGAGGTGTCCACGTG
18058
SpyCas9-NG
 92
0





 975
AG
TCCTGGGAGGTGTCCACGTG
18059
SpyCas9-
 92
0






xCas







 976
AG
TCCTGGGAGGTGTCCACGTG
18060
SpyCas9-
 92
0






xCas-NG







 977
GAC
TTTTGTTCAATCATTAAGAA
18061
SpyCas9-
 92
0






SpRY







 978
AGC
TCCTGGGAGGTGTCCACGTG
18062
SpyCas9-SpG
 92
0





 979
AGC
TCCTGGGAGGTGTCCACGTG
18063
SpyCas9-
 92
0






SpRY







 980
GACAAA
ATTTTGTTCAATCATTAAGAA
18064
cCas9-v17
 92
0





 981
GACAAA
ATTTTGTTCAATCATTAAGAA
18065
cCas9-v42
 92
0





 982
GACA
TTTTGTTCAATCATTAAGAA
18066
SpyCas9-
 92
0






3var-NRCH







 983
AGCC
TCCTGGGAGGTGTCCACGTG
18067
SpyCas9-
 92
0






3var-NRCH







 984
GAG
TTCCTGGGAGGTGTCCACGT
18068
ScaCas9
 93
0





 985
GAG
TTCCTGGGAGGTGTCCACGT
18069
ScaCas9-HiFi-
 93
0






Sc++







 986
GAG
TTCCTGGGAGGTGTCCACGT
18070
ScaCas9-
 93
0






Sc++







 987
GAG
TTCCTGGGAGGTGTCCACGT
18071
SpyCas9-
 93
0






SpRY







 988
AG
ATTTTGTTCAATCATTAAGA
18072
SpyCas9-NG
 93
0





 989
AG
ATTTTGTTCAATCATTAAGA
18073
SpyCas9-
 93
0






xCas







 990
AG
ATTTTGTTCAATCATTAAGA
18074
SpyCas9-
 93
0






xCas-NG







 991
AGA
ATTTTGTTCAATCATTAAGA
18075
SpyCas9-SpG
 93
0





 992
AGA
ATTTTGTTCAATCATTAAGA
18076
SpyCas9-
 93
0






SpRY







 993
GAGCCTTG
cgctTCCTGGGAGGTGTCCACGT
18077
BlatCas9
 93
0





 994
GAGCC
cgctTCCTGGGAGGTGTCCACGT
18078
BlatCas9
 93
0





 995
GAGCCTT
GCTTCCTGGGAGGTGTCCACGT
18079
CdiCas9
 93
0





 996
AGAC
ATTTTGTTCAATCATTAAGA
18080
SpyCas9-
 93
0






3var-NRRH







 997
AGAC
ATTTTGTTCAATCATTAAGA
18081
SpyCas9-VQR
 93
0





 998
GAGC
TTCCTGGGAGGTGTCCACGT
18082
SpyCas9-
 93
0






3var-NRRH







 999
AGACAA
ATTTTGTTCAATCATTAAGA
18083
St1Cas9-
 93
0






CNRZ1066







1000
TGAGCC
agCGCTTCCTGGGAGGTGTCCACG
18084
Nme2Cas9
 94
0





1001
TGAG
GCTTCCTGGGAGGTGTCCACG
18085
SauriCas9-
 94
0






KKH







1002
TGAG
CTTCCTGGGAGGTGTCCACG
18086
SpyCas9-VQR
 94
0





1003
AAG
TATTTTGTTCAATCATTAAG
18087
ScaCas9
 94
0





1004
AAG
TATTTTGTTCAATCATTAAG
18088
ScaCas9-HiFi-
 94
0






Sc++







1005
AAG
TATTTTGTTCAATCATTAAG
18089
ScaCas9-
 94
0






Sc++







1006
AAG
TATTTTGTTCAATCATTAAG
18090
SpyCas9-
 94
0






SpRY







1007
TG
CTTCCTGGGAGGTGTCCACG
18091
SpyCas9-NG
 94
0





1008
TG
CTTCCTGGGAGGTGTCCACG
18092
SpyCas9-
 94
0






xCas







1009
TG
CTTCCTGGGAGGTGTCCACG
18093
SpyCas9-
 94
0






xCas-NG







1010
TGA
CTTCCTGGGAGGTGTCCACG
18094
SpyCas9-SpG
 94
0





1011
TGA
CTTCCTGGGAGGTGTCCACG
18095
SpyCas9-
 94
0






SpRY







1012
AAGACAAA
tggtATTTTGTTCAATCATTAAG
18096
BlatCas9
 94
0





1013
AAGACAAA
tggtATTTTGTTCAATCATTAAG
18097
BlatCas9
 94
0





1014
AAGACAAA
tgGTATTTTGTTCAATCATTAAG
18098
GeoCas9
 94
0





1015
TGAGCCTT
gcgcTTCCTGGGAGGTGTCCACG
18099
BlatCas9
 94
0





1016
AAGAC
tggtATTTTGTTCAATCATTAAG
18100
BlatCas9
 94
0





1017
TGAGO
gcgcTTCCTGGGAGGTGTCCACG
18101
BlatCas9
 94
0





1018
AAGA
TATTTTGTTCAATCATTAAG
18102
SpyCas9-
 94
0






3var-NRRH







1019
GTGAG
agCGCTTCCTGGGAGGTGTCCAC
18103
SauCas9
 95
0





1020
GTGAG
CGCTTCCTGGGAGGTGTCCAC
18104
SauCas9KKH
 95
0





1021
GAAGA
GGTATTTTGTTCAATCATTAA
18105
SauCas9KKH
 95
0





1022
GAAG
GGTATTTTGTTCAATCATTAA
18106
SauriCas9-
 95
0






KKH







1023
GAAG
GTATTTTGTTCAATCATTAA
18107
SpyCas9-
 95
0






QQR1







1024
GAAG
ggTATTTTGTTCAATCATTAA
18108
iSpyMacCas9
 95
0





1025
GTG
GCTTCCTGGGAGGTGTCCAC
18109
ScaCas9
 95
0





1026
GTG
GCTTCCTGGGAGGTGTCCAC
18110
ScaCas9-HiFi-
 95
0






Sc++







1027
GTG
GCTTCCTGGGAGGTGTCCAC
18111
ScaCas9-
 95
0






Sc++







1028
GTG
GCTTCCTGGGAGGTGTCCAC
18112
SpyCas9-
 95
0






SpRY







1029
GAA
GTATTTTGTTCAATCATTAA
18113
SpyCas9-
 95
0






SpRY







1030
GAA
GTATTTTGTTCAATCATTAA
18114
SpyCas9-
 95
0






xCas







1031
GAAGAC
GGTATTTTGTTCAATCATTAA
18115
cCas9-v17
 95
0





1032
GAAGAC
GGTATTTTGTTCAATCATTAA
18116
cCas9-v42
 95
0





1033
GTGAGC
CGCTTCCTGGGAGGTGTCCAC
18117
cCas9-v17
 95
0





1034
GTGAGC
CGCTTCCTGGGAGGTGTCCAC
18118
cCas9-v42
 95
0





1035
AGAAG
TGGTATTTTGTTCAATCATTA
18119
SauCas9KKH
 96
0





1036
CGTGA
GCGCTTCCTGGGAGGTGTCCA
18120
SauCas9KKH
 96
0





1037
AG
GGTATTTTGTTCAATCATTA
18121
SpyCas9-NG
 96
0





1038
AG
GGTATTTTGTTCAATCATTA
18122
SpyCas9-
 96
0






xCas







1039
AG
GGTATTTTGTTCAATCATTA
18123
SpyCas9-
 96
0






xCas-NG







1040
CG
CGCTTCCTGGGAGGTGTCCA
18124
SpyCas9-NG
 96
0





1041
CG
CGCTTCCTGGGAGGTGTCCA
18125
SpyCas9-
 96
0






xCas







1042
CG
CGCTTCCTGGGAGGTGTCCA
18126
SpyCas9-
 96
0






xCas-NG







1043
AGA
GGTATTTTGTTCAATCATTA
18127
SpyCas9-SpG
 96
0





1044
AGA
GGTATTTTGTTCAATCATTA
18128
SpyCas9-
 96
0






SpRY







1045
CGT
CGCTTCCTGGGAGGTGTCCA
18129
SpyCas9-SpG
 96
0





1046
CGT
CGCTTCCTGGGAGGTGTCCA
18130
SpyCas9-
 96
0






SpRY







1047
AGAAGA
TGGTATTTTGTTCAATCATTA
18131
cCas9-v17
 96
0





1048
AGAAGA
TGGTATTTTGTTCAATCATTA
18132
cCas9-v42
 96
0





1049
AGAAGACA
acttGGTATTTTGTTCAATCATTA
18133
NmeCas9
 96
0





1050
AGAA
GGTATTTTGTTCAATCATTA
18134
SpyCas9-
 96
0






3var-NRRH







1051
AGAA
GGTATTTTGTTCAATCATTA
18135
SpyCas9-VQR
 96
0





1052
AAGAA
acTTGGTATTTTGTTCAATCATT
18136
SauCas9
 97
0





1053
AAGAA
TTGGTATTTTGTTCAATCATT
18137
SauCas9KKH
 97
0





1054
AAG
TGGTATTTTGTTCAATCATT
18138
ScaCas9
 97
0





1055
AAG
TGGTATTTTGTTCAATCATT
18139
ScaCas9-HiFi-
 97
0






Sc++







1056
AAG
TGGTATTTTGTTCAATCATT
18140
ScaCas9-
 97
0






Sc++







1057
AAG
TGGTATTTTGTTCAATCATT
18141
SpyCas9-
 97
0






SpRY







1058
ACG
GCGCTTCCTGGGAGGTGTCC
18142
ScaCas9
 97
0





1059
ACG
GCGCTTCCTGGGAGGTGTCC
18143
ScaCas9-HiFi-
 97
0






Sc++







1060
ACG
GCGCTTCCTGGGAGGTGTCC
18144
ScaCas9-
 97
0






Sc++







1061
ACG
GCGCTTCCTGGGAGGTGTCC
18145
SpyCas9-
 97
0






SpRY







1062
AAGAAG
TTGGTATTTTGTTCAATCATT
18146
cCas9-v17
 97
0





1063
AAGAAG
TTGGTATTTTGTTCAATCATT
18147
cCas9-v42
 97
0





1064
AAGA
TGGTATTTTGTTCAATCATT
18148
SpyCas9-
 97
0






3var-NRRH







1065
TAAGA
CTTGGTATTTTGTTCAATCAT
18149
SauCas9KKH
 98
0





1066
TAAG
CTTGGTATTTTGTTCAATCAT
18150
SauriCas9-
 98
0






KKH







1067
TAAG
TTGGTATTTTGTTCAATCAT
18151
SpyCas9-
 98
0






QQR1







1068
TAAG
ctTGGTATTTTGTTCAATCAT
18152
iSpyMacCas9
 98
0





1069
TAA
TTGGTATTTTGTTCAATCAT
18153
SpyCas9-
 98
0






SpRY







1070
CAC
AGCGCTTCCTGGGAGGTGTC
18154
SpyCas9-
 98
0






SpRY







1071
CACGTG
GAGCGCTTCCTGGGAGGTGTC
18155
cCas9-v16
 98
0





1072
CACGTG
GAGCGCTTCCTGGGAGGTGTC
18156
cCas9-v21
 98
0





1073
TAAGAA
CTTGGTATTTTGTTCAATCAT
18157
cCas9-v17
 98
0





1074
TAAGAA
CTTGGTATTTTGTTCAATCAT
18158
cCas9-v42
 98
0





1075
TTAAG
ACTTGGTATTTTGTTCAATCA
18159
SauCas9KKH
 99
0





1076
TTA
CTTGGTATTTTGTTCAATCA
18160
SpyCas9-
 99
0






SpRY







1077
CCA
GAGCGCTTCCTGGGAGGTGT
18161
SpyCas9-
 99
0






SpRY







1078
TTAAGA
ACTTGGTATTTTGTTCAATCA
18162
cCas9-v17
 99
0





1079
TTAAGA
ACTTGGTATTTTGTTCAATCA
18163
cCas9-v42
 99
0





1080
ATTAA
GACTTGGTATTTTGTTCAATC
18164
SauCas9KKH
100
0





1081
ATT
ACTTGGTATTTTGTTCAATC
18165
SpyCas9-
100
0






SpRY







1082
TCC
TGAGCGCTTCCTGGGAGGTG
18166
SpyCas9-
100
0






SpRY







1083
TCCACGTG
gagtGAGCGCTTCCTGGGAGGTG
18167
BlatCas9
100
0





1084
TCCAC
gagtGAGCGCTTCCTGGGAGGTG
18168
BlatCas9
100
0










In the exemplary template sequences provided herein, capital letters indicate “core nucleotides” while lower case letters indicate “flanking nucleotides.” Herein, when an RNA sequence (e.g., a template RNA sequence) is said to comprise a particular sequence (e.g., a sequence of Table 1 or a portion thereof) that comprises thymine (T), it is of course understood that the RNA sequence may (and frequently does) comprise uracil (U) in place of T. For instance, the RNA sequence may comprise U at every position shown as T in the sequence in Table 1. More specifically, the present disclosure provides an RNA sequence according to every gRNA spacer sequence shown in Table 1, wherein the RNA sequence has a U in place of each T in the sequence in Table 1.


In some embodiments of the systems and methods herein, the heterologous object sequence comprises the core nucleotides of an RT template sequence from Table 3. In some embodiments, the heterologous object sequence additionally comprises one or more (e.g., 2, 3, 4, 5, 10, 20, 30, 40, or all) consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence. In some embodiments, the heterologous object sequence comprises the core nucleotides of the RT template sequence of Table 3 that corresponds to the gRNA spacer sequence. In the context of the sequence tables, a first component “corresponds to” a second component when both components have the same ID number in the referenced table. For example, for a gRNA spacer of ID #1, the corresponding RT template would be the RT template also having ID #1. In some embodiments, the heterologous object sequence additionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the RT template sequence.


In some embodiments, the primer binding site (PBS) sequence has a sequence comprising the core nucleotides of a PBS sequence from the same row of Table 3 as the RT template sequence. In some embodiments, the PBS sequence additionally comprises one or more (e.g., 1, 2, 3, 4, 5, 6, 7, or all) consecutive nucleotides starting with the 5′ end of the flanking nucleotides of the primer region.









TABLE 3







Exemplary RT sequence (heterologous object sequence) and PBS sequence pairs


Table 3 provides exemplified PBS sequences and heterologous object


sequences (reverse transcription template regions) of a template RNA 


for correcting the pathogenic E342K mutation in SERPINA1. The gRNA


spacers from Table 1 were filtered, e.g., filtered by occurrence within 


15 nt of the desired editing location and use of a Tier 1 Cas enzyme.


PBS sequences and heterologous object sequences (reverse transcription


template regions) were designed relative to the nick site directed by 


the cognate gRNA from Table 1, as described in this application. For


exemplification, these regions were designed to be 8-17 nt (priming) 


and 1-50 nt extended beyond the location of the edit (RT). Without 


wishing to be limited by example, given variability of length, se-


quences are provided that use the maximum length parameters and


comprise all templates of shorter length within the given parameters. 


Sequences are shown with uppercase letters indicating core sequence


and lowercase letters indicating flanking sequence that may be


truncated within the described length parameters.













SEQ

SEQ




ID
PBS
ID


ID
RT template sequence
NO
sequence
NO





  1
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18169
GTCGATGGtca
18320





gcacag






  2
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18170
GTCGATGGtca
18321





gcacag






  5
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18171
GTCGATGGtca
18322





gcacag






  8
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18172
TCGATGGTcag
18323



G

cacagc






 11
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18173
GAAAGGGActg
18324



A

aagctg






 12
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18174
GAAAGGGActg
18325



A

aagctg






 13
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18175
TCGATGGTcag
18326



G

cacagc






 19
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18176
CGATGGTCagc
18327



GT

acagcc






 20
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18177
AAAGGGACtga
18328



AG

agctgc






 21
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18178
AAAGGGACtga
18329



AG

agctgc






 24
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18179
CGATGGTCagc
18330



GT

acagcc






 28
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18180
CGATGGTCagc
18331



GT

acagcc






 29
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18181
AAAGGGACtga
18332



AG

agctgc






 34
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18182
GATGGTCAgca
18333



GTC

cagcct






 35
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18183
GATGGTCAgca
18334



GTC

cagcct






 38
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18184
GATGGTCAgca
18335



GTC

cagcct






 39
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18185
GATGGTCAgca
18336



GTC

cagcct






 40
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18186
AAGGGACTgaa
18337



AGA

gctgct






 44
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18187
AAGGGACTgaa
18338



AGA

gctgct






 50
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18188
ATGGTCAGcac
18339



GTCG

agcctt






 51
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18189
ATGGTCAGcac
18340



GTCG

agcctt






 56
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18190
AGGGACTGaag
18341



AGAA

ctgctg






 57
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18191
AGGGACTGaag
18342



AGAA

ctgctg






 58
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18192
ATGGTCAGcac
18343



GTCG

agcctt






 59
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18193
ATGGTCAGcac
18344



GTCG

agcctt






 60
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18194
AGGGACTGaag
18345



AGAA

ctgctg






 61
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18195
AGGGACTGaag
18346



AGAA

ctgctg






 64
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18196
TGGTCAGCaca
18347



GTCGA

gcctta






 65
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18197
GGGACTGAagc
18348



AGAAA

tgctgg






 66
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18198
TGGTCAGCaca
18349



GTCGA

gcctta






 69
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18199
GGTCAGCAcag
18350



GTCGAT

ccttat






 70
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18200
GGTCAGCAcag
18351



GTCGAT

ccttat






 71
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18201
GGACTGAAgct
18352



AGAAAG

gctggg






 75
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18202
GTCAGCACagc
18353



GTCGATG

cttatg






 79
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18203
GTCAGCACagc
18354



GTCGATG

cttatg






 80
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18204
GACTGAAGctg
18355



AGAAAGG

ctgggg






 86
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18205
TCAGCACAgcc
18356



GTCGATGG

ttatgc






 87
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18206
TCAGCACAgcc
18357



GTCGATGG

ttatgc






 88
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18207
ACTGAAGCtgc
18358



AGAAAGGG

tggggc






 89
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18208
TCAGCACAgcc
18359



GTCGATGG

ttatgc






 90
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18209
TCAGCACAgcc
18360



GTCGATGG

ttatgc






 91
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18210
CAGCACAGcct
18361



GTCGATGGT

tatgca






 92
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18211
CTGAAGCTgct
18362



AGAAAGGGA

ggggcc






 93
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18212
CAGCACAGcct
18363



GTCGATGGT

tatgca






 94
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18213
CTGAAGCTgct
18364



AGAAAGGGA

ggggcc






 95
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18214
CTGAAGCTgct
18365



AGAAAGGGA

ggggcc






 98
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18215
AGCACAGCctt
18366



GTCGATGGTC

atgcac






 99
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18216
TGAAGCTGctg
18367



AGAAAGGGAC

gggcca






102
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18217
GAAGCTGCtgg
18368



AGAAAGGGACT

ggccat






103
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18218
GCACAGCCtta
18369



GTCGATGGTCA

tgcacg






104
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18219
GCACAGCCtta
18370



GTCGATGGTCA

tgcacg






105
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18220
AAGCTGCTggg
18371



AGAAAGGGACTG

gccatg






106
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18221
CACAGCCTtat
18372



GTCGATGGTCAG

gcacgg






107
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18222
ACAGCCTTatg
18373



GTCGATGGTCAGC

cacggc






108
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18223
AGCTGCTGggg
18374



AGAAAGGGACTGA

ccatgt






109
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18224
AGCTGCTGggg
18375



AGAAAGGGACTGA

ccatgt






110
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18225
AGCTGCTGggg
18376



AGAAAGGGACTGA

ccatgt






112
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18226
GCTGCTGGggc
18377



AGAAAGGGACTGAA

catgtt






113
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18227
GCTGCTGGggc
18378



AGAAAGGGACTGAA

catgtt






116
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18228
CAGCCTTAtgc
18379



GTCGATGGTCAGCA

acggcc






120
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18229
GCTGCTGGggc
18380



CAGAAAGGGATGAA

catgtt






122
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18230
CAGCCTTAtgc
18381



GTCGATGGTCAGCA

acggcc






123
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18231
GCTGCTGGggc
18382



AGAAAGGGACTGAA

catgtt






124
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18232
GCTGCTGGggc
18383



AGAAAGGGACTGAA

catgtt






125
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18233
CAGCCTTAtgc
18384



GTCGATGGTCAGCA

acggcc






130
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18234
CTGCTGGGgcc
18385



AGAAAGGGACTGAAG

atgttt






131
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18235
AGCCTTATgca
18386



GTCGATGGTCAGCAC

cggcct






134
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18236
CTGCTGGGgcc
18387



AGAAAGGGACTGAAG

atgttt






135
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18237
CTGCTGGGgcc
18388



AGAAAGGGACTGAAG

atgttt






138
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18238
AGCCTTATgca
18389



GTCGATGGTCAGCAC

cggcct






139
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18239
AGCCTTATgca
18390



GTCGATGGTCAGCAC

cggcct






140
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18240
AGCCTTATgca
18391



GTCGATGGTCAGCAC

cggcct






141
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18241
CTGCTGGGgcc
18392



AGAAAGGGACTGAAG

atgttt






142
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18242
AGCCTTATgca
18393



GTCGATGGTCAGCAC

cggcct






145
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18243
GCCTTATGcac
18394



GTCGATGGTCAGCACA

ggcctg






146
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18244
TGCTGGGGcca
18395



AGAAAGGGACTGAAGC

tgtttt






147
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18245
TGCTGGGGcca
18396



AGAAAGGGACTGAAGC

tgtttt






148
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18246
GCCTTATGcac
18397



GTCGATGGTCAGCACA

ggcctg






149
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18247
GCTGGGGCcat
18398



AGAAAGGGACTGAAGCT

gttttt






150
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18248
GCTGGGGCcat
18399



AGAAAGGGACTGAAGCT

gttttt






153
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18249
CCTTATGCacg
18400



GTCGATGGTCAGCACAG

gcctgg






157
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18250
CCTTATGCacg
18401



GTCGATGGTCAGCACAG

gcctgg






158
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18251
GCTGGGGCcat
18402



AGAAAGGGACTGAAGCT

gttttt






162
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18252
CTTATGCAcgg
18403



GTCGATGGTCAGCACAGC

cctgga






163
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18253
CTTATGCAcgg
18404



GTCGATGGTCAGCACAGC

cctgga






164
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18254
CTGGGGCCatg
18405



AGAAAGGGACTGAAGCTG

ttttta






165
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18255
TTATGCACggc
18406



GTCGATGGTCAGCACAGCC

ctggag






169
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18256
TTATGCACggc
18407



GTCGATGGTCAGCACAGCC

ctggag






170
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18257
TGGGGCCAtgt
18408



AGAAAGGGACTGAAGCTGC

ttttag






171
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18258
TGGGGCCAtgt
18409



AGAAAGGGACTGAAGCTGC

ttttag






172
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18259
TTATGCACggc
18410



GTCGATGGTCAGCACAGCC

ctggag






173
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18260
TGGGGCCAtgt
18411



AGAAAGGGACTGAAGCTGC

ttttag






174
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18261
TTATGCACggc
18412



GTCGATGGTCAGCACAGCC

ctggag






177
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18262
TATGCACGgcc
18413



TGTCGATGGCAGCACAGCCT

tggagg






178
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18263
TATGCACGgcc
18414



GTCGATGGTCAGCACAGCCT

tggagg






179
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18264
GGGGCCATgtt
18415



AGAAAGGGACTGAAGCTGCT

tttaga






183
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18265
GGGGCCATgtt
18416



AGAAAGGGACTGAAGCTGCT

tttaga






187
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18266
GGGCCATGttt
18417



AGAAAGGGACTGAAGCTGCTG

ttagag






188
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18267
GGGCCATGttt
18418



AGAAAGGGACTGAAGCTGCTG

ttagag






189
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18268
ATGCACGGcct
18419



GTCGATGGTCAGCACAGCCTT

ggaggg






192
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18269
GGCCATGTttt
18420



AGAAAGGGACTGAAGCTGCTGG

tagagg






193
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18270
TGCACGGCctg
18421



GTCGATGGTCAGCACAGCCTTA

gagggg






197
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18271
TGCACGGCctg
18422



GTCGATGGTCAGCACAGCCTTA

gagggg






198
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18272
GGCCATGTttt
18423



AGAAAGGGACTGAAGCTGCTGG

tagagg






199
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18273
GGCCATGTttt
18424



AGAAAGGGACTGAAGCTGCTGG

tagagg






203
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18274
GCCATGTTttt
18425



CAGAAAGGGATGAAGCTGCTGGG

agaggc






206
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18275
GCACGGCCtgg
18426



GTCGATGGTCAGCACAGCCTTAT

agggga






207
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18276
GCACGGCCtgg
18427



GTCGATGGTCAGCACAGCCTTAT

agggga






210
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18277
GCACGGCCtgg
18428



GTCGATGGTCAGCACAGCCTTAT

agggga






211
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18278
GCCATGTTttt
18429



AGAAAGGGACTGAAGCTGCTGGG

agaggc






214
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18279
GCACGGCCtgg
18430



GTCGATGGTCAGCACAGCCTTAT

agggga






218
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18280
GCCATGTTttt
18431



AGAAAGGGACTGAAGCTGCTGGG

agaggc






224
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18281
CACGGCCTgga
18432



GTCGATGGTCAGCACAGCCTTATG

ggggag






225
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18282
CACGGCCTgga
18433



GTCGATGGTCAGCACAGCCTTATG

ggggag






228
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18283
CCATGTTTtta
18434



AGAAAGGGACTGAAGCTGCTGGGG

gaggcc






229
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18284
CCATGTTTtta
18435



AGAAAGGGACTGAAGCTGCTGGGG

gaggcc






232
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18285
CACGGCCTgga
18436



GTCGATGGTCAGCACAGCCTTATG

ggggag






233
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18286
CACGGCCTgga
18437



GTCGATGGTCAGCACAGCCTTATG

ggggag






234
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18287
CACGGCCTgga
18438



GTCGATGGTCAGCACAGCCTTATG

ggggag






235
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18288
CACGGCCTgga
18439



GTCGATGGTCAGCACAGCCTTATG

ggggag






239
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18289
ACGGCCTGgag
18440



GTCGATGGTCAGCACAGCCTTATGC

gggaga






240
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18290
CATGTTTTtag
18441



AGAAAGGGACTGAAGCTGCTGGGGC

aggcca






241
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18291
ACGGCCTGgag
18442



GTCGATGGTCAGCACAGCCTTATGC

gggaga






244
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18292
ACGGCCTGgag
18443



GTCGATGGTCAGCACAGCCTTATGC

gggaga






245
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18293
CATGTTTTtag
18444



AGAAAGGGACTGAAGCTGCTGGGGC

aggcca






246
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18294
CATGTTTTtag
18445



AGAAAGGGACTGAAGCTGCTGGGGC

aggcca






249
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18295
ATGTTTTTaga
18446



AGAAAGGGACTGAAGCTGCTGGGGCC

ggccat






250
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18296
CGGCCTGGagg
18447



GTCGATGGTCAGCACAGCCTTATGCA

ggagag






251
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18297
ATGTTTTTaga
18448



AGAAAGGGACTGAAGCTGCTGGGGCC

ggccat






252
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18298
CGGCCTGGagg
18449



GTCGATGGTCAGCACAGCCTTATGCA

ggagag






257
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18299
GGCCTGGAggg
18450



GTCGATGGTCAGCACAGCCTTATGCAC

gagaga






258
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18300
GGCCTGGAggg
18451



GTCGATGGTCAGCACAGCCTTATGCAC

gagaga






259
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18301
TGTTTTTAgag
18452



CAGAAAGGGATGAAGCTGCTGGGGCCA

gccata






261
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18302
GTTTTTAGagg
18453



AGAAAGGGACTGAAGCTGCTGGGGCCAT

ccatac






262
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18303
GCCTGGAGggg
18454



GTCGATGGTCAGCACAGCCTTATGCACG

agagaa






263
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18304
GTTTTTAGagg
18455



AGAAAGGGACTGAAGCTGCTGGGGCCAT

ccatac






264
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18305
TTTTTAGAggc
18456



AGAAAGGGACTGAAGCTGCTGGGGCCATG

catacc






265
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18306
TTTTTAGAggc
18457



AGAAAGGGACTGAAGCTGCTGGGGCCATG

catacc






268
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18307
CCTGGAGGgga
18458



GTCGATGGTCAGCACAGCCTTATGCACGG

gagaag






272
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18308
TTTTTAGAggc
18459



AGAAAGGGACTGAAGCTGCTGGGGCCATG

catacc






274
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18309
CCTGGAGGgga
18460



GTCGATGGTCAGCACAGCCTTATGCACGG

gagaag






275
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18310
TTTTTAGAggc
18461



AGAAAGGGACTGAAGCTGCTGGGGCCATG

catacc






276
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18311
TTTTTAGAggc
18462



AGAAAGGGACTGAAGCTGCTGGGGCCATG

catacc






279
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18312
TTTTAGAGgcc
18463



AGAAAGGGACTGAAGCTGCTGGGGCCATGT

ataccc






282
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18313
TTTTAGAGgcc
18464



CTGAAGCTGCTGGGGCCATGT

ataccc






283
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18314
TTTTAGAGgcc
18465



CAGAAAGGGATGAAGCTGCTGGGGCCATGT

ataccc






286
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18315
TTTTAGAGgcc
18466



AGAAAGGGACTGAAGCTGCTGGGGCCATGT

ataccc






289
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18316
CTGGAGGGgag
18467



GTCGATGGTCAGCACAGCCTTATGAGAAAGGGACACGGC

agaagc






290
catgggtatggcctctaaaaacatggccccagcagcttcagtccctttcTC
18317
CTGGAGGGgag
18468



GTCGATGGTCAGCACAGCCTTATGCACGGC

agaagc






291
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18318
TTTTAGAGgcc
18469



CAGAAAGGGATGAAGCTGCTGGGGCCATGT

ataccc






294
tctgcttctctcccctccaggccgtgcataaggctgtgctgaccatcgaCG
18319
TTTTAGAGgcc
18470



CAGAAAGGGATGAAGCTGCTGGGGCCATGT

ataccc









Capital letters indicate “core nucleotides” while lower case letters indicate “flanking nucleotides.” Herein, when an RNA sequence (e.g., a template RNA sequence) is said to comprise a particular sequence (e.g., a sequence of Table 3 or a portion thereof) that comprises thymine (T), it is of course understood that the RNA sequence may (and frequently does) comprise uracil (U) in place of T. For instance, the RNA sequence may comprise U at every position shown as T in the sequence in Table 3. More specifically, the present disclosure provides an RNA sequence according to every heterologous object sequence and PBS sequence shown in Table 3, wherein the RNA sequence has a U in place of each T in the sequence of Table 3.


In some embodiments of the systems and methods herein, the template RNA comprises a gRNA scaffold (e.g., that binds a gene modifying polypeptide, e.g., a Cas polypeptide) that comprises a sequence of a gRNA scaffold of Table 12. In some embodiments, the gRNA scaffold comprises a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to a gRNA scaffold of Table 12. In some embodiments, the gRNA scaffold comprises a sequence of a scaffold region of Table 12 that corresponds to the RT template sequence, the spacer sequence, or both, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.


In some embodiments of the systems and methods herein, the system further comprises a second strand-targeting gRNA that directs a nick to the second strand of the human SERPINA1 gene. In some embodiments, the second strand-targeting gRNA comprises a left gRNA spacer sequence or a right gRNA spacer sequence from Table 2. In some embodiments, the gRNA spacer additionally comprises one or more (e.g., 2, 3, or all) consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the left gRNA spacer sequence or right gRNA spacer sequence. In some embodiments, the second strand-targeting gRNA comprises a sequence comprising the core nucleotides of a second nick gRNA sequence from Table 4, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto. In some embodiments, the second nick gRNA sequence additionally comprises one or more consecutive nucleotides starting with the 3′ end of the flanking nucleotides of the second nick gRNA sequence. In some embodiments, the second nick gRNA comprises a gRNA scaffold sequence that is orthogonal to the Cas domain of the gene modifying polypeptide. In some embodiments, the second nick gRNA comprises a gRNA scaffold sequence of Table 12.









TABLE 2







Exemplary left gRNA spacer and right gRNA spacer pairs


Table 2 provides exemplified second strand-targeting gRNA species for 


optional use for correcting the pathogenic E342K mutation in SERPINA1.


The gRNA spacers from Table 1 were filtered, e.g., filtered by occur-


rence within 15 nt of the desired editing location and use of a Tier 


1 Cas enzyme. Second strand-targeting gRNAs were generated by search-


ing the opposite strand of DNA in the regions −40 to −140 (“left”)


and +40 to +140 (“right”), relative to the first nick site defined 


by the first gRNA, for the PAM utilized by the corresponding Cas


variant. One exemplary spacer is shown for each side of the target 


nick site.















SEQ
 

SEQ





ID
left

ID
right


ID
left gRNA spacer
NO
pam
right gRNA spacer
NO
pam





  1
ACTTGGTATTTTGTTCAATCA
1847
TTAAG
CAAGGCTCACGTGGACACCTC
1877
CCAGG




   1


   3






  2
CTTGGTATTTTGTTCAATCAT
1847
TAAG
CAAGGCTCACGTGGACACCTC
1877
CCAG




   2


   4






  5
CTTGGTATTTTGTTCAATCA
1847
TTA
AAGGCTCACGTGGACACCTC
1877
CCA




   3


   5






  8
CTTGGTATTTTGTTCAATCAT
1847
TAAGA
AAGGCTCACGTGGACACCTCC
1877
CAGGA




   4


   6






 11
ACCCTTTGTCTTCTTAATGA
1847
TTG
GCGCTTCCTGGGAGGTGTCC
1877
ACG




   5


   7






 12
TTGTCTTCTTAATGATTGAA
1847
CAA
AGCGCTTCCTGGGAGGTGTC
1877
CAC




   6


   8






 13
TTGGTATTTTGTTCAATCAT
1847
TAA
AGGCTCACGTGGACACCTCC
1877
CAG




   7


   9






 19
TTGGTATTTTGTTCAATCATT
1847
AAGAA
AGGCTCACGTGGACACCTCCC
1878
AGGAA




   8


   0






 20
TTTGTCTTCTTAATGATTGAA
1847
CAAAA
GCGCTTCCTGGGAGGTGTCCA
1878
CGTGA




   9


   1






 21
TTTGTCTTCTTAATGATTGAA
1848
CAAAA
GCGCTTCCTGGGAGGTGTCCA
1878
CGTGA




   0


   2






 24
GGTATTTTGTTCAATCATTA
1848
AG
GGCTCACGTGGACACCTCCC
1878
AG




   1


   3






 28
TGGTATTTTGTTCAATCATT
1848
AAG
GGCTCACGTGGACACCTCCC
1878
AGG




   2


   4






 29
TGTCTTCTTAATGATTGAAC
1848
AAA
GCGCTTCCTGGGAGGTGTCC
1878
ACG




   3


   5






 34
acTTGGTATTTTGTTCAATCATT
1848
AAGAA
caAGGCTCACGTGGACACCTCC
1878
AGGAA




   4

C
   6






 35
TGGTATTTTGTTCAATCATTA
1848
AGAAG
GGCTCACGTGGACACCTCCCA
1878
GGAAG




   5


   7






 38
TGGTATTTTGTTCAATCATT
1848
AAG
GGCTCACGTGGACACCTCCC
1878
AGG




   6


   8






 39
GGTATTTTGTTCAATCATTA
1848
AGA
GCTCACGTGGACACCTCCCA
1878
GGA




   7


   9






 40
CCCTTTGTCTTCTTAATGAT
1848
TG
CGCTTCCTGGGAGGTGTCCA
1879
CG




   8


   0






 44
GTCTTCTTAATGATTGAACA
1848
AAA
CGCTTCCTGGGAGGTGTCCA
1879
CGT




   9


   1






 50
GGTATTTTGTTCAATCATTAA
1849
GAAGA
GGCTCACGTGGACACCTCCCA
1879
GGAAG




   0


   2






 51
GGTATTTTGTTCAATCATTAA
1849
GAAG
GCTCACGTGGACACCTCCCAG
1879
GAAG




   1


   3






 56
TAATGATTGAACAAAATACC
1849
AAG
GCTTCCTGGGAGGTGTCCAC
1879
GTG




   2


   4






 57
TCTTCTTAATGATTGAACAA
1849
AAT
GCTTCCTGGGAGGTGTCCAC
1879
GTG




   3


   5






 58
GTATTTTGTTCAATCATTAA
1849
GAA
CTCACGTGGACACCTCCCAG
1879
GAA




   4


   6






 59



AAGGCTCACGTGGACACCTC
1879
CCAGGAA







   7






 60
ttgtCTTCTTAATGATTGAACAA
1849
AATAC
gcgcTTCCTGGGAGGTGTCCAC
1879
TGAGCCT




   6

G
   8
T





 61
ttgtCTTCTTAATGATTGAACAA
1849
AATAC
gcgcTTCCTGGGAGGTGTCCAC
1879
TGAGCCT




   7

G
   9
T





 64
GGTATTTTGTTCAATCATTAA
1849
GAAGA
GGCTCACGTGGACACCTCCCA
1880
GGAAG




   8


   0






 65
CTTCTTAATGATTGAACAAA
1849
ATA
CTTCCTGGGAGGTGTCCACG
1880
TGA




   9


   1






 66
TATTTTGTTCAATCATTAAG
1850
AAG
TCACGTGGACACCTCCCAGG
1880
AAG




   0


   2






 69
ATTTTGTTCAATCATTAAGAA
1850
GACAA
GGCTCACGTGGACACCTCCCA
1880
GGAAG




   1


   3






 70
ATTTTGTTCAATCATTAAGA
1850
AGA
CACGTGGACACCTCCCAGGA
1880
AGC




   2


   4






 71
TTCTTAATGATTGAACAAAA
1850
TAC
TTCCTGGGAGGTGTCCACGT
1880
GAG




   3


   5






 75
ATTTTGTTCAATCATTAAGA
1850
AG
CACGTGGACACCTCCCAGGA
1880
AG




   4


   6






 79
TTTTGTTCAATCATTAAGAA
1850
GAC
ACGTGGACACCTCCCAGGAA
1880
GCG




   5


   7






 80
TCTTAATGATTGAACAAAAT
1850
ACC
TCCTGGGAGGTGTCCACGTG
1880
AGC




   6


   8






 86
TATTTTGTTCAATCATTAAG
1850
AAG
ACGTGGACACCTCCCAGGAA
1880
GCG




   7


   9






 87
TTTGTTCAATCATTAAGAAG
1850
ACA
CGTGGACACCTCCCAGGAAG
1881
CGC




   8


   0






 88
CTTAATGATTGAACAAAATA
1850
CCA
CCTGGGAGGTGTCCACGTGA
1881
GCC




   9


   1






 89
tggtATTTTGTTCAATCATTAAG
1851
AAGACAA
tcacGTGGACACCTCCCAGGAA
1881
CGCTC




   0
A
G
   2






 90
tggtATTTTGTTCAATCATTAAG
1851
AAGACAA
tcacGTGGACACCTCCCAGGAA
1881
CGCTC




   1
A
G
   3






 91
TTTGTTCAATCATTAAGAAGA
1851
CAAAG
GGCTCACGTGGACACCTCCCA
1881
GGAAG




   2


   4






 92
TTAATGATTGAACAAAATAC
1851
CAA
CTGGGAGGTGTCCACGTGAG
1881
CCT




   3


   5






 93
TTGTTCAATCATTAAGAAGA
1851
CAA
GTGGACACCTCCCAGGAAGC
1881
GCT




   4


   6






 94
tcttAATGATTGAACAAAATACC
1851
AAGTC
tcctGGGAGGTGTCCACGTGAG
1881
CTTGC




   5

C
   7






 95
tcttAATGATTGAACAAAATACC
1851
AAGTC
tcctGGGAGGTGTCCACGTGAG
1881
CTTGC




   6

C
   8






 98
TGTTCAATCATTAAGAAGAC
1851
AAA
TGGACACCTCCCAGGAAGCG
1881
CTC




   7


   9






 99
TAATGATTGAACAAAATACC
1851
AAG
TGGGAGGTGTCCACGTGAGC
1882
CTT




   8


   0






102
AATGATTGAACAAAATACCA
1851
AGT
GGGAGGTGTCCACGTGAGCC
1882
TTG




   9


   1






103
GTTCAATCATTAAGAAGACA
1852
AAG
GGACACCTCCCAGGAAGCGC
1882
TCA




   0


   2






104
tggtATTTTGTTCAATCATTAAG
1852
AAGACAA
acgtGGACACCTCCCAGGAAGC
1882
CTCAC




   1
A
G
   3






105
ATGATTGAACAAAATACCAA
1852
GTC
GGAGGTGTCCACGTGAGCCT
1882
TGC




   2


   4






106
TTCAATCATTAAGAAGACAA
1852
AGG
GACACCTCCCAGGAAGCGCT
1882
CAC




   3


   5






107
TCAATCATTAAGAAGACAAA
1852
GGG
ACACCTCCCAGGAAGCGCTC
1882
ACT




   4


   6






108
TGATTGAACAAAATACCAAG
1852
TCT
GAGGTGTCCACGTGAGCCTT
1882
GCT




   5


   7






109
taatGATTGAACAAAATACCAA
1852
TCTCC
ctggGAGGTGTCCACGTGAGCC
1882
TGCTC



G
   6

T
   8






110
taatGATTGAACAAAATACCAA
1852
TCTCC
ctggGAGGTGTCCACGTGAGCC
1882
TGCTC



G
   7

T
   9






112
taATGATTGAACAAAATACCAA
1852
CTCCCC
gaGGTGTCCACGTGAGCCTTG
1883
GAGGCC



GT
   8

CTC
   0






113
AATGATTGAACAAAATACCA
1852
AG
GGAGGTGTCCACGTGAGCCT
1883
TG




   9


   1






116
CAATCATTAAGAAGACAAAG
1853
GG
CGTGGACACCTCCCAGGAAG
1883
CG




   0


   2






120
GATTGAACAAAATACCAAGT
1853
CTC
AGGTGTCCACGTGAGCCTTG
1883
CTC




   1


   3






122
CAATCATTAAGAAGACAAAG
1853
GGT
CACCTCCCAGGAAGCGCTCA
1883
CTC




   2


   4






123
aatgATTGAACAAAATACCAAG
1853
CTCCC
ctggGAGGTGTCCACGTGAGCC
1883
TGCTC



T
   3

T
   5






124
aatgATTGAACAAAATACCAAG
1853
CTCCC
ctggGAGGTGTCCACGTGAGCC
1883
TGCTC



T
   4

T
   6






125
cattAAGAAGACAAAGGGTTTG
1853
TGAACTT
ggacACCTCCCAGGAAGCGCTC
1883
CTCCC



T
   5
G
A
   7






130
taATGATTGAACAAAATACCAA
1853
CTCCCC
gaGGTGTCCACGTGAGCCTTG
1883
GAGGCC



GT
   6

CTC
   8






131
aaGAAGACAAAGGGTTTGTTG
1853
TTGACC
ggACACCTCCCAGGAAGCGCT
1883
TCCCCC



AAC
   7

CAC
   9






134
TAATGATTGAACAAAATACC
1853
AAG
GGTGTCCACGTGAGCCTTGC
1884
TCG




   8


   0






135
ATTGAACAAAATACCAAGTC
1853
TCC
GGTGTCCACGTGAGCCTTGC
1884
TCG




   9


   1






138
TCATTAAGAAGACAAAGGGT
1854
TTG
CCAGGAAGCGCTCACTCCCC
1884
CTG




   0


   2






139
AATCATTAAGAAGACAAAGG
1854
GTT
ACCTCCCAGGAAGCGCTCAC
1884
TCC




   1


   3






140
cattAAGAAGACAAAGGGTTTG
1854
TGAACTT
gacaCCTCCCAGGAAGCGCTCA
1884
TCCCC



T
   2
G
C
   4






141
atgaTTGAACAAAATACCAAGT
1854
TCCCC
aggtGTCCACGTGAGCCTTGCT
1884
GAGGC



C
   3

C
   5






142
cattAAGAAGACAAAGGGTTTG
1854
TGAACTT
gacaCCTCCCAGGAAGCGCTCA
1884
TCCCC



T
   4
G
C
   6






145
CATTAAGAAGACAAAGGGTTT
1854
GTTGA
TCCCAGGAAGCGCTCACTCCC
1884
CCTGG




   5


   7






146
CTTAATGATTGAACAAAATAC
1854
CAAG
GGTGTCCACGTGAGCCTTGCT
1884
CGAG




   6


   8






147
TTGAACAAAATACCAAGTCT
1854
CCC
GTGTCCACGTGAGCCTTGCT
1884
CGA




   7


   9






148
ATCATTAAGAAGACAAAGGG
1854
TT
CCTCCCAGGAAGCGCTCACT
1885
CCC




   8


   0






149
AAATACCAAGTCTCCCCTCTT
1854
CATGG
GGTGTCCACGTGAGCCTTGCT
1885
CGAGG




   9


   1






150
AAATACCAAGTCTCCCCTCTT
1855
CATGG
GGTGTCCACGTGAGCCTTGCT
1885
CGAGG




   0


   2






153
CATTAAGAAGACAAAGGGTT
1855
TG
CAGGAAGCGCTCACTCCCCC
1885
TG




   1


   3






157
TCATTAAGAAGACAAAGGGT
1855
TTG
CTCCCAGGAAGCGCTCACTC
1885
CCC




   2


   4






158
TGAACAAAATACCAAGTCTC
1855
CCC
TGTCCACGTGAGCCTTGCTC
1885
GAG




   3


   5






162
TCATTAAGAAGACAAAGGGT
1855
TTG
CCAGGAAGCGCTCACTCCCC
1885
CTG




   4


   6






163
CATTAAGAAGACAAAGGGTT
1855
TGT
TCCCAGGAAGCGCTCACTCC
1885
CCC




   5


   7






164
GAACAAAATACCAAGTCTCC
1855
CCT
GTCCACGTGAGCCTTGCTCG
1885
AGG




   6


   8






165
CATTAAGAAGACAAAGGGTT
1855
TG
CAGGAAGCGCTCACTCCCCC
1885
TG




   7


   9






169
ATTAAGAAGACAAAGGGTTT
1855
GTT
CCCAGGAAGCGCTCACTCCC
1886
CCT




   8


   0






170
AACAAAATACCAAGTCTCCC
1855
CTC
TCCACGTGAGCCTTGCTCGA
1886
GGC




   9


   1






171
tgaaCAAAATACCAAGTCTCCCC
1856
TCTTCAT
ggtgTCCACGTGAGCCTTGCTC
1886
AGGCCTG




   0
G
G
   2
G





172
cattAAGAAGACAAAGGGTTTG
1856
TGAACTT
tcccAGGAAGCGCTCACTCCCC
1886
TGGAC



T
   1
G
C
   3






173
tgaaCAAAATACCAAGTCTCCCC
1856
TCTTCAT
ggtgTCCACGTGAGCCTTGCTC
1886
AGGCCTG




   2
G
G
   4
G





174
cattAAGAAGACAAAGGGTTTG
1856
TGAACTT
tcccAGGAAGCGCTCACTCCCC
1886
TGGAC



T
   3
G
C
   5






177
TTAAGAAGACAAAGGGTTTG
1856
TTG
CCAGGAAGCGCTCACTCCCC
1886
CTG




   4


   6






178
TTAAGAAGACAAAGGGTTTG
1856
TTG
CCAGGAAGCGCTCACTCCCC
1886
CTG




   5


   7






179
TACCAAGTCTCCCCTCTTCA
1856
TG
TCCACGTGAGCCTTGCTCGA
1886
GG




   6


   8






183
ACAAAATACCAAGTCTCCCC
1856
TCT
CCACGTGAGCCTTGCTCGAG
1886
GCC




   7


   9






187
ATACCAAGTCTCCCCTCTTC
1856
ATG
ACGTGAGCCTTGCTCGAGGC
1887
CTG




   8


   0






188
CAAAATACCAAGTCTCCCCT
1856
CTT
CACGTGAGCCTTGCTCGAGG
1887
CCT




   9


   1






189
TAAGAAGACAAAGGGTTTGT
1857
TGA
CAGGAAGCGCTCACTCCCCC
1887
TGG




   0


   2






192
AATACCAAGTCTCCCCTCTTC
1857
ATGG
CACGTGAGCCTTGCTCGAGGC
1887
CTGG




   1


   3






193
TAAGAAGACAAAGGGTTTGT
1857
TG
AGGAAGCGCTCACTCCCCCT
1887
GG




   2


   4






197
AAGAAGACAAAGGGTTTGTT
1857
GAA
AGGAAGCGCTCACTCCCCCT
1887
GGA




   3


   5






198
AAAATACCAAGTCTCCCCTC
1857
TTC
ACGTGAGCCTTGCTCGAGGC
1887
CTG




   4


   6






199
tgaaCAAAATACCAAGTCTCCCC
1857
TCTTCAT
acgtGAGCCTTGCTCGAGGCCT
1887
GGATC




   5
G
G
   7






203
AAATACCAAGTCTCCCCTCTT
1857
CATGG
ACGTGAGCCTTGCTCGAGGCC
1887
TGGGAT




   6


   8






206
TTAAGAAGACAAAGGGTTTG
1857
TTG
GAAGCGCTCACTCCCCCTGG
1887
ACG




   7


   9






207
GGGTTTGTTGAACTTGACCT
1857
CGG
CAGGAAGCGCTCACTCCCCC
1888
TGG




   8


   0






210
AGAAGACAAAGGGTTTGTTG
1857
AAC
GGAAGCGCTCACTCCCCCTG
1888
GAC




   9


   1






211
TACCAAGTCTCCCCTCTTCA
1858
TG
CGTGAGCCTTGCTCGAGGCC
1888
TG




   0


   2






214
TAAGAAGACAAAGGGTTTGT
1858
TG
AGGAAGCGCTCACTCCCCCT
1888
GG




   1


   3






218
AAATACCAAGTCTCCCCTCT
1858
TCA
CGTGAGCCTTGCTCGAGGCC
1888
TGG




   2


   4






224
AAGGGTTTGTTGAACTTGACC
1858
TCGG
GGAAGCGCTCACTCCCCCTGG
1888
ACGG




   3


   5






225
AAGGGTTTGTTGAACTTGACC
1858
TCGG
GGAAGCGCTCACTCCCCCTGG
1888
ACGG




   4


   6






228
ATACCAAGTCTCCCCTCTTC
1858
ATG
GTGAGCCTTGCTCGAGGCCT
1888
GGG




   5


   7






229
AATACCAAGTCTCCCCTCTT
1858
CAT
GTGAGCCTTGCTCGAGGCCT
1888
GGG




   6


   8






232
AGACAAAGGGTTTGTTGAAC
1858
TTG
GAAGCGCTCACTCCCCCTGG
1888
ACG




   7


   9






233
GAAGACAAAGGGTTTGTTGA
1858
ACT
GAAGCGCTCACTCCCCCTGG
1889
ACG




   8


   0






234
agaaGACAAAGGGTTTGTTGAA
1858
TTGAC
caggAAGCGCTCACTCCCCCTG
1889
ACGGC



C
   9

G
   1






235
agaaGACAAAGGGTTTGTTGAA
1859
TTGAC
caggAAGCGCTCACTCCCCCTG
1889
ACGGC



C
   0

G
   2






239
GAAGACAAAGGGTTTGTTGAA
1859
CTTGA
AGGAAGCGCTCACTCCCCCTG
1889
GACGG




   1


   3






240
AATACCAAGTCTCCCCTCTTC
1859
ATGG
ACGTGAGCCTTGCTCGAGGCC
1889
TGGG




   2


   4






241
AAGGGTTTGTTGAACTTGACC
1859
TCGG
GGAAGCGCTCACTCCCCCTGG
1889
ACGG




   3


   5






244
AAGACAAAGGGTTTGTTGAA
1859
CTT
AAGCGCTCACTCCCCCTGGA
1889
CGG




   4


   6






245
ATACCAAGTCTCCCCTCTTC
1859
ATG
TGAGCCTTGCTCGAGGCCTG
1889
GGA




   5


   7






246
tgaaCAAAATACCAAGTCTCCCC
1859
TCTTCAT
acgtGAGCCTTGCTCGAGGCCT
1889
GGATC




   6
G
G
   8






249
ATACCAAGTCTCCCCTCTTCA
1859
TGGGA
GAGCCTTGCTCGAGGCCTGGG
1889
ATCAG




   7


   9






250
GAAGACAAAGGGTTTGTTGAA
1859
CTTGA
CGCTCACTCCCCCTGGACGGC
1890
CCTGG




   8


   0






251
TACCAAGTCTCCCCTCTTCA
1859
TGG
GAGCCTTGCTCGAGGCCTGG
1890
GAT




   9


   1






252
AGACAAAGGGTTTGTTGAAC
1860
TTG
AGCGCTCACTCCCCCTGGAC
1890
GGC




   0


   2






257
GAAGACAAAGGGTTTGTTGAA
1860
CTTGA
CGCTCACTCCCCCTGGACGGC
1890
CCTGG




   1


   3






258
GACAAAGGGTTTGTTGAACT
1860
TGA
GCGCTCACTCCCCCTGGACG
1890
GCC




   2


   4






259
ACCAAGTCTCCCCTCTTCAT
1860
GGG
AGCCTTGCTCGAGGCCTGGG
1890
ATC




   3


   5






261
CCAAGTCTCCCCTCTTCATG
1860
GGA
GCCTTGCTCGAGGCCTGGGA
1890
TCA




   4


   6






262
ACAAAGGGTTTGTTGAACTT
1860
GAC
CGCTCACTCCCCCTGGACGG
1890
CCC




   5


   7






263
tgaaCAAAATACCAAGTCTCCCC
1860
TCTTCAT
tgagCCTTGCTCGAGGCCTGGG
1890
TCAGC




   6
G
A
   8






264
tcCCCTCTTCATGGGAAAAGTG
1860
GAATCC
gtGAGCCTTGCTCGAGGCCTG
1890
TCAGCC



GT
   7

GGA
   9






265
CCAAGTCTCCCCTCTTCATG
1860
GG
CTTGCTCGAGGCCTGGGATC
1891
AG




   8


   0






268
GACAAAGGGTTTGTTGAACT
1860
TG
TCACTCCCCCTGGACGGCCC
1891
TG




   9


   1






272
CAAGTCTCCCCTCTTCATGG
1861
GAA
CCTTGCTCGAGGCCTGGGAT
1891
CAG




   0


   2






274
CAAAGGGTTTGTTGAACTTG
1861
ACC
GCTCACTCCCCCTGGACGGC
1891
CCT




   1


   3






275
tgaaCAAAATACCAAGTCTCCCC
1861
TCTTCAT
gagcCTTGCTCGAGGCCTGGGA
1891
CAGCCTT




   2
G
T
   4
A





276
tgaaCAAAATACCAAGTCTCCCC
1861
TCTTCAT
gagcCTTGCTCGAGGCCTGGGA
1891
CAGCCTT




   3
G
T
   5
A





279
tcCCCTCTTCATGGGAAAAGTG
1861
GAATCC
gtGAGCCTTGCTCGAGGCCTG
1891
TCAGCC



GT
   4

GGA
   6






282
GTCTCCCCTCTTCATGGGAA
1861
AAG
CCTTGCTCGAGGCCTGGGAT
1891
CAG




   5


   7






283
ACCAAGTCTCCCCTCTTCAT
1861
GGG
GTGAGCCTTGCTCGAGGCCT
1891
GGG




   6


   8






286
AAGTCTCCCCTCTTCATGGG
1861
AAA
CTTGCTCGAGGCCTGGGATC
1891
AGC




   7


   9






289
AGGGTTTGTTGAACTTGACC
1861
TCG
CTCACTCCCCCTGGACGGCC
1892
CTG




   8


   0






290
AAAGGGTTTGTTGAACTTGA
1861
CCT
CTCACTCCCCCTGGACGGCC
1892
CTG




   9


   1






291
CCAAGTCTCCCCTCTTCATG
1862
GG
CTTGCTCGAGGCCTGGGATC
1892
AG




   0


   2






294
ccccTCTTCATGGGAAAAGTGG
1862
GAATC
gagcCTTGCTCGAGGCCTGGGA
1892
CAGCCTT



T
   1

T
   3
A









Capital letters indicate “core nucleotides” while lower case letters indicate “flanking nucleotides.” Herein, when an RNA sequence (e.g., a gRNA to produce a second nick) is said to comprise a particular sequence (e.g., a sequence of Table 2 or a portion thereof) that comprises thymine (T), it is of course understood that the RNA sequence may (and frequently does) comprise uracil (U) in place of T. For instance, the RNA sequence may comprise U at every position shown as T in the sequence in Table 2. More specifically, the present disclosure provides an RNA sequence according to every gRNA spacer sequence shown in Table 2, wherein the RNA sequence has a U in place of each T in the sequence in Table 2.


In some embodiments, the systems and methods provided herein may comprise a template sequence listed in Table 4. Table 4 provides exemplary template RNA sequences (column 4) and optional second strand-targeting gRNA sequences (column 5) designed to be paired with a gene modifying polypeptide to correct a mutation in the SERPINA1 gene. The templates in Table 4 are meant to exemplify the total sequence of: (1) gRNA spacer (e.g., for targeting for first strand nick), (2) gRNA scaffold, (3) heterologous object sequence, and (4) PBS sequence (e.g., for initiating TPRT at first strand nick).









TABLE 4







Exemplary template RNA sequences and second nick gRNA sequences


Table 4 provides design of RNA components of gene modifying systems for correcting


the pathogenic E342K mutation in SERPINA1. The gRNA spacers from Table 1 were


filtered, e.g., filtered by occurrence within 15 nt of the desired editing location


and use of a Tier 1 Cas enzyme. For each gRNA ID, this table details the sequence of


a complete template RNA, optional second strand-targeting gRNA, and Cas variant for


use in a Cas-RT fusion gene modifying polypeptide. For exemplification, PBS sequences


and post-edit homology regions (after the location of the edit) are set to 12 nt and


30 nt, respectively. Additionally, a second strand-targeting gRNA is selected with


preference for a distance near 100 nt from the first nick and a first preference for


a design resulting in a PAM-in system, as described elsewhere in this application.














Cas


SEQ

SEQ


ID
species
Strand
Template RNA
ID NO
Second strand-targeting gRNA
ID NO
















1
SauCas9
−
GCTGTGCTGACCATCGACAAGGTTTTAGTACTCTGGAAACAGA
19075
ACTTGGTATTTTGTTCAATCAGTTTTAGT
19226



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAcatggccccagcagcttcagtccctttcTCGT

GCAAAATGCCGTGTTTATCTCGTCAACTT






CGATGGtcag

GTTGGCGAGA






2
Sauri-
−
GCTGTGCTGACCATCGACAAGGTTTTAGTACTCTGGAAACAGA
19076
CTTGGTATTTTGTTCAATCATGTTTTAGT
19227



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAcatggccccagcagcttcagtccctttcTCGT

GCAAAATGCCGTGTTTATCTCGTCAACTT






CGATGGtcag

GTTGGCGAGA






5
SpyCas9-
−
CTGTGCTGACCATCGACAAGGTTTTAGAGCTAGAAATAGCAAG
19077
CTTGGTATTTTGTTCAATCAGTTTTAGAG
19228



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcatggccccagcagcttcagtccctttcTCGTC

TCCGTTATCAACTTGAAAAAGTGGCACCG






GATGGtcag

AGTCGGTGC






8
SauCas9-
−
GGCTGTGCTGACCATCGACAAGTTTTAGTACTCTGGAAACAGA
19078
CTTGGTATTTTGTTCAATCATGTTTTAGT
19229



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAatggccccagcagcttcagtccctttcTCGTC

GCAAAATGCCGTGTTTATCTCGTCAACTT






GATGGTcagc

GTTGGCGAGA






11
ScaCas9-
+
CAGCTTCAGTCCCTTTCTTGGTTTTAGAGCTAGAAATAGCAAG
19079
GCGCTTCCTGGGAGGTGTCCGTTTTAGAG
19230



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcgtgcataaggctgtgctgaccatcgaCGAGAA

TCCGTTATCAACTTGAAAAAGTGGCACCG






AGGGActga

AGTCGGTGC






12
SpyCas9-
+
CAGCTTCAGTCCCTTTCTTGGTTTTAGAGCTAGAAATAGCAAG
19080
AGCGCTTCCTGGGAGGTGTCGTTTTAGAG
19231



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcgtgcataaggctgtgctgaccatcgaCGAGAA

TCCGTTATCAACTTGAAAAAGTGGCACCG






AGGGActga

AGTCGGTGC






13
SpyCas9-
−
GCTGTGCTGACCATCGACAAGTTTTAGAGCTAGAAATAGCAAG
19081
TTGGTATTTTGTTCAATCATGTTTTAGAG
19232



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCatggccccagcagcttcagtccctttcTCGTCG

TCCGTTATCAACTTGAAAAAGTGGCACCG






ATGGTcagc

AGTCGGTGC






19
SauCas9
−
AGGCTGTGCTGACCATCGACAGTTTTAGTACTCTGGAAACAGA
19082
TTGGTATTTTGTTCAATCATTGTTTTAGT
19233



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAtggccccagcagcttcagtccctttcTCGTCG

GCAAAATGCCGTGTTTATCTCGTCAACTT






ATGGTCagca

GTTGGCGAGA






20
SauCas9
+
AGCAGCTTCAGTCCCTTTCTTGTTTTAGTACTCTGGAAACAGA
19083
GCGCTTCCTGGGAGGTGTCCAGTTTTAGT
19234



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAgtgcataaggctgtgctgaccatcgaCGAGAA

GCAAAATGCCGTGTTTATCTCGTCAACTT






AGGGACtgaa

GTTGGCGAGA






21
SauCas9
+
AGCAGCTTCAGTCCCTTTCTTGTTTTAGTACTCTGGAAACAGA
19084
GCGCTTCCTGGGAGGTGTCCAGTTTTAGT
19235



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAgtgcataaggctgtgctgaccatcgaCGAGAA

GCAAAATGCCGTGTTTATCTCGTCAACTT






AGGGACtgaa

GTTGGCGAGA






24
SpyCas9-
−
GGCTGTGCTGACCATCGACAGTTTTAGAGCTAGAAATAGCAAG
19085
GGTATTTTGTTCAATCATTAGTTTTAGAG
19236



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtggccccagcagcttcagtccctttcTCGTCGA

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGGTCagca

AGTCGGTGC






28
SpyCas9-
−
GGCTGTGCTGACCATCGACAGTTTTAGAGCTAGAAATAGCAAG
19086
TGGTATTTTGTTCAATCATTGTTTTAGAG
19237



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtggccccagcagcttcagtccctttcTCGTCGA

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGGTCagca

AGTCGGTGC






29
SpyCas9-
+
GCAGCTTCAGTCCCTTTCTTGTTTTAGAGCTAGAAATAGCAAG
19087
GCGCTTCCTGGGAGGTGTCCGTTTTAGAG
19238



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgtgcataaggctgtgctgaccatcgaCGAGAAA

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGGACtgaa

AGTCGGTGC






34
SauCas9
−
atAAGGCTGTGCTGACCATCGACGTTTTAGTACTCTGGAAACA
19088
acTTGGTATTTTGTTCAATCATTGTTTTA
19239





GAATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAA

GTACTCTGGAAACAGAATCTACTAAAACA






CTTGTTGGCGAGAggccccagcagcttcagtccctttcTCGTC

AGGCAAAATGCCGTGTTTATCTCGTCAAC






GATGGTCAgcac

TTGTTGGCGAGA






35
SauCas9
−
AAGGCTGTGCTGACCATCGACGTTTTAGTACTCTGGAAACAGA
19089
TGGTATTTTGTTCAATCATTAGTTTTAGT
19240



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAggccccagcagcttcagtccctttcTCGTCGA

GCAAAATGCCGTGTTTATCTCGTCAACTT






TGGTCAgcac

GTTGGCGAGA






38
ScaCas9-
−
AGGCTGTGCTGACCATCGACGTTTTAGAGCTAGAAATAGCAAG
19090
TGGTATTTTGTTCAATCATTGTTTTAGAG
19241



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCggccccagcagcttcagtccctttcTCGTCGAT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGTCAgcac

AGTCGGTGC






39
SpyCas9-
−
AGGCTGTGCTGACCATCGACGTTTTAGAGCTAGAAATAGCAAG
19091
GGTATTTTGTTCAATCATTAGTTTTAGAG
19242



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCggccccagcagcttcagtccctttcTCGTCGAT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGTCAgcac

AGTCGGTGC






40
SpyCas9-
+
AGCAGCTTCAGTCCCTTTCTGTTTTAGAGCTAGAAATAGCAAG
19092
CGCTTCCTGGGAGGTGTCCAGTTTTAGAG
19243



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtgcataaggctgtgctgaccatcgaCGAGAAAG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGACTgaag

AGTCGGTGC






44
SpyCas9-
+
AGCAGCTTCAGTCCCTTTCTGTTTTAGAGCTAGAAATAGCAAG
19093
CGCTTCCTGGGAGGTGTCCAGTTTTAGAG
19244



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtgcataaggctgtgctgaccatcgaCGAGAAAG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGACTgaag

AGTCGGTGC






50
SauCas9
−
TAAGGCTGTGCTGACCATCGAGTTTTAGTACTCTGGAAACAGA
19094
GGTATTTTGTTCAATCATTAAGTTTTAGT
19245



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAgccccagcagcttcagtccctttcTCGTCGAT

GCAAAATGCCGTGTTTATCTCGTCAACTT






GGTCAGcaca

GTTGGCGAGA






51
Sauri-
−
TAAGGCTGTGCTGACCATCGAGTTTTAGTACTCTGGAAACAGA
19095
GGTATTTTGTTCAATCATTAAGTTTTAGT
19246



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAgccccagcagcttcagtccctttcTCGTCGAT

GCAAAATGCCGTGTTTATCTCGTCAACTT






GGTCAGcaca

GTTGGCGAGA






56
ScaCas9-
+
CAGCAGCTTCAGTCCCTTTCGTTTTAGAGCTAGAAATAGCAAG
19096
GCTTCCTGGGAGGTGTCCACGTTTTAGAG
19247



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgcataaggctgtgctgaccatcgaCGAGAAAGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GACTGaagc

AGTCGGTGC






57
SpyCas9-
+
CAGCAGCTTCAGTCCCTTTCGTTTTAGAGCTAGAAATAGCAAG
19097
GCTTCCTGGGAGGTGTCCACGTTTTAGAG
19248



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgcataaggctgtgctgaccatcgaCGAGAAAGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GACTGaagc

AGTCGGTGC






58
SpyCas9-
−
AAGGCTGTGCTGACCATCGAGTTTTAGAGCTAGAAATAGCAAG
19098
GTATTTTGTTCAATCATTAAGTTTTAGAG
19249



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgccccagcagcttcagtccctttcTCGTCGATG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GTCAGcaca

AGTCGGTGC






59
St1Cas9
−
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCTGGTACCAGAA
19099
NAGTCTTTGTACTCTGGTACCAGAAGCTA
19250





GCTACAAAGATAAGGCTTCATGCCGAAATCAACACCCTGTCAT

CAAAGATAAGGCTTCATGCCGAAATCAAC






TTTATGGCAGGGTGTTTTgccccagcagcttcagtccctttcT

ACCCTGTCATTTTATGGCAGGGTGTTTT






CGTCGATGGTCAGcaca








60
BlatCas9
+
ccccAGCAGCTTCAGTCCCTTTCGCTATAGTTCCTTACTGAAA
19100
gcgcTTCCTGGGAGGTGTCCACGGCTATA
19251





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTgc

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ataaggctgtgctgaccatcgaCGAGAAAGGGACTGaagc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






61
BlatCas9
+
ccccAGCAGCTTCAGTCCCTTTCGCTATAGTTCCTTACTGAAA
19101
gcgcTTCCTGGGAGGTGTCCACGGCTATA
19252





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTgc

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ataaggctgtgctgaccatcgaCGAGAAAGGGACTGaagc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






64
SauCas9
−
ATAAGGCTGTGCTGACCATCGGTTTTAGTACTCTGGAAACAGA
19102
GGTATTTTGTTCAATCATTAAGTTTTAGT
19253



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAccccagcagcttcagtccctttcTCGTCGATG

GCAAAATGCCGTGTTTATCTCGTCAACTT






GTCAGCacag

GTTGGCGAGA






65
SpyCas9-
+
CCAGCAGCTTCAGTCCCTTTGTTTTAGAGCTAGAAATAGCAAG
19103
CTTCCTGGGAGGTGTCCACGGTTTTAGAG
19254



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcataaggctgtgctgaccatcgaCGAGAAAGGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






ACTGAagct

AGTCGGTGC






66
SpyCas9-
−
TAAGGCTGTGCTGACCATCGGTTTTAGAGCTAGAAATAGCAAG
19104
TATTTTGTTCAATCATTAAGGTTTTAGAG
19255



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCccccagcagcttcagtccctttcTCGTCGATGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






TCAGCacag

AGTCGGTGC






69
SauCas9
−
CATAAGGCTGTGCTGACCATCGTTTTAGTACTCTGGAAACAGA
19105
ATTTTGTTCAATCATTAAGAAGTTTTAGT
19256



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAcccagcagcttcagtccctttcTCGTCGATGG

GCAAAATGCCGTGTTTATCTCGTCAACTT






TCAGCAcagc

GTTGGCGAGA






70
SpyCas9-
−
ATAAGGCTGTGCTGACCATCGTTTTAGAGCTAGAAATAGCAAG
19106
ATTTTGTTCAATCATTAAGAGTTTTAGAG
19257



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcccagcagcttcagtccctttcTCGTCGATGGT

TCCGTTATCAACTTGAAAAAGTGGCACCG






CAGCAcagc

AGTCGGTGC






71
SpyCas9-
+
CCCAGCAGCTTCAGTCCCTTGTTTTAGAGCTAGAAATAGCAAG
19107
TTCCTGGGAGGTGTCCACGTGTTTTAGAG
19258



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCataaggctgtgctgaccatcgaCGAGAAAGGGA

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTGAAgctg

AGTCGGTGC






75
SpyCas9-
−
CATAAGGCTGTGCTGACCATGTTTTAGAGCTAGAAATAGCAAG
19108
ATTTTGTTCAATCATTAAGAGTTTTAGAG
19259



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCccagcagcttcagtccctttcTCGTCGATGGTC

TCCGTTATCAACTTGAAAAAGTGGCACCG






AGCACagCC

AGTCGGTGC






79
SpyCas9-
−
CATAAGGCTGTGCTGACCATGTTTTAGAGCTAGAAATAGCAAG
19109
TTTTGTTCAATCATTAAGAAGTTTTAGAG
19260



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCccagcagcttcagtccctttcTCGTCGATGGTC

TCCGTTATCAACTTGAAAAAGTGGCACCG






AGCACagcc

AGTCGGTGC






80
SpyCas9-
+
CCCCAGCAGCTTCAGTCCCTGTTTTAGAGCTAGAAATAGCAAG
19110
TCCTGGGAGGTGTCCACGTGGTTTTAGAG
19261



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtaaggctgtgctgaccatcgaCGAGAAAGGGAC

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGAAGctgc

AGTCGGTGC






86
ScaCas9-
−
GCATAAGGCTGTGCTGACCAGTTTTAGAGCTAGAAATAGCAAG
19111
TATTTTGTTCAATCATTAAGGTTTTAGAG
19262



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcagcagcttcagtccctttcTCGTCGATGGTCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCACAgcct

AGTCGGTGC






87
SpyCas9-
−
GCATAAGGCTGTGCTGACCAGTTTTAGAGCTAGAAATAGCAAG
19112
TTTGTTCAATCATTAAGAAGGTTTTAGAG
19263



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcagcagcttcagtccctttcTCGTCGATGGTCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCACAgcct

AGTCGGTGC






88
SpyCas9-
+
GCCCCAGCAGCTTCAGTCCCGTTTTAGAGCTAGAAATAGCAAG
19113
CCTGGGAGGTGTCCACGTGAGTTTTAGAG
19264



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCaaggctgtgctgaccatcgaCGAGAAAGGGACT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GAAGCtgct

AGTCGGTGC






89
BlatCas9
−
cgtgCATAAGGCTGTGCTGACCAGCTATAGTTCCTTACTGAAA
19114
tggtATTTTGTTCAATCATTAAGGCTATA
19265





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTca

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gcagcttcagtccctttcTCGTCGATGGTCAGCACAgcct

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






90
BlatCas9
−
cgtgCATAAGGCTGTGCTGACCAGCTATAGTTCCTTACTGAAA
19115
tggtATTTTGTTCAATCATTAAGGCTATA
19266





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTca

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gcagcttcagtccctttcTCGTCGATGGTCAGCACAgcct

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






91
SauCas9
−
GTGCATAAGGCTGTGCTGACCGTTTTAGTACTCTGGAAACAGA
19116
TTTGTTCAATCATTAAGAAGAGTTTTAGT
19267



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAagcagcttcagtccctttcTCGTCGATGGTCA

GCAAAATGCCGTGTTTATCTCGTCAACTT






GCACAGcctt

GTTGGCGAGA






92
SpyCas9-
+
GGCCCCAGCAGCTTCAGTCCGTTTTAGAGCTAGAAATAGCAAG
19117
CTGGGAGGTGTCCACGTGAGGTTTTAGAG
19268



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCaggctgtgctgaccatcgaCGAGAAAGGGACTG

TCCGTTATCAACTTGAAAAAGTGGCACCG






AAGCTgctg

AGTCGGTGC






93
SpyCas9-
−
TGCATAAGGCTGTGCTGACCGTTTTAGAGCTAGAAATAGCAAG
19118
TTGTTCAATCATTAAGAAGAGTTTTAGAG
19269



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCagcagcttcagtccctttcTCGTCGATGGTCAG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CACAGcctt

AGTCGGTGC






94
BlatCas9
+
catgGCCCCAGCAGCTTCAGTCCGCTATAGTTCCTTACTGAAA
19119
tcctGGGAGGTGTCCACGTGAGCGCTATA
19270





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTag

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gctgtgctgaccatcgaCGAGAAAGGGACTGAAGCTgctg

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






95
BlatCas9
+
catgGCCCCAGCAGCTTCAGTCCGCTATAGTTCCTTACTGAAA
19120
tcctGGGAGGTGTCCACGTGAGCGCTATA
19271





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTag

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gctgtgctgaccatcgaCGAGAAAGGGACTGAAGCTgctg

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






98
SpyCas9-
−
GTGCATAAGGCTGTGCTGACGTTTTAGAGCTAGAAATAGCAAG
19121
TGTTCAATCATTAAGAAGACGTTTTAGAG
19272



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgcagcttcagtccctttcTCGTCGATGGTCAGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






ACAGCctta

AGTCGGTGC






99
SpyCas9-
+
TGGCCCCAGCAGCTTCAGTCGTTTTAGAGCTAGAAATAGCAAG
19122
TGGGAGGTGTCCACGTGAGCGTTTTAGAG
19273



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCggctgtgctgaccatcgaCGAGAAAGGGACTGA

TCCGTTATCAACTTGAAAAAGTGGCACCG






AGCTGctgg

AGTCGGTGC






102
SpyCas9-
+
ATGGCCCCAGCAGCTTCAGTGTTTTAGAGCTAGAAATAGCAAG
19123
GGGAGGTGTCCACGTGAGCCGTTTTAGAG
19274



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgctgtgctgaccatcgaCGAGAAAGGGACTGAA

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCTGCtggg

AGTCGGTGC






103
SpyCas9-
−
CGTGCATAAGGCTGTGCTGAGTTTTAGAGCTAGAAATAGCAAG
19124
GTTCAATCATTAAGAAGACAGTTTTAGAG
19275



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcagcttcagtccctttcTCGTCGATGGTCAGCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






CAGCCttat

AGTCGGTGC






104
BlatCas9
−
ggccGTGCATAAGGCTGTGCTGAGCTATAGTTCCTTACTGAAA
19125
tggtATTTTGTTCAATCATTAAGGCTATA
19276





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTca

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gcttcagtccctttcTCGTCGATGGTCAGCACAGCCttat

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






105
SpyCas9-
+
CATGGCCCCAGCAGCTTCAGGTTTTAGAGCTAGAAATAGCAAG
19126
GGAGGTGTCCACGTGAGCCTGTTTTAGAG
19277



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCctgtgctgaccatcgaCGAGAAAGGGACTGAAG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTGCTgggg

AGTCGGTGC






106
SpyCas9-
−
CCGTGCATAAGGCTGTGCTGGTTTTAGAGCTAGAAATAGCAAG
19127
TTCAATCATTAAGAAGACAAGTTTTAGAG
19278



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCagcttcagtccctttcTCGTCGATGGTCAGCAC

TCCGTTATCAACTTGAAAAAGTGGCACCG






AGCCTtatg

AGTCGGTGC






107
SpyCas9-
−
GCCGTGCATAAGGCTGTGCTGTTTTAGAGCTAGAAATAGCAAG
19128
TCAATCATTAAGAAGACAAAGTTTTAGAG
19279



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgcttcagtccctttcTCGTCGATGGTCAGCACA

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCCTTatgc

AGTCGGTGC






108
SpyCas9-
+
ACATGGCCCCAGCAGCTTCAGTTTTAGAGCTAGAAATAGCAAG
19129
GAGGTGTCCACGTGAGCCTTGTTTTAGAG
19280



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtgtgctgaccatcgaCGAGAAAGGGACTGAAGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGCTGgggc

AGTCGGTGC






109
BlatCas9
+
aaaaCATGGCCCCAGCAGCTTCAGCTATAGTTCCTTACTGAAA
19130
ctggGAGGTGTCCACGTGAGCCTGCTATA
19281





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtg

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






tgctgaccatcgaCGAGAAAGGGACTGAAGCTGCTGgggc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






110
BlatCas9
+
aaaaCATGGCCCCAGCAGCTTCAGCTATAGTTCCTTACTGAAA
19131
ctggGAGGTGTCCACGTGAGCCTGCTATA
19282





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtg

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






tgctgaccatcgaCGAGAAAGGGACTGAAGCTGCTGgggc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






112
Nme2Cas9
+
taAAAACATGGCCCCAGCAGCTTCGTTGTAGCTCCCTTTCTCA
19132
gaGGTGTCCACGTGAGCCTTGCTCGTTGT
19283





TTTCGGAAACGAAATGAGAACCGTTGCTACAATAAGGCCGTCT

AGCTCCCTTTCTCATTTCGGAAACGAAAT






GAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT

GAGAACCGTTGCTACAATAAGGCCGTCTG






TAAGGGGCATCGTTTAgtgctgaccatcgaCGAGAAAGGGACT

AAAAGATGTGCCGCAACGCTCTGCCCCTT






GAAGCTGCTGGggcc

AAAGCTTCTGCTTTAAGGGGCATCGTTTA






113
SpyCas9-
+
AACATGGCCCCAGCAGCTTCGTTTTAGAGCTAGAAATAGCAAG
19133
GGAGGTGTCCACGTGAGCCTGTTTTAGAG
19284



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgtgctgaccatcgaCGAGAAAGGGACTGAAGCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCTGGggcc

AGTCGGTGC






116
SpyCas9-
−
GGCCGTGCATAAGGCTGTGCGTTTTAGAGCTAGAAATAGCAAG
19134
CAATCATTAAGAAGACAAAGGTTTTAGAG
19285



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcttcagtccctttcTCGTCGATGGTCAGCACAG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CCTTAtgca

AGTCGGTGC






120
SpyCas9-
+
AACATGGCCCCAGCAGCTTCGTTTTAGAGCTAGAAATAGCAAG
19135
AGGTGTCCACGTGAGCCTTGGTTTTAGAG
19286



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgtgctgaccatcgaCGAGAAAGGGACTGAAGCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCTGGggcc

AGTCGGTGC






122
SpyCas9-
−
GGCCGTGCATAAGGCTGTGCGTTTTAGAGCTAGAAATAGCAAG
19136
CAATCATTAAGAAGACAAAGGTTTTAGAG
19287



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcttcagtccctttcTCGTCGATGGTCAGCACAG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CCTTAtgca

AGTCGGTGC






123
BlatCas9
+
aaaaACATGGCCCCAGCAGCTTCGCTATAGTTCCTTACTGAAA
19137
ctggGAGGTGTCCACGTGAGCCTGCTATA
19288





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTgt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gctgaccatcgaCGAGAAAGGGACTGAAGCTGCTGGggcc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






124
BlatCas9
+
aaaaACATGGCCCCAGCAGCTTCGCTATAGTTCCTTACTGAAA
19138
ctggGAGGTGTCCACGTGAGCCTGCTATA
19289





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTgt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






gctgaccatcgaCGAGAAAGGGACTGAAGCTGCTGGggcc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






125
BlatCas9
−
ccagGCCGTGCATAAGGCTGTGCGCTATAGTTCCTTACTGAAA
19139
cattAAGAAGACAAAGGGTTTGTGCTATA
19290





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTct

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






tcagtccctttcTCGTCGATGGTCAGCACAGCCTTAtgca

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






130
Nme2Cas9
+
ctAAAAACATGGCCCCAGCAGCTTGTTGTAGCTCCCTTTCTCA
19140
gaGGTGTCCACGTGAGCCTTGCTCGTTGT
19291





TTTCGGAAACGAAATGAGAACCGTTGCTACAATAAGGCCGTCT

AGCTCCCTTTCTCATTTCGGAAACGAAAT






GAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT

GAGAACCGTTGCTACAATAAGGCCGTCTG






TAAGGGGCATCGTTTAtgctgaccatcgaCGAGAAAGGGACTG

AAAAGATGTGCCGCAACGCTCTGCCCCTT






AAGCTGCTGGGgcca

AAAGCTTCTGCTTTAAGGGGCATCGTTTA 






131
Nme2Cas9
−
ctCCAGGCCGTGCATAAGGCTGTGGTTGTAGCTCCCTTTCTCA
19141
aaGAAGACAAAGGGTTTGTTGAACGTTGT
19292





TTTCGGAAACGAAATGAGAACCGTTGCTACAATAAGGCCGTCT

AGCTCCCTTTCTCATTTCGGAAACGAAAT






GAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT

GAGAACCGTTGCTACAATAAGGCCGTCTG






TAAGGGGCATCGTTTAttcagtccctttcTCGTCGATGGTCAG

AAAAGATGTGCCGCAACGCTCTGCCCCTT






CACAGCCTTATgcac

AAAGCTTCTGCTTTAAGGGGCATCGTTTA






134
ScaCas9-
+
AAACATGGCCCCAGCAGCTTGTTTTAGAGCTAGAAATAGCAAG
19142
GGTGTCCACGTGAGCCTTGCGTTTTAGAG
19293



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtgctgaccatcgaCGAGAAAGGGACTGAAGCTG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTGGGgcca

AGTCGGTGC






135
SpyCas9-
+
AAACATGGCCCCAGCAGCTTGTTTTAGAGCTAGAAATAGCAAG
19143
GGTGTCCACGTGAGCCTTGCGTTTTAGAG
19294



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtgctgaccatcgaCGAGAAAGGGACTGAAGCTG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTGGGgcca

AGTCGGTGC






138
ScaCas9-
−
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTAGAAATAGCAAG
19144
TCATTAAGAAGACAAAGGGTGTTTTAGAG
19295



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCttcagtccctttcTCGTCGATGGTCAGCACAGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTTATgcac

AGTCGGTGC






139
SpyCas9-
−
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTAGAAATAGCAAG
19145
AATCATTAAGAAGACAAAGGGTTTTAGAG
19296



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCttcagtccctttcTCGTCGATGGTCAGCACAGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTTATgcac

AGTCGGTGC






140
BlatCas9
−
tccaGGCCGTGCATAAGGCTGTGGCTATAGTTCCTTACTGAAA
19146
cattAAGAAGACAAAGGGTTTGTGCTATA
19297





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






cagtccctttcTCGTCGATGGTCAGCACAGCCTTATgcac

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






141
BlatCas9
+
taaaAACATGGCCCCAGCAGCTTGCTATAGTTCCTTACTGAAA
19147
aggtGTCCACGTGAGCCTTGCTCGCTATA
19298





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtg

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ctgaccatcgaCGAGAAAGGGACTGAAGCTGCTGGGgcca

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






142
BlatCas9
−
tccaGGCCGTGCATAAGGCTGTGGCTATAGTTCCTTACTGAAA
19148
cattAAGAAGACAAAGGGTTTGTGCTATA
19299





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






cagtccctttcTCGTCGATGGTCAGCACAGCCTTATgcac

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






145
SauCas9
−
CCAGGCCGTGCATAAGGCTGTGTTTTAGTACTCTGGAAACAGA
19149
CATTAAGAAGACAAAGGGTTTGTTTTAGT
19300



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAtcagtccctttcTCGTCGATGGTCAGCACAGC

GCAAAATGCCGTGTTTATCTCGTCAACTT






CTTATGcacg

GTTGGCGAGA






146
Sauri-
+
AAAAACATGGCCCCAGCAGCTGTTTTAGTACTCTGGAAACAGA
19150
GGTGTCCACGTGAGCCTTGCTGTTTTAGT
19301



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAgctgaccatcgaCGAGAAAGGGACTGAAGCTG

GCAAAATGCCGTGTTTATCTCGTCAACTT






CTGGGGccat

GTTGGCGAGA






147
SpyCas9-
+
AAAACATGGCCCCAGCAGCTGTTTTAGAGCTAGAAATAGCAAG
19151
GTGTCCACGTGAGCCTTGCTGTTTTAGAG
19302



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgctgaccatcgaCGAGAAAGGGACTGAAGCTGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGGGGccat

AGTCGGTGC






148
SpyCas9-
−
CAGGCCGTGCATAAGGCTGTGTTTTAGAGCTAGAAATAGCAAG
19152
ATCATTAAGAAGACAAAGGGGTTTTAGAG
19303



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtcagtccctttcTCGTCGATGGTCAGCACAGCC

TCCGTTATCAACTTGAAAAAGTGGCACCG






TTATGcacg

AGTCGGTGC






149
SauCas9
+
TAAAAACATGGCCCCAGCAGCGTTTTAGTACTCTGGAAACAGA
19153
GGTGTCCACGTGAGCCTTGCTGTTTTAGT
19304



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGActgaccatcgaCGAGAAAGGGACTGAAGCTGC

GCAAAATGCCGTGTTTATCTCGTCAACTT






TGGGGCcatg

GTTGGCGAGA






150
SauCas9
+
TAAAAACATGGCCCCAGCAGCGTTTTAGTACTCTGGAAACAGA
19154
GGTGTCCACGTGAGCCTTGCTGTTTTAGT
19305



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGActgaccatcgaCGAGAAAGGGACTGAAGCTGC

GCAAAATGCCGTGTTTATCTCGTCAACTT






TGGGGCcatg

GTTGGCGAGA






153
SpyCas9-
−
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTAGAAATAGCAAG
19155
CATTAAGAAGACAAAGGGTTGTTTTAGAG
19306



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcagtccctttcTCGTCGATGGTCAGCACAGCCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TATGCacgg

AGTCGGTGC






157
SpyCas9-
−
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTAGAAATAGCAAG
19156
TCATTAAGAAGACAAAGGGTGTTTTAGAG
19307



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcagtccctttcTCGTCGATGGTCAGCACAGCCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TATGCacgg

AGTCGGTGC






158
SpyCas9-
+
AAAAACATGGCCCCAGCAGCGTTTTAGAGCTAGAAATAGCAAG
19157
TGTCCACGTGAGCCTTGCTCGTTTTAGAG
19308



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCctgaccatcgaCGAGAAAGGGACTGAAGCTGCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGGGCcatg

AGTCGGTGC






162
ScaCas9-
−
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTAGAAATAGCAAG
19158
TCATTAAGAAGACAAAGGGTGTTTTAGAG
19309



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCagtccctttcTCGTCGATGGTCAGCACAGCCTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






ATGCAcggc

AGTCGGTGC






163
SpyCas9-
−
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTAGAAATAGCAAG
19159
CATTAAGAAGACAAAGGGTTGTTTTAGAG
19310



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCagtccctttcTCGTCGATGGTCAGCACAGCCTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






ATGCAcggc

AGTCGGTGC






164
SpyCas9-
+
TAAAAACATGGCCCCAGCAGGTTTTAGAGCTAGAAATAGCAAG
19160
GTCCACGTGAGCCTTGCTCGGTTTTAGAG
19311



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtgaccatcgaCGAGAAAGGGACTGAAGCTGCTG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGGCCatgt

AGTCGGTGC






165
SpyCas9-
−
CTCCAGGCCGTGCATAAGGCGTTTTAGAGCTAGAAATAGCAAG
19161
CATTAAGAAGACAAAGGGTTGTTTTAGAG
19312



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgtccctttcTCGTCGATGGTCAGCACAGCCTTA

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGCACggcc

AGTCGGTGC






169
SpyCas9-
−
CTCCAGGCCGTGCATAAGGCGTTTTAGAGCTAGAAATAGCAAG
19162
ATTAAGAAGACAAAGGGTTTGTTTTAGAG
19313



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgtccctttcTCGTCGATGGTCAGCACAGCCTTA

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGCACggcc

AGTCGGTGC






170
SpyCas9-
+
CTAAAAACATGGCCCCAGCAGTTTTAGAGCTAGAAATAGCAAG
19163
TCCACGTGAGCCTTGCTCGAGTTTTAGAG
19314



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgaccatcgaCGAGAAAGGGACTGAAGCTGCTGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGCCAtgtt

AGTCGGTGC






171
BlatCas9
+
cctcTAAAAACATGGCCCCAGCAGCTATAGTTCCTTACTGAAA
19164
ggtgTCCACGTGAGCCTTGCTCGGCTATA
19315





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTga

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ccatcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCAtgtt

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






172
BlatCas9
−
ccccTCCAGGCCGTGCATAAGGCGCTATAGTTCCTTACTGAAA
19165
cattAAGAAGACAAAGGGTTTGTGCTATA
19316





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTgt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ccctttcTCGTCGATGGTCAGCACAGCCTTATGCACggcc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






173
BlatCas9
+
cctcTAAAAACATGGCCCCAGCAGCTATAGTTCCTTACTGAAA
19166
ggtgTCCACGTGAGCCTTGCTCGGCTATA
19317





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTga

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ccatcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCAtgtt

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






174
BlatCas9
−
ccccTCCAGGCCGTGCATAAGGCGCTATAGTTCCTTACTGAAA
19167
cattAAGAAGACAAAGGGTTTGTGCTATA
19318





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTgt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






ccctttcTCGTCGATGGTCAGCACAGCCTTATGCACggcc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






177
ScaCas9-
−
CCTCCAGGCCGTGCATAAGGGTTTTAGAGCTAGAAATAGCAAG
19168
TTAAGAAGACAAAGGGTTTGGTTTTAGAG
19319



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtccctttcTCGTCGATGGTCAGCACAGCCTTAT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCACGgcct

AGTCGGTGC






178
SpyCas9-
−
CCTCCAGGCCGTGCATAAGGGTTTTAGAGCTAGAAATAGCAAG
19169
TTAAGAAGACAAAGGGTTTGGTTTTAGAG
19320



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtccctttcTCGTCGATGGTCAGCACAGCCTTAT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCACGgcct

AGTCGGTGC






179
SpyCas9-
+
TCTAAAAACATGGCCCCAGCGTTTTAGAGCTAGAAATAGCAAG
19170
TCCACGTGAGCCTTGCTCGAGTTTTAGAG
19321



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCaccatcgaCGAGAAAGGGACTGAAGCTGCTGGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCCATgttt

AGTCGGTGC






183
SpyCas9-
+
TCTAAAAACATGGCCCCAGCGTTTTAGAGCTAGAAATAGCAAG
19171
CCACGTGAGCCTTGCTCGAGGTTTTAGAG
19322



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCaccatcgaCGAGAAAGGGACTGAAGCTGCTGGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCCATgttt

AGTCGGTGC






187
ScaCas9-
+
CTCTAAAAACATGGCCCCAGGTTTTAGAGCTAGAAATAGCAAG
19172
ACGTGAGCCTTGCTCGAGGCGTTTTAGAG
19323



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCccatcgaCGAGAAAGGGACTGAAGCTGCTGGGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CCATGtttt

AGTCGGTGC






188
SpyCas9-
+
CTCTAAAAACATGGCCCCAGGTTTTAGAGCTAGAAATAGCAAG
19173
CACGTGAGCCTTGCTCGAGGGTTTTAGAG
19324



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCccatcgaCGAGAAAGGGACTGAAGCTGCTGGGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CCATGtttt

AGTCGGTGC






189
SpyCas9-
−
CCCTCCAGGCCGTGCATAAGGTTTTAGAGCTAGAAATAGCAAG
19174
TAAGAAGACAAAGGGTTTGTGTTTTAGAG
19325



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCccctttcTCGTCGATGGTCAGCACAGCCTTATG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CACGGcctg

AGTCGGTGC






192
Sauri-
+
GCCTCTAAAAACATGGCCCCAGTTTTAGTACTCTGGAAACAGA
19175
CACGTGAGCCTTGCTCGAGGCGTTTTAGT
19326



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAcatcgaCGAGAAAGGGACTGAAGCTGCTGGGG

GCAAAATGCCGTGTTTATCTCGTCAACTT






CCATGTtttt

GTTGGCGAGA






193
SpyCas9-
−
CCCCTCCAGGCCGTGCATAAGTTTTAGAGCTAGAAATAGCAAG
19176
TAAGAAGACAAAGGGTTTGTGTTTTAGAG
19327



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcctttcTCGTCGATGGTCAGCACAGCCTTATGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






ACGGCctgg

AGTCGGTGC






197
SpyCas9-
−
CCCCTCCAGGCCGTGCATAAGTTTTAGAGCTAGAAATAGCAAG
19177
AAGAAGACAAAGGGTTTGTTGTTTTAGAG
19328



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcctttcTCGTCGATGGTCAGCACAGCCTTATGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






ACGGCctgg

AGTCGGTGC






198
SpyCas9-
+
CCTCTAAAAACATGGCCCCAGTTTTAGAGCTAGAAATAGCAAG
19178
ACGTGAGCCTTGCTCGAGGCGTTTTAGAG
19329



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcatcgaCGAGAAAGGGACTGAAGCTGCTGGGGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






CATGTtttt

AGTCGGTGC






199
BlatCas9
+
tggcCTCTAAAAACATGGCCCCAGCTATAGTTCCTTACTGAAA
19179
acgtGAGCCTTGCTCGAGGCCTGGCTATA
19330





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTca

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






tcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCATGTtttt

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






203
SauCas9
+
GGCCTCTAAAAACATGGCCCCGTTTTAGTACTCTGGAAACAGA
19180
ACGTGAGCCTTGCTCGAGGCCGTTTTAGT
19331



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAatcgaCGAGAAAGGGACTGAAGCTGCTGGGGC

GCAAAATGCCGTGTTTATCTCGTCAACTT






CATGTTttta

GTTGGCGAGA






206
ScaCas9-
−
TCCCCTCCAGGCCGTGCATAGTTTTAGAGCTAGAAATAGCAAG
19181
TTAAGAAGACAAAGGGTTTGGTTTTAGAG
19332



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCctttcTCGTCGATGGTCAGCACAGCCTTATGCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






CGGCCtgga

AGTCGGTGC






207
SpyCas9
−
TCCCCTCCAGGCCGTGCATAGTTTTAGAGCTAGAAATAGCAAG
19182
GGGTTTGTTGAACTTGACCTGTTTTAGAG
19333





TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCctttcTCGTCGATGGTCAGCACAGCCTTATGCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






CGGCCtgga

AGTCGGTGC






210
SpyCas9-
−
TCCCCTCCAGGCCGTGCATAGTTTTAGAGCTAGAAATAGCAAG
19183
AGAAGACAAAGGGTTTGTTGGTTTTAGAG
19334



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCctttcTCGTCGATGGTCAGCACAGCCTTATGCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






CGGCCtgga

AGTCGGTGC






211
SpyCas9-
+
GCCTCTAAAAACATGGCCCCGTTTTAGAGCTAGAAATAGCAAG
19184
CGTGAGCCTTGCTCGAGGCCGTTTTAGAG
19335



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCatcgaCGAGAAAGGGACTGAAGCTGCTGGGGCC

TCCGTTATCAACTTGAAAAAGTGGCACCG






ATGTTttta

AGTCGGTGC






214
SpyCas9-
−
TCCCCTCCAGGCCGTGCATAGTTTTAGAGCTAGAAATAGCAAG
19185
TAAGAAGACAAAGGGTTTGTGTTTTAGAG
19336



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCctttcTCGTCGATGGTCAGCACAGCCTTATGCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






CGGCCtgga

AGTCGGTGC






218
SpyCas9-
+
GCCTCTAAAAACATGGCCCCGTTTTAGAGCTAGAAATAGCAAG
19186
CGTGAGCCTTGCTCGAGGCCGTTTTAGAG
19337



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCatcgaCGAGAAAGGGACTGAAGCTGCTGGGGCC

TCCGTTATCAACTTGAAAAAGTGGCACCG






ATGTTttta

AGTCGGTGC






224
Sauri-
−
TCTCCCCTCCAGGCCGTGCATGTTTTAGTACTCTGGAAACAGA
19187
AAGGGTTTGTTGAACTTGACCGTTTTAGT
19338



Cas9

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAtttcTCGTCGATGGTCAGCACAGCCTTATGCA

GCAAAATGCCGTGTTTATCTCGTCAACTT






CGGCCTggag

GTTGGCGAGA






225
Sauri-
−
TCTCCCCTCCAGGCCGTGCATGTTTTAGTACTCTGGAAACAGA
19188
AAGGGTTTGTTGAACTTGACCGTTTTAGT
19339



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAtttcTCGTCGATGGTCAGCACAGCCTTATGCA

GCAAAATGCCGTGTTTATCTCGTCAACTT






CGGCCTggag

GTTGGCGAGA






228
ScaCas9-
+
GGCCTCTAAAAACATGGCCCGTTTTAGAGCTAGAAATAGCAAG
19189
GTGAGCCTTGCTCGAGGCCTGTTTTAGAG
19340



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGTTTttag

AGTCGGTGC






229
SpyCas9-
+
GGCCTCTAAAAACATGGCCCGTTTTAGAGCTAGAAATAGCAAG
19190
GTGAGCCTTGCTCGAGGCCTGTTTTAGAG
19341



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCA

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGTTTttag

AGTCGGTGC






232
ScaCas9-
−
CTCCCCTCCAGGCCGTGCATGTTTTAGAGCTAGAAATAGCAAG
19191
AGACAAAGGGTTTGTTGAACGTTTTAGAG
19342



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtttcTCGTCGATGGTCAGCACAGCCTTATGCAC

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGCCTggag

AGTCGGTGC






233
SpyCas9-
−
CTCCCCTCCAGGCCGTGCATGTTTTAGAGCTAGAAATAGCAAG
19192
GAAGACAAAGGGTTTGTTGAGTTTTAGAG
19343



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtttcTCGTCGATGGTCAGCACAGCCTTATGCAC

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGCCTggag

AGTCGGTGC






234
BlatCas9
−
tctcTCCCCTCCAGGCCGTGCATGCTATAGTTCCTTACTGAAA
19193
agaaGACAAAGGGTTTGTTGAACGCTATA
19344





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






tcTCGTCGATGGTCAGCACAGCCTTATGCACGGCCTggag

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






235
BlatCas9
−
tctcTCCCCTCCAGGCCGTGCATGCTATAGTTCCTTACTGAAA
19194
agaaGACAAAGGGTTTGTTGAACGCTATA
19345





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTtt

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






tcTCGTCGATGGTCAGCACAGCCTTATGCACGGCCTggag

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






239
SauCas9
−
CTCTCCCCTCCAGGCCGTGCAGTTTTAGTACTCTGGAAACAGA
19195
GAAGACAAAGGGTTTGTTGAAGTTTTAGT
19346



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAttcTCGTCGATGGTCAGCACAGCCTTATGCAC

GCAAAATGCCGTGTTTATCTCGTCAACTT






GGCCTGgagg

GTTGGCGAGA






240
Sauri-
+
ATGGCCTCTAAAAACATGGCCGTTTTAGTACTCTGGAAACAGA
19196
ACGTGAGCCTTGCTCGAGGCCGTTTTAGT
19347



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCA

GCAAAATGCCGTGTTTATCTCGTCAACTT






TGTTTTtaga

GTTGGCGAGA






241
Sauri-
−
CTCTCCCCTCCAGGCCGTGCAGTTTTAGTACTCTGGAAACAGA
19197
AAGGGTTTGTTGAACTTGACCGTTTTAGT
19348



Cas9-

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG




KKH

TGTTGGCGAGAttcTCGTCGATGGTCAGCACAGCCTTATGCAC

GCAAAATGCCGTGTTTATCTCGTCAACTT






GGCCTGgagg

GTTGGCGAGA






244
SpyCas9-
−
TCTCCCCTCCAGGCCGTGCAGTTTTAGAGCTAGAAATAGCAAG
19198
AAGACAAAGGGTTTGTTGAAGTTTTAGAG
19349



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCttcTCGTCGATGGTCAGCACAGCCTTATGCACG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GCCTGgagg

AGTCGGTGC






245
SpyCas9-
+
TGGCCTCTAAAAACATGGCCGTTTTAGAGCTAGAAATAGCAAG
19199
TGAGCCTTGCTCGAGGCCTGGTTTTAGAG
19350



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcgaCGAGAAAGGGACTGAAGCTGCTGGGGCCAT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GTTTTtaga

AGTCGGTGC






246
BlatCas9
+
gtatGGCCTCTAAAAACATGGCCGCTATAGTTCCTTACTGAAA
19200
acgtGAGCCTTGCTCGAGGCCTGGCTATA
19351





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTcg

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






aCGAGAAAGGGACTGAAGCTGCTGGGGCCATGTTTTtaga

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






249
SauCas9
+
TATGGCCTCTAAAAACATGGCGTTTTAGTACTCTGGAAACAGA
19201
GAGCCTTGCTCGAGGCCTGGGGTTTTAGT
19352



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAgaCGAGAAAGGGACTGAAGCTGCTGGGGCCAT

GCAAAATGCCGTGTTTATCTCGTCAACTT






GTTTTTagag

GTTGGCGAGA






250
SauCas9
−
TCTCTCCCCTCCAGGCCGTGCGTTTTAGTACTCTGGAAACAGA
19202
GAAGACAAAGGGTTTGTTGAAGTTTTAGT
19353



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGAtcTCGTCGATGGTCAGCACAGCCTTATGCACG

GCAAAATGCCGTGTTTATCTCGTCAACTT






GCCTGGaggg

GTTGGCGAGA






251
SpyCas9-
+
ATGGCCTCTAAAAACATGGCGTTTTAGAGCTAGAAATAGCAAG
19203
GAGCCTTGCTCGAGGCCTGGGTTTTAGAG
19354



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCgaCGAGAAAGGGACTGAAGCTGCTGGGGCCATG

TCCGTTATCAACTTGAAAAAGTGGCACCG






TTTTTagag

AGTCGGTGC






252
SpyCas9-
−
CTCTCCCCTCCAGGCCGTGCGTTTTAGAGCTAGAAATAGCAAG
19204
AGACAAAGGGTTTGTTGAACGTTTTAGAG
19355



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCtcTCGTCGATGGTCAGCACAGCCTTATGCACGG

TCCGTTATCAACTTGAAAAAGTGGCACCG






CCTGGaggg

AGTCGGTGC






257
SauCas9
−
TTCTCTCCCCTCCAGGCCGTGGTTTTAGTACTCTGGAAACAGA
19205
GAAGACAAAGGGTTTGTTGAAGTTTTAGT
19356



KKH

ATCTACTAAAACAAGGCAAAATGCCGTGTTTATCTCGTCAACT

ACTCTGGAAACAGAATCTACTAAAACAAG






TGTTGGCGAGACTCGTCGATGGTCAGCACAGCCTTATGCACGG

GCAAAATGCCGTGTTTATCTCGTCAACTT






CCTGGAgggg

GTTGGCGAGA






258
SpyCas9-
−
TCTCTCCCCTCCAGGCCGTGGTTTTAGAGCTAGAAATAGCAAG
19206
GACAAAGGGTTTGTTGAACTGTTTTAGAG
19357



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCcTCGTCGATGGTCAGCACAGCCTTATGCACGGC

TCCGTTATCAACTTGAAAAAGTGGCACCG






CTGGAgggg

AGTCGGTGC






259
SpyCas9-
+
TATGGCCTCTAAAAACATGGGTTTTAGAGCTAGAAATAGCAAG
19207
AGCCTTGCTCGAGGCCTGGGGTTTTAGAG
19358



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCaCGAGAAAGGGACTGAAGCTGCTGGGGCCATGT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TTTTAgagg

AGTCGGTGC






261
SpyCas9-
+
GTATGGCCTCTAAAAACATGGTTTTAGAGCTAGAAATAGCAAG
19208
GCCTTGCTCGAGGCCTGGGAGTTTTAGAG
19359



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCCGAGAAAGGGACTGAAGCTGCTGGGGCCATGTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TTTAGaggc

AGTCGGTGC






262
SpyCas9-
−
TTCTCTCCCCTCCAGGCCGTGTTTTAGAGCTAGAAATAGCAAG
19209
ACAAAGGGTTTGTTGAACTTGTTTTAGAG
19360



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCTCGTCGATGGTCAGCACAGCCTTATGCACGGCC

TCCGTTATCAACTTGAAAAAGTGGCACCG






TGGAGggga

AGTCGGTGC






263
BlatCas9
+
tgggTATGGCCTCTAAAAACATGGCTATAGTTCCTTACTGAAA
19210
tgagCCTTGCTCGAGGCCTGGGAGCTATA
19361





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTCG

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






AGAAAGGGACTGAAGCTGCTGGGGCCATGTTTTTAGaggc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






264
Nme2Cas9
+
caTGGGTATGGCCTCTAAAAACATGTTGTAGCTCCCTTTCTCA
19211
gtGAGCCTTGCTCGAGGCCTGGGAGTTGT
19362





TTTCGGAAACGAAATGAGAACCGTTGCTACAATAAGGCCGTCT

AGCTCCCTTTCTCATTTCGGAAACGAAAT






GAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT

GAGAACCGTTGCTACAATAAGGCCGTCTG






TAAGGGGCATCGTTTAGAGAAAGGGACTGAAGCTGCTGGGGCC

AAAAGATGTGCCGCAACGCTCTGCCCCTT






ATGTTTTTAGAggcc

AAAGCTTCTGCTTTAAGGGGCATCGTTTA






265
SpyCas9-
+
GGTATGGCCTCTAAAAACATGTTTTAGAGCTAGAAATAGCAAG
19212
CTTGCTCGAGGCCTGGGATCGTTTTAGAG
19363



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCGAGAAAGGGACTGAAGCTGCTGGGGCCATGTTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TTAGAggcc

AGTCGGTGC






268
SpyCas9-
−
CTTCTCTCCCCTCCAGGCCGGTTTTAGAGCTAGAAATAGCAAG
19213
GACAAAGGGTTTGTTGAACTGTTTTAGAG
19364



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCCGTCGATGGTCAGCACAGCCTTATGCACGGCCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGAGGggag

AGTCGGTGC






272
SpyCas9-
+
GGTATGGCCTCTAAAAACATGTTTTAGAGCTAGAAATAGCAAG
19214
CCTTGCTCGAGGCCTGGGATGTTTTAGAG
19365



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCGAGAAAGGGACTGAAGCTGCTGGGGCCATGTTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TTAGAggcc

AGTCGGTGC






274
SpyCas9-
−
CTTCTCTCCCCTCCAGGCCGGTTTTAGAGCTAGAAATAGCAAG
19215
CAAAGGGTTTGTTGAACTTGGTTTTAGAG
19366



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCCGTCGATGGTCAGCACAGCCTTATGCACGGCCT

TCCGTTATCAACTTGAAAAAGTGGCACCG






GGAGGggag

AGTCGGTGC






275
BlatCas9
+
atggGTATGGCCTCTAAAAACATGCTATAGTTCCTTACTGAAA
19216
gagcCTTGCTCGAGGCCTGGGATGCTATA
19367





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTGA

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






GAAAGGGACTGAAGCTGCTGGGGCCATGTTTTTAGAggcc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






276
BlatCas9
+
atggGTATGGCCTCTAAAAACATGCTATAGTTCCTTACTGAAA
19217
gagcCTTGCTCGAGGCCTGGGATGCTATA
19368





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTGA

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






GAAAGGGACTGAAGCTGCTGGGGCCATGTTTTTAGAggcc

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT






279
Nme2Cas9
+
acATGGGTATGGCCTCTAAAAACAGTTGTAGCTCCCTTTCTCA
19218
gtGAGCCTTGCTCGAGGCCTGGGAGTTGT
19369





TTTCGGAAACGAAATGAGAACCGTTGCTACAATAAGGCCGTCT

AGCTCCCTTTCTCATTTCGGAAACGAAAT






GAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT

GAGAACCGTTGCTACAATAAGGCCGTCTG






TAAGGGGCATCGTTTAAGAAAGGGACTGAAGCTGCTGGGGCCA

AAAAGATGTGCCGCAACGCTCTGCCCCTT






TGTTTTTAGAGgcca

AAAGCTTCTGCTTTAAGGGGCATCGTTTA






282
ScaCas9-
+
GGGTATGGCCTCTAAAAACAGTTTTAGAGCTAGAAATAGCAAG
19219
CCTTGCTCGAGGCCTGGGATGTTTTAGAG
19370



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCAGAAAGGGACTGAAGCTGCTGGGGCCATGTTTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TAGAGgcca

AGTCGGTGC






283
SpyCas9
+
GGGTATGGCCTCTAAAAACAGTTTTAGAGCTAGAAATAGCAAG
19220
GTGAGCCTTGCTCGAGGCCTGTTTTAGAG
19371





TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCAGAAAGGGACTGAAGCTGCTGGGGCCATGTTTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TAGAGgcca

AGTCGGTGC






286
SpyCas9-
+
GGGTATGGCCTCTAAAAACAGTTTTAGAGCTAGAAATAGCAAG
19221
CTTGCTCGAGGCCTGGGATCGTTTTAGAG
19372



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCAGAAAGGGACTGAAGCTGCTGGGGCCATGTTTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TAGAGgcca

AGTCGGTGC






289
ScaCas9-
−
GCTTCTCTCCCCTCCAGGCCGTTTTAGAGCTAGAAATAGCAAG
19222
AGGGTTTGTTGAACTTGACCGTTTTAGAG
19373



Sc++

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCGTCGATGGTCAGCACAGCCTTATGCACGGCCTG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GAGGGgaga

AGTCGGTGC






290
SpyCas9-
−
GCTTCTCTCCCCTCCAGGCCGTTTTAGAGCTAGAAATAGCAAG
19223
AAAGGGTTTGTTGAACTTGAGTTTTAGAG
19374



SpRY

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCGTCGATGGTCAGCACAGCCTTATGCACGGCCTG

TCCGTTATCAACTTGAAAAAGTGGCACCG






GAGGGgaga

AGTCGGTGC






291
SpyCas9-
+
GGGTATGGCCTCTAAAAACAGTTTTAGAGCTAGAAATAGCAAG
19224
CTTGCTCGAGGCCTGGGATCGTTTTAGAG
19375



NG

TTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACC

CTAGAAATAGCAAGTTAAAATAAGGCTAG






GAGTCGGTGCAGAAAGGGACTGAAGCTGCTGGGGCCATGTTTT

TCCGTTATCAACTTGAAAAAGTGGCACCG






TAGAGgcca

AGTCGGTGC






294
BlatCas9
+
catgGGTATGGCCTCTAAAAACAGCTATAGTTCCTTACTGAAA
19225
gagcCTTGCTCGAGGCCTGGGATGCTATA
19376





GGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGGCGTTGGGG

GTTCCTTACTGAAAGGTAAGTTGCTATAG






ATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATA

TAAGGGCAACAGACCCGAGGCGTTGGGGA






ATGACAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCTAG

TCGCCTAGCCCGTGTTTACGGGCTCTCCC






AAAGGGACTGAAGCTGCTGGGGCCATGTTTTTAGAGgcca

CATATTCAAAATAATGACAGACGAGCACC








TTGGAGCATTTATCTCCGAGGTGCT









Capital letters indicate “core nucleotides” while lower case letters indicate “flanking nucleotides.” Herein, when an RNA sequence (e.g., a template RNA sequence) is said to comprise a particular sequence (e.g., a sequence of Table 4 or a portion thereof) that comprises thymine (T), it is of course understood that the RNA sequence may (and frequently does) comprise uracil (U) in place of T. For instance, the RNA sequence may comprise U at every position shown as T in the sequence in Table 4. More specifically, the present disclosure provides an RNA sequence according to every template sequence shown in Table 4, wherein the RNA sequence has a U in place of each T in the sequence of Table 4.









TABLE 5







Exemplary template RNA sequences comprising PAM-inactivating sites


Table 5 provides select sequences from Table 4, with annotation illustrating


inactivation of PAM sites. Column “ID” contains a unique identifier for the


template RNA that corresponds to the ID used in Tables 1-4 and can be used,


e.g., to identify the corresponding gRNA spacer sequence in Table 1. Column


“Cas species” indicates a type of Cas domain suitable for inclusion in a


gene modifying polypeptide for use with the template RNA. Column “consensus”


indicates a consensus PAM motif recognized by the Cas. Column “PAM sequence”


indicates a particular PAM sequence recognized by the Cas, e.g., in the


SERPINA1 gene. Column “PAM mutation” indicates a mutation that can be produced


in the PAM by a template RNA described on the same row of the table; mutated


nucleotides are indicated with bold and underlining. Column “strand” indicates


the + or 1 strand of the target nucleic acid. Column “distance” indicates the


number of nucleotides in the pre-edit homology region. Column “PBS sequence”


indicates a PBS sequence for partial or full inclusion in the template RNA,


wherein core nucleotides are capitalized and flanking nucleotides are lower


case. Column “RT template sequence” indicates a heterologous object sequence


for partial or full inclusion in the template RNA, wherein core nucleotides


are capitalized, flanking nucleotides are lower case, and nucleotide


differences from the target nucleic acid are shown in bold and underline.


























RT Template




Cas
con-
PAM
PAM


PBS
SEQ
Sequence
SEQ


ID
species
sensus
sequence
mutation
strand
distance
sequence
ID NO
(Mutation)
ID NO




















5
SpyCas9-
NRN
AAA


0
GTCGATGGt
19432
catgggtatggcctcta
19463



SpRY





cagcacag

aaaacatggccccagca












gcttcagtccctttcTC






11
ScaCas9-
NNG
TCG
TCA
+
1
GAAAGGGAc
19433
tctgcttctctcccctc
19464



Sc++





tgaagctg

caggccgtgcataaggc












tgtgctgaccattgaCG












A






12
SpyCas9-
NYN
TCG

+
1
GAAAGGGAc
19434
tctgcttctctcccctc
19465



SpRY





tgaagctg

caggccgtgcataaggc












tgtgctgaccatcgaCG












A






13
SpyCas9-
NRN
GAA


1
TCGATGGTc
19435
catgggtatggcctcta
19466



SpRY





agcacagc

aaaacatggccccagca












gcttcagtccctttcTC












G






20
SauCas9KKH
NNNRR
GTCGA
GTCTA
+
2
AAAGGGACt
19436
tctgcttctctcccctc
19467









gaagctgc

caggccgtgcataaggc












tgtgctgaccatagaCG












AG






21
SauCas9KKH
NNNRR
GTCGAT
GTCTAT
+
2
AAAGGGACt
19437
tctgcttctctcccctc
19468




T




gaagctgc

caggccgtgcataaggc












tgtgctgaccatagaCG












AG






24
SpyCas9-NG
NG
AG
AA
−
2
CGATGGTCa
19438
catgggtatggcctcta
19469









gcacagcc

aaaacatggccccagca












gcttcagtcccttttTC












GT






28
SpyCas9-
NRN
AGA

−
2
CGATGGTCa
19439
catgggtatggcctcta
19470



SpRY





gcacagcc

aaaacatggccccagca












gcttcagtccctttcTC












GT






29
SpyCas9-
NYN
GTC

+
2
AAAGGGACt
19440
tctgcttctctcccctc
19471



SpRY





gaagctgc

caggccgtgcataaggc












tgtgctgaccatcgaCG












AG






34
SauCas9
NNGRR
aAGAA
gAAAA
−
3
GATGGTCAg
19441
catgggtatggcctcta
19472









cacagcct

aaaacatggccccagca












gcttcagtcccttttTC












GTC






38
ScaCas9-
NNG
aAG
gAA
−
3
GATGGTCAg
19442
catgggtatggcctcta
19473



Sc++





cacagcct

aaaacatggccccagca












gcttcagtcccttttTC












GTC






39
SpyCas9-
NRN
aAG

−
3
GATGGTCAg
19443
catgggtatggcctcta
19474



SpRY





cacagcct

aaaacatggccccagca












gcttcagtccctttcTC












GTC






40
SpyCas9-NG
NG
tG
cA
+
3
AAGGGACTg
19444
tctgcttctctcccctc
19475









aagctgct

caggccgtgcataaggc












tgtgctgaccatcgaTG












AGA






44
SpyCas9-
NRN
tGT

+
3
AAGGGACTg
19445
tctgcttctctcccctc
19476



SpRY





aagctgct

caggccgtgcataaggc












tgtgctgaccatcgaCG












AGA






51
SauriCas9-
NNRG
CaAG
CgAA
−
4
ATGGTCAGC
19446
catgggtatggcctcta
19477



KKH





acagcctt

aaaacatggccccagca












gcttcagtcccttttTC












GTCG






56
ScaCas9-
NNG
TtG
TcA
+
4
AGGGACTGa
19447
tctgcttctctcccctc
19478



Sc++





agctgctg

caggccgtgcataaggc












tgtgctgaccatcgaTG












AGAA






59
St1Cas9
NNAGA
CaAGAA
CgAAAAG
−
4
ATGGTCAGc
19448
catgggtatggcctcta
19479




AW
A



acagcctt

aaaacatggccccagca












gcttcagtcccctttTC












GTCG






89
BlatCas9
NNNNC
TCGACa
TCGATgA
−
8
TCAGCACAg
19449
catgggtatggcctcta
19480




NDD
AG
G


ccttatgc

aaaacatggccccagca












gcttcagtccctttcTC














A
TCGATGG






90
BlatCas9
NNNNC
TCGAC
TCGAT
−
8
TCAGCACAg
19450
catgggtatggcctcta
19481









ccttatgc

aaaacatggccccagca












gcttcagtccctttcTC














A
TCGATGG






94
BlatCas9
NNNNC
CTTTCTt
CTTTTTcG
+
9
CTGAAGCTg
19451
tctgcttctctcccctc
19482




NDD
G



ctggggcc

caggccgtgcataaggc












tgtgctgaccatcgaCG












AAAAAGGGA






95
BlatCas9
NNNNC
CTTTC
CTTTT
+
9
CTGAAGCTg
19452
tctgcttctctcccctc
19483









ctggggcc

caggccgtgcataaggc












tgtgctgaccatcgaCG












AAAAAGGGA






104
BlatCas9
NNNNC
CCATC
CCATT
−
11
GCACAGCCt
19453
catgggtatggcctcta
19484









tatgcacg

aaaacatggccccagca












gcttcagtccctttcTC












GTCAATGGTCA






116
SpyCas9-NG
NG
TG
TC
−
14
CAGCCTTAt
19454
catgggtatggcctcta
19485









gcacggcc

aaaacatggccccagca












gcttcagtccctttcTC












GTCGATGGTGAGCA






125
BlatCas9
NNNNC
TGACC
TGACA
−
14
CAGCCTTAt
19455
catgggtatggcctcta
19486









gcacggcc

aaaacatggccccagca












gcttcagtccctttcTC












GTCGATTGTCAGCA






130
Nme2Cas9
NNNNC
CAGTCC
CAGTGC
+
15
CTGCTGGGg
19456
tctgcttctctcccctc
19487




C




ccatgttt

caggccgtgcataaggc












tgtgctgaccatcgaCG












AGAAAGGCACTGAAG






131
Nme2Cas9
NNNNC
CTGACC
CTGACA

15
AGCCTTATg
19457
catgggtatggcctcta
19488




C




cacggcct

aaaacatggccccagca












gcttcagtccctttcTC












GTCGATTGTCAGC












AC






138
ScaCas9-
NNG
CTG
CTC
−
15
AGCCTTATg
19458
catgggtatggcctcta
19489



Sc++





cacggcct

aaaacatggccccagca












gcttcagtccctttcTC












GTCGATGGTGAGCAC






141
BlatCas9
NNNNC
CAGTC
CAGTG
+
15
CTGCTGGGg
19459
tctgcttctctcccctc
19490









ccatgttt

caggccgtgcataaggc












tgtgctgaccatcgaCG












AGAAAGGCACTGAAG






207
SpyCas9
NGG
AGG
AAG
−
23
GCACGGCCt
19460
catgggtatggcctcta
19491









ggagggga

aaaacatggccccagca












gcttcagtccctttcTC












GTCGATGGTCAGCACAG












CTTTAT






283
SpyCas9
NGG
TGG
TAG
+
30
TTTTAGAGg
19461
tctgcttctctcccctc
19492









ccataccc

caggccgtgcataaggc












tgtgctgaccatcgaCG












AGAAAGGGACTGAAGCT












GCTGGGGCTATGT






333
SpyCas9
NGG
AGG
AAG
−
35
GGGGAGAGa
19462
catgggtatggcctcta
19493









agcagaga

aaaacatggccccagca












gcttcagtccctttcTC












GTCGATGGTCAGCACAG












CCTTATGCACGGCTTGG












A









Herein, when an RNA sequence (e.g., a template RNA sequence) is said to comprise a particular sequence (e.g., a sequence of Table 5 or a portion thereof) that comprises thymine (T), it is of course understood that the RNA sequence may (and frequently does) comprise uracil (U) in place of T. For instance, the RNA sequence may comprise U at every position shown as T in the sequence in Table 5. More specifically, the present disclosure provides an RNA sequence according to every template sequence shown in Table 5, wherein the RNA sequence has a U in place of each T in the sequence of Table 5.


In some embodiments, a gRNA scaffold described herein comprises a nucleic acid sequence comprising, in the 5′ to 3′ direction, a crRNA of Table 6A, a tetraloop from the same row of Table 6A, and a tracrRNA from the same row of Table 6A, or a sequence having at least 70%, 80%, 85%, 90%, 95%, or 99% identity thereto. In some embodiments, the gRNA or template RNA having a sequence according to Table 6A is comprised by a system that further comprises a gene modifying polypeptide, and a spacer, wherein the spacer comprises a gRNA spacer described in the same row of Table 6A.









TABLE 6A







Exemplary spacer and scaffold pairs.

















gRNA
SEQ ID

SEQ ID
Tetra

SEQ ID

SEQ ID


Name
Spacer
NO
crRNA
NO
loop
tracrRNA
NO
Full Scaffold
NO





pU6-
CTGTGC
19951
GTTTT
20070
GAA
TAGCAAGTTAA
20308
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20427


Spy-
TGACCA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
TCGACA

TA


TCCGTTATCAA

CGGTGC



sgRNA-
AG




CTTGAAAAAGT





1





GGCACCGAGTC











GGTGC








pU6-
GCTGTG
19952
GTTTT
20071
GAA
TAGCAAGTTAA
20309
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20428


Spy-
CTGACC

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
ATCGAC

TA


TCCGTTATCAA

CGGTGC



sgRNA-
AAG




CTTGAAAAAGT





1G





GGCACCGAGTC











GGTGC








pU6-
CAGCTT
19953
GTTTT
20072
GAA
TAGCAAGTTAA
20310
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20429


Spy-
CAGTCC

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CTTTCT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TG




CTTGAAAAAGT





2





GGCACCGAGTC











GGTGC








pU6-
GCAGCT
19954
GTTTT
20073
GAA
TAGCAAGTTAA
20311
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20430


Spy-
TCAGTC

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CCTTTC

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TTG




CTTGAAAAAGT





2G





GGCACCGAGTC











GGTGC








pU6-
GGCTGT
19955
GTTTT
20074
GAA
TAGCAAGTTAA
20312
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20431


Spy-
GCTGAC

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CATCGA

TA


TCCGTTATCAA

CGGTGC



sgRNA-
CA




CTTGAAAAAGT





3





GGCACCGAGTC











GGTGC








pU6-
AGGCTG
19956
GTTTT
20075
GAA
TAGCAAGTTAA
20313
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20432


Spy-
TGCTGA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CCATCG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
AC




CTTGAAAAAGT





4





GGCACCGAGTC











GGTGC








pU6-
GAGGCT
19957
GTTTT
20076
GAA
TAGCAAGTTAA
20314
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20433


Spy-
GTGCTG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
ACCATC

TA


TCCGTTATCAA

CGGTGC



sgRNA-
GAC




CTTGAAAAAGT





4G





GGCACCGAGTC











GGTGC








pU6-
AGCAGC
19958
GTTTT
20077
GAA
TAGCAAGTTAA
20315
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20434


Spy-
TTCAGT

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CCCTTT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
CT




CTTGAAAAAGT





5





GGCACCGAGTC











GGTGC








pU6-
GAGCAG
19959
GTTTT
20078
GAA
TAGCAAGTTAA
20316
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20435


Spy-
CTTCAG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
TCCCTT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TCT




CTTGAAAAAGT





5G





GGCACCGAGTC











GGTGC








pU6-
GGCCGT
19960
GTTTT
20079
GAA
TAGCAAGTTAA
20317
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20436


Spy-
GCATAA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
GGCTGT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
GC




CTTGAAAAAGT





6





GGCACCGAGTC











GGTGC








pU6-
CCAGGC
19961
GTTTT
20080
GAA
TAGCAAGTTAA
20318
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20437


Spy-
CGTGCA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
TAAGGC

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TG




CTTGAAAAAGT





7





GGCACCGAGTC











GGTGC








pU6-
GCCAGG
19962
GTTTT
20081
GAA
TAGCAAGTTAA
20319
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20438


Spy-
CCGTGC

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
ATAAGG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
CTG




CTTGAAAAAGT





7G





GGCACCGAGTC











GGTGC








pU6-
CAGCAG
19963
GTTTT
20082
GAA
TAGCAAGTTAA
20320
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20439


Spy-
CTTCAG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
TCCCTT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TC




CTTGAAAAAGT





8





GGCACCGAGTC











GGTGC








pU6-
GCAGCA
19964
GTTTT
20083
GAA
TAGCAAGTTAA
20321
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20440


Spy-
GCTTCA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
GTCCCT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TTC




CTTGAAAAAGT





8G





GGCACCGAGTC











GGTGC








pU6-
AGGCCG
19965
GTTTT
20084
GAA
TAGCAAGTTAA
20322
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20441


Spy-
TGCATA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
AGGCTG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
TG




CTTGAAAAAGT





9





GGCACCGAGTC











GGTGC








pU6-
GAGGCC
19966
GTTTT
20085
GAA
TAGCAAGTTAA
20323
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20442


Spy-
GTGCAT

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
AAGGCT

TA


TCCGTTATCAA

CGGTGC



sgRNA-
GTG




CTTGAAAAAGT





9G





GGCACCGAGTC











GGTGC








pU6-
TCCAGG
19967
GTTTT
20086
GAA
TAGCAAGTTAA
20324
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20443


Spy-
CCGTGC

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
ATAAGG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
CT




CTTGAAAAAGT





10





GGCACCGAGTC











GGTGC








pU6-
GTCCAG
19968
GTTTT
20087
GAA
TAGCAAGTTAA
20325
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20444


Spy-
GCCGTG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CATAAG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
GCT




CTTGAAAAAGT





10G





GGCACCGAGTC











GGTGC








pU6-
ACCTCG
19969
GTTTT
20088
GAA
TAGCAAGTTAA
20326
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20445


Spy-
GGGGGG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
ATAGAC

TA


TCCGTTATCAA

CGGTGC



sgRNA-
AT




CTTGAAAAAGT





11





GGCACCGAGTC











GGTGC








pU6-
GACCTC
19970
GTTTT
20089
GAA
TAGCAAGTTAA
20327
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20446


Spy-
GGGGGG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
GATAGA

TA


TCCGTTATCAA

CGGTGC



sgRNA-
CAT




CTTGAAAAAGT





11G





GGCACCGAGTC











GGTGC








pU6-
TGTTGA
19971
GTTTT
20090
GAA
TAGCAAGTTAA
20328
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20447


Spy-
ACTTGA

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
CCTCGG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
GG




CTTGAAAAAGT





12





GGCACCGAGTC











GGTGC








pU6-
GTGTTG
19972
GTTTT
20091
GAA
TAGCAAGTTAA
20329
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
20448


Spy-
AACTTG

AGAGC

A
AATAAGGCTAG

TAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGT



A1AT-
ACCTCG

TA


TCCGTTATCAA

CGGTGC



sgRNA-
GGG




CTTGAAAAAGT





12G





GGCACCGAGTC











GGTGC








pU6-
AAGGCT
19973
GTTTT
20092
GAA
CAGAATCTACT
20330
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20449


Sau-
GTGCTG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
ACCATC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GAC




TTATCTCGTCA





1





ACTTGTTGGCG











AGA








pU6-
GAAGGC
19974
GTTTT
20093
GAA
CAGAATCTACT
20331
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20450


Sau-
TGTGCT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GACCAT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CGAC




TTATCTCGTCA





1G





ACTTGTTGGCG











AGA








pU6-
AGCAGC
19975
GTTTT
20094
GAA
CAGAATCTACT
20332
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20451


Sau-
TTCAGT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CCCTTT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CTT




TTATCTCGTCA





2





ACTTGTTGGCG











AGA








pU6-
GAGCAG
19976
GTTTT
20095
GAA
CAGAATCTACT
20333
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20452


Sau-
CTTCAG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TCCCTT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TCTTCC




TTATCTCGTCA





2G
AGGC




ACTTGTTGGCG











AGA








pU6-
CGTGCA
19977
GTTTT
20096
GAA
CAGAATCTACT
20334
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20453


Sau-
TAAGGC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TGT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-





TTATCTCGTCA





3





ACTTGTTGGCG











AGA








pU6-
GCCAGG
19978
GTTTT
20097
GAA
CAGAATCTACT
20335
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20454


Sau-
CCGTGC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
ATAAGG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CTGT




TTATCTCGTCA





3G





ACTTGTTGGCG











AGA








pU6-
TAAAAA
19979
GTTTT
20098
GAA
CAGAATCTACT
20336
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20455


Sau-
CATGGC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CCCAGC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
AGC




TTATCTCGTCA





4





ACTTGTTGGCG











AGA








pU6-
GTAAAA
19980
GTTTT
20099
GAA
CAGAATCTACT
20337
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20456


Sau-
ACATGG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CCCCAG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CAGC




TTATCTCGTCA





4G





ACTTGTTGGCG











AGA








pU6-
GGCCTC
19981
GTTTT
20100
GAA
CAGAATCTACT
20338
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20457


Sau-
TAAAAA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CATGGC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CCC




TTATCTCGTCA





5





ACTTGTTGGCG











AGA








pU6-
TATGGC
19982
GTTTT
20101
GAA
CAGAATCTACT
20339
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20458


Sau-
CTCTAA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AAACAT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GGC




TTATCTCGTCA





6





ACTTGTTGGCG











AGA








pU6-
GTATGG
19983
GTTTT
20102
GAA
CAGAATCTACT
20340
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20459


Sau-
CCTCTA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AAAACA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TGGC




TTATCTCGTCA





6G





ACTTGTTGGCG











AGA








pU6-
TTGACC
19984
GTTTT
20103
GAA
CAGAATCTACT
20341
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20460


Sau-
TCGGGG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGGATA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GAC




TTATCTCGTCA





7





ACTTGTTGGCG











AGA








pU6-
GTTGAC
19985
GTTTT
20104
GAA
CAGAATCTACT
20342
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20461


Sau-
CTCGGG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGGGAT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
AGAC




TTATCTCGTCA





7G





ACTTGTTGGCG











AGA








pU6-
TTTGTT
19986
GTTTT
20105
GAA
CAGAATCTACT
20343
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20462


Sau-
GAACTT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GACCTC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GGG




TTATCTCGTCA





8





ACTTGTTGGCG











AGA








pU6-
GTTTGT
19987
GTTTT
20106
GAA
CAGAATCTACT
20344
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20463


Sau-
TGAACT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TGACCT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CGGG




TTATCTCGTCA





8G





ACTTGTTGGCG











AGA








pU6-
ACGTGA
19988
GTTTT
20107
GAA
CAGAATCTACT
20345
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20464


Sau-
GCCTTG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CTCGAG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GCC




TTATCTCGTCA





9





ACTTGTTGGCG











AGA








pU6-
GACGTG
19989
GTTTT
20108
GAA
CAGAATCTACT
20346
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20465


Sau-
AGCCTT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GCTCGA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GGCC




TTATCTCGTCA





9G





ACTTGTTGGCG











AGA








pU6-
ATTAAG
19990
GTTTT
20109
GAA
CAGAATCTACT
20347
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20466


Sau-
AAGACA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AAGGGT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TTG




TTATCTCGTCA





10





ACTTGTTGGCG











AGA








pU6-
GATTAA
19991
GTTTT
20110
GAA
CAGAATCTACT
20348
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20467


Sau-
GAAGAC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AAAGGG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TTTG




TTATCTCGTCA





10G





ACTTGTTGGCG











AGA








pU6-
AGGTGT
19992
GTTTT
20111
GAA
CAGAATCTACT
20349
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20468


Sau-
CCACGT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GAGCCT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TGC




TTATCTCGTCA





11





ACTTGTTGGCG











AGA








pU6-
GAGGTG
19993
GTTTT
20112
GAA
CAGAATCTACT
20350
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20469


Sau-
TCCACG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TGAGCC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TTGC




TTATCTCGTCA





11G





ACTTGTTGGCG











AGA








pU6-
TGTTCA
19994
GTTTT
20113
GAA
CAGAATCTACT
20351
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20470


Sau-
ATCATT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AAGAA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GACA




TTATCTCGTCA





12





ACTTGTTGGCG











AGA








pU6-
GTGTTC
19995
GTTTT
20114
GAA
CAGAATCTACT
20352
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20471


Sau-
AATCAT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TAAGAA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GACA




TTATCTCGTCA





12G





ACTTGTTGGCG











AGA








pU6-
CGCTTC
19996
GTTTT
20115
GAA
CAGAATCTACT
20353
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20472


Sau-
CTGGGA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGTGTC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CAC




TTATCTCGTCA





13





ACTTGTTGGCG











AGA








pU6-
GCGCTT
19997
GTTTT
20116
GAA
CAGAATCTACT
20354
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20473


Sau-
CCTGGG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AGGTGT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CCAC




TTATCTCGTCA





13G





ACTTGTTGGCG











AGA








pU6-
TCTCCC
19998
GTTTT
20117
GAA
CAGAATCTACT
20355
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20474


Sauri-
CTCCAG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GCCGTG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CAT




TTATCTCGTCA





1





ACTTGTTGGCG











AGA








pU6-
GTCTCC
19999
GTTTT
20118
GAA
CAGAATCTACT
20356
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20475


Sauri-
CCTCCA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGCCGT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GCAT




TTATCTCGTCA





1G





ACTTGTTGGCG











AGA








pU6-
ATGGGT
20000
GTTTT
20119
GAA
CAGAATCTACT
20357
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20476


Sauri-
ATGGCC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TCTAAA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
AAC




TTATCTCGTCA





2





ACTTGTTGGCG











AGA








pU6-
GATGGG
20001
GTTTT
20120
GAA
CAGAATCTACT
20358
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20477


Sauri-
TATGGC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CTCTAA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
AAAC




TTATCTCGTCA





2G





ACTTGTTGGCG





pU6-





AGA








Sauri-
TAAGGC
20002
GTTTT
20121
GAA
CAGAATCTACT
20359
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20478


A1AT-
TGTGCT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



sgRNA-
GACCAT

TCTG


AAATGCCGTGT

GCGAGA



3
CGA




TTATCTCGTCA











ACTTGTTGGCG











AGA








pU6-
GTAAGG
20003
GTTTT
20122
GAA
CAGAATCTACT
20360
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20479


Sauri-
CTGTGC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TGACCA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TCGA




TTATCTCGTCA





3G





ACTTGTTGGCG











AGA








pU6-
AAAAAC
20004
GTTTT
20123
GAA
CAGAATCTACT
20361
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20480


Sauri-
ATGGCC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CCAGCA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GCT




TTATCTCGTCA





4





ACTTGTTGGCG











AGA










 
















pU6-
GAAAAA
20005
GTTTT
20124
GAA
CAGAATCTACT
20362
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20481


Sauri-
CATGGC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CCCAGC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
AGCT




TTATCTCGTCA





4G





ACTTGTTGGCG











AGA








pU6-
GCCTCT
20006
GTTTT
20125
GAA
CAGAATCTACT
20363
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20482


Sauri-
AAAAAC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
ATGGCC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CCA




TTATCTCGTCA





5





ACTTGTTGGCG











AGA








pU6-
ATGGCC
20007
GTTTT
20126
GAA
CAGAATCTACT
20364
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20483


Sauri-
TCTAAA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AACATG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GCC




TTATCTCGTCA





6





ACTTGTTGGCG











AGA








pU6-
GATGGC
20008
GTTTT
20127
GAA
CAGAATCTACT
20365
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20484


Sauri-
CTCTAA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AAACAT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GGCC




TTATCTCGTCA





6G





ACTTGTTGGCG











AGA








pU6-
CTCTCC
20009
GTTTT
20128
GAA
CAGAATCTACT
20366
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20485


Sauri-
CCTCCA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGCCGT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GCA




TTATCTCGTCA





7





ACTTGTTGGCG











AGA








pU6-
GCTCTC
20010
GTTTT
20129
GAA
CAGAATCTACT
20367
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20486


Sauri-
CCCTCC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AGGCCG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
TGCA




TTATCTCGTCA





7G





ACTTGTTGGCG











AGA








pU6-
TGTCTC
20011
GTTTT
20130
GAA
CAGAATCTACT
20368
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20487


Sauri-
TGCTTC

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
TCTCCC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CTC




TTATCTCGTCA





8





ACTTGTTGGCG











AGA








pU6-
GTGTCT
20012
GTTTT
20131
GAA
CAGAATCTACT
20369
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20488


Sauri-
CTGCTT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
CTCTCC

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CCTC




TTATCTCGTCA





8G





ACTTGTTGGCG











AGA








pU6-
TGACCT
20013
GTTTT
20132
GAA
CAGAATCTACT
20370
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20489


Sauri-
CGGGGG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGATAG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
ACA




TTATCTCGTCA





9





ACTTGTTGGCG











AGA








pU6-
GTGACC
20014
GTTTT
20133
GAA
CAGAATCTACT
20371
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20490


Sauri-
TCGGGG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GGGATA

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GACA




TTATCTCGTCA





9G





ACTTGTTGGCG











AGA








pU6-
AAGGGT
20015
GTTTT
20134
GAA
CAGAATCTACT
20372
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20491


Sauri-
TTGTTG

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AACTTG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
ACC




TTATCTCGTCA





10





ACTTGTTGGCG











AGA








pU6-
GAAGGG
20016
GTTTT
20135
GAA
CAGAATCTACT
20373
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20492


Sauri-
TTTGTT

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GAACTT

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
GACC




TTATCTCGTCA





10G





ACTTGTTGGCG











AGA








pU6-
GTGTCC
20017
GTTTT
20136
GAA
CAGAATCTACT
20374
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20493


Sauri-
ACGTGA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
GCCTTG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
CTC




TTATCTCGTCA





11





ACTTGTTGGCG











AGA








pU6-
GTTCAA
20018
GTTTT
20137
GAA
CAGAATCTACT
20375
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACA
20494


Sauri-
TCATTA

AGTAC

A
AAAACAAGGCA

AGGCAAAATGCCGTGTTTATCTCGTCAACTTGTTG



A1AT-
AGAAG

TCTG


AAATGCCGTGT

GCGAGA



sgRNA-
ACAA




TTATCTCGTCA





12





ACTTGTTGGCG











AGA








pU6-
GTAAAA
20019
GTTGT
20138
GAA
CGAAATGAGAA
20376
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20495


Nme2-
ACATGG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
CCCCAG

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
CAGCTT

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



1


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GTCCAG
20020
GTTGT
20139
GAA
CGAAATGAGAA
20377
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20496


Nme2-
GCCGTG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
CATAAG

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
GCTGTG

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



2


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GCATGG
20021
GTTGT
20140
GAA
CGAAATGAGAA
20378
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20497


Nme2-
GTATGG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
CCTCTA

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
AAAACA

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



3


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GACATG
20022
GTTGT
20141
GAA
CGAAATGAGAA
20379
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20498


Nme2-
GGTATG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
GCCTCT

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
AAAAAC

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



4


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GCGTGT
20023
GTTGT
20142
GAA
CGAAATGAGAA
20380
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20499


Nme2-
CTCTGC

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
TTCTCT

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
CCCCTC

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



5


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GCCTTA
20024
GTTGT
20143
GAA
CGAAATGAGAA
20381
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20500


Nme2-
CAACGT

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
GTCTCT

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
GCTTCT

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



6


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GCCTCG
20025
GTTGT
20144
GAA
CGAAATGAGAA
20382
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20501


Nme2-
GGGGGG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
ATAGAC

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
ATGGGT

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



7


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GTGAGC
20026
GTTGT
20145
GAA
CGAAATGAGAA
20383
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20502


Nme2-
CTTGCT

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
CGAGGC

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
CTGGGA

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



8


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GAGAAG
20027
GTTGT
20146
GAA
CGAAATGAGAA
20384
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20503


Nme2-
ACAAAG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
GGTTTG

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
TTGAAC

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



9


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
GAGGTG
20028
GTTGT
20147
GAA
CGAAATGAGAA
20385
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATG
20504


Nme2-
TCCACG

AGCTC

A
CCGTTGCTACA

AGAACCGTTGCTACAATAAGGCCGTCTGAAAAGAT



A1AT-
TGAGCC

CCTTT


ATAAGGCCGTC

GTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTT



sgRNA-
TTGCTC

CTCAT


TGAAAAGATGT

TAAGGGGCATCGTTTA



10


TTCG


GCCGCAACGCT











CTGCCCCTTAA











AGCTTCTGCTT











TAAGGGGCATC











GTTTA








pU6-
TGCATA
20029
GCTAT
20148
GAA
GGTAAGTTGCT
20386
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20505


Blat-
AGGCTG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
TGCTGA

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
CCA

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



1





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GTGCAT
20030
GCTAT
20149
GAA
GGTAAGTTGCT
20387
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20506


Blat-
AAGGCT

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
GTGCTG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
ACCA

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



1G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
TGGCCC
20031
GCTAT
20150
GAA
GGTAAGTTGCT
20388
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20507


Blat-
CAGCAG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
CTTCAG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
TCC

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



2





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GTGGCC
20032
GCTAT
20151
GAA
GGTAAGTTGCT
20389
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20508


Blat-
CCAGCA

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
GCTTCA

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
GTCC

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



2G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
CCGTGC
20033
GCTAT
20152
GAA
GGTAAGTTGCT
20390
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20509


Blat-
ATAAGG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
CTGTGC

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
TGA

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



3





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GCCGTG
20034
GCTAT
20153
GAA
GGTAAGTTGCT
20391
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20510


Blat-
CATAAG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
GCTGTG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
CTGA

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



3G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
AGGCCG
20035
GCTAT
20154
GAA
GGTAAGTTGCT
20392
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20511


Blat-
TGCATA

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
AGGCTG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
TGC

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



4





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GAGGC
20036
GCTAT
20155
GAA
GGTAAGTTGCT
20393
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20512


Blat-
CGTGCA

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
TAAGGC

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
TGTGC

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



4G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
AAAACA
20037
GCTAT
20156
GAA
GGTAAGTTGCT
20394
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20513


Blat-
TGGCCC

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
CAGCAG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
CTT

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



5





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GAAAAC
20038
GCTAT
20157
GAA
GGTAAGTTGCT
20395
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20514


Blat-
ATGGCC

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
CCAGCA

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
GCTT

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



5G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GGGGGG
20039
GCTAT
20158
GAA
GGTAAGTTGCT
20396
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20515


Blat-
ATAGAC

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
ATGGGT

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
ATG

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



6





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
TGAACT
20040
GCTAT
20159
GAA
GGTAAGTTGCT
20397
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20516


Blat-
TGACCT

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
CGGGGG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
GGA

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



7





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GTGAAC
20041
GCTAT
20160
GAA
GGTAAGTTGCT
20398
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20517


Blat-
TTGACC

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
TCGGGG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
GGGA

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



7G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
TCGAGG
20042
GCTAT
20161
GAA
GGTAAGTTGCT
20399
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20518


Blat-
CCTGGG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
ATCAGC

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
CTT

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



8





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GTCGAG
20043
GCTAT
20162
GAA
GGTAAGTTGCT
20400
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20519


Blat-
GCCTGG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
GATCAG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
CCTT

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



8G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
ACAAAG
20044
GCTAT
20163
GAA
GGTAAGTTGCT
20401
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20520


Blat-
GGTTTG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
TTGAAC

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
TTG

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



9





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GACAAA
20045
GCTAT
20164
GAA
GGTAAGTTGCT
20402
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20521


Blat-
GGGTTT

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
GTTGAA

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
CTTG

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



9G





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
GCCTTG
20046
GCTAT
20165
GAA
GGTAAGTTGCT
20403
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAG
20522


Blat-
CTCGAG

AGTTC

A
ATAGTAAGGGC

TAAGGGCAACAGACCCGAGGCGTTGGGGATCGCCT



A1AT-
GCCTGG

CTTAC


AACAGACCCGA

AGCCCGTGTTTACGGGCTCTCCCCATATTCAAAAT



sgRNA-
GAT

T


GGCGTTGGGGA

AATGACAGACGAGCACCTTGGAGCATTTATCTCCG



10





TCGCCTAGCCC

AGGTGCT









GTGTTTACGGG











CTCTCCCCATA











TTCAAAATAAT











GACAGACGAGC











ACCTTGGAGCA











TTTATCTCCGA











GGTGCT








pU6-
AAAGGG
20047
GTTGT
20166
GAA
GCGAAATGAAA
20404
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20523


Ppn-
ACTGAA

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
GCTGCT

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
GGGG

TTCAT


AATTTCTCGCA

GGCATC



1


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GAAAGG
20048
GTTGT
20167
GAA
GCGAAATGAAA
20405
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20524


Ppn-
GACTGA

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
AGCTGC

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
TGGGG

TTCAT


AATTTCTCGCA

GGCATC



1G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
CCTGGA
20049
GTTGT
20168
GAA
GCGAAATGAAA
20406
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20525


Ppn-
GGGGAG

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
AGAAGC

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
AGAG

TTCAT


AATTTCTCGCA

GGCATC



2


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GCCTGG
20050
GTTGT
20169
GAA
GCGAAATGAAA
20407
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20526


Ppn-
AGGGGA

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
GAGAAG

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
CAGAG

TTCAT


AATTTCTCGCA

GGCATC



2G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
CCCATG
20051
GTTGT
20170
GAA
GCGAAATGAAA
20408
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20527


Ppn-
TCTATC

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
CCCCCC

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
GAGG

TTCAT


AATTTCTCGCA

GGCATC



3


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GCCCAT
20052
GTTGT
20171
GAA
GCGAAATGAAA
20409
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20528


Ppn-
GTCTAT

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
CCCCCC

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
CGAGG

TTCAT


AATTTCTCGCA

GGCATC



3G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
TCAATC
20053
GTTGT
20172
GAA
GCGAAATGAAA
20410
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20529


Ppn-
ATTAAG

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
AAGACA

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
AAGG

TTCAT


AATTTCTCGCA

GGCATC



4


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GTCAAT
20054
GTTGT
20173
GAA
GCGAAATGAAA
20411
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20530


Ppn-
CATTAA

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
GAAGAC

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
AAAGG

TTCAT


AATTTCTCGCA

GGCATC



4G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
TTGTTC
20055
GTTGT
20174
GAA
GCGAAATGAAA
20412
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20531


Ppn-
AATCAT

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
TAAGAA

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
GACA

TTCAT


AATTTCTCGCA

GGCATC



5


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GTTGTT
20056
GTTGT
20175
GAA
GCGAAATGAAA
20413
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20532


Ppn-
CAATCA

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
TTAAGA

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
AGACA

TTCAT


AATTTCTCGCA

GGCATC



5G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
TCAACA
20057
GTTGT
20176
GAA
GCGAAATGAAA
20414
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20533


Ppn-
AACCCT

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
TTGTCT

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
TCTT

TTCAT


AATTTCTCGCA

GGCATC



6


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GTCAAC
20058
GTTGT
20177
GAA
GCGAAATGAAA
20415
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20534


Ppn-
AAACCC

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
TTTGTC

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
TTCTT

TTCAT


AATTTCTCGCA

GGCATC



6G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GGGGAG
20059
GTTGT
20178
GAA
GCGAAATGAAA
20416
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20535


Ppn-
ACTTGG

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
TATTTT

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
GTTC

TTCAT


AATTTCTCGCA

GGCATC



7


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
CATGAA
20060
GTTGT
20179
GAA
GCGAAATGAAA
20417
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20536


Ppn-
GAGGGG

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
AGACTT

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
GGTA

TTCAT


AATTTCTCGCA

GGCATC



8


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GCATGA
20061
GTTGT
20180
GAA
GCGAAATGAAA
20418
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20537


Ppn-
AGAGGG

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
GAGACT

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
TGGTA

TTCAT


AATTTCTCGCA

GGCATC



8G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
TTTCCC
20062
GTTG
20181
GAA
GCGAAATGAAA
20419
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20538


Ppn-
ATGAAG

TAGC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
AGGGGA

TCCCT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
GACT

TTTTC


AATTTCTCGCA

GGCATC



9


ATTTC


AAGCTCTGCCT








GC


CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
GTTTCC
20063
GTTGT
20182
GAA
GCGAAATGAAA
20420
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAA
20539


Ppn-
CATGAA

AGCTC

A
AACGTTGTTAC

TGAAAAACGTTGTTACAATAAGAGATGAATTTCTC



A1AT-
GAGGGG

CCTTT


AATAAGAGATG

GCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGA



sgRNA-
AGACT

TTCAT


AATTTCTCGCA

GGCATC



9G


TTCGC


AAGCTCTGCCT











CTTGAAATTTC











GGTTTCAAGAG











GCATC








pU6-
AAGGCT
20064
GTCTT
20183
GTA
CAGAAGCTACA
20421
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATA
20540


St1-
GTGCTG

TGTAC

C
AAGATAAGGCT

AGGCTTCATGCCGAAATCAACACCCTGTCATTTTA



A1AT-
ACCATC

TCTG


TCATGCCGAAA

TGGCAGGGTGTTTT



sgRNA-
GA




TCAACACCCTG





1





TCATTTTATGG











CAGGGTGTTTT








pU6-
GAAGGC
20065
GTCTT
20184
GTA
CAGAAGCTACA
20422
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATA
20541


St1-
TGTGCT

TGTAC

C
AAGATAAGGCT

AGGCTTCATGCCGAAATCAACACCCTGTCATTTTA



A1AT-
GACCAT

TCTG


TCATGCCGAAA

TGGCAGGGTGTTTT



sgRNA-
CGA




TCAACACCCTG





1G





TCATTTTATGG











CAGGGTGTTTT








pU6-
AAGGCT
20066
GTCTT
20185
GTA
CAGAAGCTACA
20423
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATA
20542


St1-
CACGTG

TGTAC

C
AAGATAAGGCT

AGGCTTCATGCCGAAATCAACACCCTGTCATTTTA



A1AT-
GACACC

TCTG


TCATGCCGAAA

TGGCAGGGTGTTTT



sgRNA-
TC




TCAACACCCTG





2





TCATTTTATGG











CAGGGTGTTTT








pU6-
GAAGGC
20067
GTCTT
20186
GTA
CAGAAGCTACA
20424
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATA
20543


St1-
TCACGT

TGTAC

C
AAGATAAGGCT

AGGCTTCATGCCGAAATCAACACCCTGTCATTTTA



A1AT-
GGACAC

TCTG


TCATGCCGAAA

TGGCAGGGTGTTTT



sgRNA-
CTC




TCAACACCCTG





2G





TCATTTTATGG











CAGGGTGTTTT








pU6-
TACCAA
20068
GTCTT
20187
GTA
CAGAAGCTACA
20425
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATA
20544


St1-
GTCTCC

TGTAC

C
AAGATAAGGCT

AGGCTTCATGCCGAAATCAACACCCTGTCATTTTA



A1AT-
CCTCTT

TCTG


TCATGCCGAAA

TGGCAGGGTGTTTT



sgRNA-
CA




TCAACACCCTG





3





TCATTTTATGG











CAGGGTGTTTT








pU6-
GTACCA
20069
GTCTT
20188
GTA
CAGAAGCTACA
20426
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATA
20545


St1-
AGTCTC

TGTAC

C
AAGATAAGGCT

AGGCTTCATGCCGAAATCAACACCCTGTCATTTTA



A1AT-
CCCTCT

TCTG


TCATGCCGAAA

TGGCAGGGTGTTTT



sgRNA-
TCA




TCAACACCCTG





3G





TCATTTTATGG











CAGGGTGTTTT









In some embodiments, the systems and methods provided herein may comprise a template sequence, or component thereof, listed in Table 6B, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto. Table 6B provides exemplary template RNA sequences designed to be paired with a gene modifying polypeptide to correct a mutation in the SERPINA1 gene.









TABLE 6B







Exemplary template RNA sequences


Table 6B provides design of exemplary DNA components of gene modifying systems for correcting the pathogenic E342K mutation in


SERPINA1 to the wild-type form. This table details the sequence of a complete template RNA for use in exemplary gene modifying systems


comprising a gene modifying polypeptide.

















SEQ ID

SEQ

SEQ ID

SEQ ID


Name
Spacer
NO
PBS
ID NO
RT
NO
tgRNA sequence
NO





pU6_A1AT_
CTGTGC
20546
GTCGATGG
21356
ACATGGCCC
22166
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22976


SpRY_
TGACCA

TCAGCACA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G

CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB17






CGATGGTCAGCACAG






pU6_A1AT_
CTGTGC
20547
GTCGATGG
21357
ACATGGCCC
22167
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22977


SpRY_
TGACCA

TCAGCACA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB16






CGATGGTCAGCACA






pU6_A1AT_
CTGTGC
20548
GTCGATGG
21358
ACATGGCCC
22168
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22978


SpRY_
TGACCA

TCAGCAC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB15






CGATGGTCAGCAC






pU6_A1AT_
CTGTGC
20549
GTCGATGG
21359
ACATGGCCC
22169
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22979


SpRY_
TGACCA

TCAGCA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB14






CGATGGTCAGCA






pU6_A1AT_
CTGTGC
20550
GTCGATGG
21360
ACATGGCCC
22170
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22980


SpRY_
TGACCA

TCAGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB13






CGATGGTCAGC






pU6_A1AT_
CTGTGC
20551
GTCGATGG
21361
ACATGGCCC
22171
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22981


SpRY_
TGACCA

TCAG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB12






CGATGGTCAG






pU6_A1AT_
CTGTGC
20552
GTCGATGG
21362
ACATGGCCC
22172
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22982


SpRY_
TGACCA

TCA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB11






CGATGGTCA






pU6_A1AT_
CTGTGC
20553
GTCGATGG
21363
ACATGGCCC
22173
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22983


SpRY_
TGACCA

TC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB10






CGATGGTC






pU6_A1AT_
CTGTGC
20554
GTCGATGG

ACATGGCCC
22174
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22984


SpRY_
TGACCA

T

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB9






CGATGGT






pU6_A1AT_
CTGTGC
20555
GTCGATGG

ACATGGCCC
22175
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22985


SpRY_
TGACCA



CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
AG



TCTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB8






CGATGG






pU6_A1AT_
CTGTGC
20556
GTCGATGG
21366
GCCCCAGCA
22176
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22986


SpRY_
TGACCA

TCAGCACA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G

CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB17






GTCAGCACAG






pU6_A1AT_
CTGTGC
20557
GTCGATGG
21367
GCCCCAGCA
22177
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22987


SpRY_
TGACCA

TCAGCACA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB16






GTCAGCACA






pU6_A1AT_
CTGTGC
20558
GTCGATGG
21368
GCCCCAGCA
22178
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22988


SpRY_
TGACCA

TCAGCAC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB15






GTCAGCAC






pU6_A1AT_
CTGTGC
20559
GTCGATGG
21369
GCCCCAGCA
22179
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA



SpRY_
TGACCA

TCAGCA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT
22989


ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



 G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB14






GTCAGCA






pU6_A1AT_
CTGTGC
20560
GTCGATGG
21370
GCCCCAGCA
22180
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22990


SpRY_
TGACCA

TCAGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB13






GTCAGC






pU6_A1AT_
CTGTGC
20561
GTCGATGG
21371
GCCCCAGCA
22181
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22991


SpRY_
TGACCA

TCAG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB12






GTCAG






pU6_A1AT_
CTGTGC
20562
GTCGATGG
21372
GCCCCAGCA
22182
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22992


SpRY_
TGACCA

TCA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB11






GTCA






pU6_A1
CTGTGC
20563
GTCGATGG
21373
GCCCCAGCA
22183
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22993


AT_SpR
TGACCA

TC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25F
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



E_PB10






GTC






pU6_A1AT_
CTGTGC
20564
GTCGATGG

GCCCCAGCA
22184
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22994


SpRY_
TGACCA

T

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB9






GT






pU6_A1AT_
CTGTGC
20565
GTCGATGG

GCCCCAGCA
22185
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22995


SpRY_
TGACCA



GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CCTTTCTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
AG





GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB8






G






pU6_A1AT_
CTGTGC
20566
GTCGATGG
21376
AGCAGCTTC
22186
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22996


SpRY_
TGACCA

TCAGCACA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G

CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB17






CACAG






pU6_A1AT_
CTGTGC
20567
GTCGATGG
21377
AGCAGCTTC
22187
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22997


SpRY_
TGACCA

TCAGCACA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB16






CACA






pU6_A1AT_
CTGTGC
20568
GTCGATGG
21378
AGCAGCTTC
22188
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22998


SpRY_
TGACCA

TCAGCAC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB15






CAC






pU6_A1
CTGTGC
20569
GTCGATGG
21379
AGCAGCTTC
22189
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
22999


AT_SpR
TGACCA

TCAGCA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



G_20F
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



E_PB14






CA






pU6_A1AT_
CTGTGC
20570
GTCGATGG
21380
AGCAGCTTC
22190
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23000


SpRY_
TGACCA

TCAGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB13






C






pU6_A1AT_
CTGTGC
20571
GTCGATGG
21381
AGCAGCTTC
22191
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23001


SpRY_
TGACCA

TCAG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20572
GTCGATGG
21382
AGCAGCTTC
22192
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23002


SpRY_
TGACCA

TCA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20573
GTCGATGG
21383
AGCAGCTTC
22193
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23003


SpRY_
TGACCA

TC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20574
GTCGATGG

AGCAGCTTC
22194
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23004


SpRY_
TGACCA

T

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGGT



PB9













pU6_A1AT_
CTGTGC
20575
GTCGATGG

AGCAGCTTC
22195
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23005


SpRY_
TGACCA



AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA



CTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
AG





AGCAGCTTCAGTCCCTTTCTCGTCGATGG



PB8













pU6_A1AT_
CTGTGC
20576
GTCGATGG
21386
TTCAGTCCC
22196
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23006


SpRY_
TGACCA

TCAGCACA

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTCAGCACAG



PB17













pU6_A1AT_
CTGTGC
20577
GTCGATGG
21387
TTCAGTCCC
22197
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23007


SpRY_
TGACCA

TCAGCACA

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTCAGCACA



PB16













pU6_A1AT_
CTGTGC
20578
GTCGATGG
21388
TTCAGTCCC
22198
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23008


SpRY_
TGACCA

TCAGCAC

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14F
AG





TTCAGTCCCTTTCTCGTCGATGGTCAGCAC



E_PB15













pU6_A1AT_
CTGTGC
20579
GTCGATGG
21389
TTCAGTCCC
22199
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23009


SpRY_
TGACCA

TCAGCA

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTCAGCA



PB14













pU6_A1AT_
CTGTGC
20580
GTCGATGG
21390
TTCAGTCCC
22200
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23010


SpRY_
TGACCA

TCAGC

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTCAGC



PB13













pU6_A1AT_
CTGTGC
20581
GTCGATGG
21391
TTCAGTCCC
22201
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23011


SpRY_
TGACCA

TCAG

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20582
GTCGATGG
21392
TTCAGTCCC
22202
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23012


SpRY_
TGACCA

TCA

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20583
GTCGATGG
21393
TTCAGTCCC
22203
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23013


SpRY_
TGACCA

TC

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20584
GTCGATGG

TTCAGTCCC
22204
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23014


SpRY_
TGACCA

T

TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGGT



PB9










pU6_A1AT_
CTGTGC
20585
GTCGATGG

TTCAGTCCC
22205
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23015


SpRY_
TGACCA



TTTCTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
AG





TTCAGTCCCTTTCTCGTCGATGG



PB8













pU6_A1
CTGTGC
20586
GTCGATGG
21396
AGTCCCTTT
22206
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23016


AT_SpR
TGACCA

TCAGCACA

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED0-
TCGACA

G



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



G 11F
AG





AGTCCCTTTCTCGTCGATGGTCAGCACAG



E_PB17













pU6_A1AT_
CTGTGC
20587
GTCGATGG
21397
AGTCCCTTT
22207
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23017


SpRY_
TGACCA

TCAGCACA

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTCAGCACA



PB16













pU6_A1AT_
CTGTGC
20588
GTCGATGG
21398
AGTCCCTTT
22208
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23018


SpRY_
TGACCA

TCAGCAC

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTCAGCAC



PB15













pU6_A1AT_
CTGTGC
20589
GTCGATGG
21399
AGTCCCTTT
22209
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23019


SpRY_
TGACCA

TCAGCA

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTCAGCA



PB14













pU6_A1AT_
CTGTGC
20590
GTCGATGG
21400
AGTCCCTTT
22210
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23020


SpRY_
TGACCA

TCAGC

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTCAGC



PB13













pU6_A1AT_
CTGTGC
20591
GTCGATGG
21401
AGTCCCTTT
22211
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23021


SpRY_
TGACCA

TCAG

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20592
GTCGATGG
21402
AGTCCCTTT
22212
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23022


SpRY_
TGACCA

TCA

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20593
GTCGATGG
21403
AGTCCCTTT
22213
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23023


SpR
TGACCA

TC

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20594
GTCGATGG

AGTCCCTTT
22214
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23024


SpRY_
TGACCA

T

CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGGT



PB9













pU6_A1AT_
CTGTGC
20595
GTCGATGG

AGTCCCTTT
22215
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23025


SpRY_
TGACCA



CTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
AG





AGTCCCTTTCTCGTCGATGG



PB8













pU6_A1AT_
CTGTGC
20596
GTCGATGG
21406
TCCCTTTCTC
22216
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23026


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCAGCACAG



PB17













pU6_A1AT_
CTGTGC
20597
GTCGATGG
21407
TCCCTTTCTC
22217
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23027


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCAGCACA



PB16













pU6_A1AT_
CTGTGC
20598
GTCGATGG
21408
TCCCTTTCTC
22218
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23028


SpRY_
TGACCA

TCAGCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCAGCAC



PB15













pU6_A1AT_
CTGTGC
20599
GTCGATGG
21409
TCCCTTTCTC
22219
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23029


SpRY_
TGACCA

TCAGCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCAGCA



PB14













pU6_A1AT_
CTGTGC
20600
GTCGATGG
21410
TCCCTTTCTC
22220
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23030


SpRY_
TGACCA

TCAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCAGC



PB13













pU6_A1AT_
CTGTGC
20601
GTCGATGG
21411
TCCCTTTCTC
22221
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23031


SpRY_
TGACCA

TCAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20602
GTCGATGG
21412
TCCCTTTCTC
22222
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23032


SpRY_
TGACCA

TCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20603
GTCGATGG
21413
TCCCTTTCTC
22223
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23033


SpRY_
TGACCA

TC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20604
GTCGATGG

TCCCTTTCTC
22224
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23034


SpRY_
TGACCA

T



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGGT



PB9













pU6_A1AT_
CTGTGC
20605
GTCGATGG

TCCCTTTCTC
22225
CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23035


SpRY_
TGACCA





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
AG





TCCCTTTCTCGTCGATGG



PB8













pU6_A1AT_
CTGTGC
20606
GTCGATGG
21416
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23036


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCAGCACAG



PB17













pU6_A1AT_
CTGTGC
20607
GTCGATGG
21417
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23037


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCAGCACA



PB16













pU6_A1AT_
CTGTGC
20608
GTCGATGG
21418
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23038


SpRY_
TGACCA

TCAGCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCAGCAC



PB15













pU6_A1AT_
CTGTGC
20609
GTCGATGG
21419
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23039


SpRY_
TGACCA

TCAGCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCAGCA



PB14













pU6_A1AT_
CTGTGC
20610
GTCGATGG
21420
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23040


SpRY_
TGACCA

TCAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCAGC



PB13













pU6_A1AT_
CTGTGC
20611
GTCGATGG
21421
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23041


SpRY_
TGACCA

TCAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20612
GTCGATGG
21422
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23042


SpRY_
TGACCA

TCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20613
GTCGATGG
21423
CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23043


SpRY_
TGACCA

TC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20614
GTCGATGG

CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23044


SpR
TGACCA

T



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGGT



PB9













pU6_A1AT_
CTGTGC
20615
GTCGATGG

CCTTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23045


SpRY_
TGACCA





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
AG





CCTTTCTCGTCGATGG



PB8













pU6_A1AT_
CTGTGC
20616
GTCGATGG
21426
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23046


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCAGCACAG



PB17













pU6_A1AT_
CTGTGC
20617
GTCGATGG
21427
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23047


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCAGCACA



PB16













pU6_A1AT_
CTGTGC
20618
GTCGATGG
21428
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23048


SpRY_
TGACCA

TCAGCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCAGCAC



PB15













pU6_A1AT_
CTGTGC
20619
GTCGATGG
21429
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23049


SpRY_
TGACCA

TCAGCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCAGCA



PB14













pU6_A1AT_
CTGTGC
20620
GTCGATGG
21430
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23050


SpRY_
TGACCA

TCAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCAGC



PB13













pU6_A1AT_
CTGTGC
20621
GTCGATGG
21431
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23051


SpRY_
TGACCA

TCAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20622
GTCGATGG
21432
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23052


SpRY_
TGACCA

TCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20623
GTCGATGG
21433
TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23053


SpRY_
TGACCA

TC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20624
GTCGATGG

TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23054


SpRY_
TGACCA

T



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGGT



PB9













pU6_A1AT_
CTGTGC
20625
GTCGATGG

TTTCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23055


SpRY_
TGACCA





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
AG





TTTCTCGTCGATGG



_PB8













pU6_A1AT_
CTGTGC
20626
GTCGATGG
21436
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23056


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA

G



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCAGCACAG



PB17













pU6_A1AT_
CTGTGC
20627
GTCGATGG
21437
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23057


SpRY_
TGACCA

TCAGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCAGCACA



PB16













pU6_A1AT_
CTGTGC
20628
GTCGATGG
21438
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23058


SpRY_
TGACCA

TCAGCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCAGCAC



PB15













pU6_A1AT_
CTGTGC
20629
GTCGATGG
21439
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23059


SpRY_
TGACCA

TCAGCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCAGCA



_PB14













pU6_A1AT_
CTGTGC
20630
GTCGATGG
21440
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23060


SpRY_
TGACCA

TCAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCAGC



PB13













pU6_A1AT_
CTGTGC
20631
GTCGATGG
21441
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23061


SpRY_
TGACCA

TCAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCAG



PB12













pU6_A1AT_
CTGTGC
20632
GTCGATGG
21442
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23062


SpRY_
TGACCA

TCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTCA



PB11













pU6_A1AT_
CTGTGC
20633
GTCGATGG
21443
TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23063


SpRY_
TGACCA

TC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGTC



PB10













pU6_A1AT_
CTGTGC
20634
GTCGATGG

TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23064


SpRY_
TGACCA

T



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGGT



PB9













pU6_A1AT_
CTGTGC
20635
GTCGATGG

TCTC

CTGTGCTGACCATCGACAAGGTTTTAGAGCTA
23065


SpRY_
TGACCA





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED0-
TCGACA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
AG





TCTCGTCGATGG



PB8













pU6_A1AT_
GGCTGT
20636
CGATGGTC
21446
ACATGGCCC
22256
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23066


SpRY_
GCTGAC

AGCACAGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C

CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB17






CGATGGTCAGCACAGCC






pU6_A1AT_
GGCTGT
20637
CGATGGTC
21447
ACATGGCCC
22257
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23067


SpRY_
GCTGAC

AGCACAGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB16






CGATGGTCAGCACAGC






pU6_A1AT_
GGCTGT
20638
CGATGGTC
21448
ACATGGCCC
22258
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23068


SpRY_
GCTGAC

AGCACAG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB15






CGATGGTCAGCACAG






pU6_A1AT_
GGCTGT
20639
CGATGGTC
21449
ACATGGCCC
22259
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23069


SpRY_
GCTGAC

AGCACA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB14






CGATGGTCAGCACA






pU6_A1AT_
GGCTGT
20640
CGATGGTC
21450
ACATGGCCC
22260
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23070


SpRY_
GCTGAC

AGCAC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB13






CGATGGTCAGCAC






pU6_A1AT_
GGCTGT
20641
CGATGGTC
21451
ACATGGCCC
22261
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23071


SpRY_
GCTGAC

AGCA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB12






CGATGGTCAGCA






pU6_A1AT_
GGCTGT
20642
CGATGGTC
21452
ACATGGCCC
22262
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23072


SpRY_
GCTGAC

AGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB11






CGATGGTCAGC






pU6_A1AT_
GGCTGT
20643
CGATGGTC
21453
ACATGGCCC
22263
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23073


SpRY_
GCTGAC

AG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



BP10






CGATGGTCAG






pU6_A1AT_
GGCTGT
20644
CGATGGTC

ACATGGCCC
22264
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23074


SpRY_
GCTGAC

A

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB9






CGATGGTCA






pU6_A1AT_
GGCTGT
20645
CGATGGTC

ACATGGCCC
22265
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23075


SpRY_
GCTGAC



CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
CA



TCTCGT

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



PB8






CGATGGTC






pU6_A1AT_
GGCTGT
20646
CGATGGTC
21456
GCCCCAGCA
22266
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23076


SpRY_
GCTGAC

AGCACAGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C

CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



BP17






GTCAGCACAGCC






pU6_A1AT_
GGCTGT
20647
CGATGGTC
21457
GCCCCAGCA
22267
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23077


SpRY_
GCTGAC

AGCACAGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB16






GTCAGCACAGC






pU6_A1AT_
GGCTGT
20648
CGATGGTC
21458
GCCCCAGCA
22268
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23078


SpRY_
GCTGAC

AGCACAG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB15






GTCAGCACAG






pU6_A1AT_
GGCTGT
20649
CGATGGTC
21459
GCCCCAGCA
22269
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23079


SpRY_
GCTGAC

AGCACA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB14






GTCAGCACA






pU6_A1AT_
GGCTGT
20650
CGATGGTC
21460
GCCCCAGCA
22270
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23080


SpRY_
GCTGAC

AGCAC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB13






GTCAGCAC






pU6_A1AT_
GGCTGT
20651
CGATGGTC
21461
GCCCCAGCA
22271
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23081


SpRY_
GCTGAC

AGCA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB12






GTCAGCA






pU6_A1AT_
GGCTGT
20652
CGATGGTC
21462
GCCCCAGCA
22272
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23082


SpRY_
GCTGAC

AGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB11






GTCAGC






pU6_A1AT_
GGCTGT
20653
CGATGGTC
21463
GCCCCAGCA
22273
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23083


SpRY_
GCTGAC

AG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB10






GTCAG






pU6_A1AT_
GGCTGT
20654
CGATGGTC

GCCCCAGCA
22274
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23084


SpRY_
GCTGAC

A

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB9






GTCA






pU6_A1AT_
GGCTGT
20655
CGATGGTC

GCCCCAGCA
22275
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23085


SpRY_
GCTGAC



GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CCTTTCTCG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_25FE_
CA



T

GCCCCAGCAGCTTCAGTCCCTTTCTCGTCGATG



PB8






GTC






pU6_A1AT_
GGCTGT
20656
CGATGGTC
21466
AGCAGCTTC
22276
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23086


SpRY_
GCTGAC

AGCACAGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C

CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB17






CACAGCC






pU6_A1AT_
GGCTGT
20657
CGATGGTC
21467
AGCAGCTTC
22277
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23087


SpRY_
GCTGAC

AGCACAGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB16






CACAGC






pU6_A1AT_
GGCTGT
20658
CGATGGTC
21468
AGCAGCTTC
22278
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23088


SpRY_
GCTGAC

AGCACAG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB15






CACAG






pU6_A1AT_
GGCTGT
20659
CGATGGTC
21469
AGCAGCTTC
22279
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23089


SpRY_
GCTGAC

AGCACA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB14






CACA






pU6_A1
GGCTGT
20660
CGATGGTC
21470
AGCAGCTTC
22280
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23090


AT_SpR
GCTGAC

AGCAC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



20FE_P
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



B13






CAC






pU6_A1AT_
GGCTGT
20661
CGATGGTC
21471
AGCAGCTTC
22281
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23091


SpRY_
GCTGAC

AGCA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB12






CA






pU6_A1AT_
GGCTGT
20662
CGATGGTC
21472
AGCAGCTTC
22282
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23092


SpRY_
GCTGAC

AGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB11






C






pU6_A1AT_
GGCTGT
20663
CGATGGTC
21473
AGCAGCTTC
22283
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23093


SpRY_
GCTGAC

AG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20664
CGATGGTC

AGCAGCTTC
22284
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23094


SpRY_
GCTGAC

A

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20665
CGATGGTC

AGCAGCTTC
22285
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23095


SpRY_
GCTGAC



AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA



CTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_20FE_
CA





AGCAGCTTCAGTCCCTTTCTCGTCGATGGTC



PB8













pU6_A1AT_
GGCTGT
20666
CGATGGTC
21476
TTCAGTCCC
22286
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23096


SpRY_
GCTGAC

AGCACAGC

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGCACAGC



PB17






C






pU6_A1AT_
GGCTGT
20667
CGATGGTC
21477
TTCAGTCCC
22287
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23097


SpRY_
GCTGAC

AGCACAGC

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGCACAGC



PB16













pU6_A1AT_
GGCTGT
20668
CGATGGTC
21478
TTCAGTCCC
22288
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23098


SpRY_
GCTGAC

AGCACAG

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGCACAG



PB15













pU6_A1AT_
GGCTGT
20669
CGATGGTC
21479
TTCAGTCCC
22289
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23099


SpRY_
GCTGAC

AGCACA

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGCACA



PB14













pU6_A1AT_
GGCTGT
20670
CGATGGTC
21480
TTCAGTCCC
22290
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23100


SpRY_
GCTGAC

AGCAC

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGCAC



PB13













pU6_A1AT_
GGCTGT
20671
CGATGGTC
21481
TTCAGTCCC
22291
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23101


SpRY_
GCTGAC

AGCA

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGCA



PB12













pU6_A1AT_
GGCTGT
20672
CGATGGTC
21482
TTCAGTCCC
22292
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23102


SpRY_
GCTGAC

AGC

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAGC



PB11













pU6_A1AT_
GGCTGT
20673
CGATGGTC
21483
TTCAGTCCC
22293
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23103


SpRY_
GCTGAC

AG

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20674
CGATGGTC

TTCAGTCCC
22294
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23104


SpRY_
GCTGAC

A

TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20675
CGATGGTC

TTCAGTCCC
22295
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23105


SpRY_
GCTGAC



TTTCTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_14FE_
CA





TTCAGTCCCTTTCTCGTCGATGGTC



PB8













pU6_A1AT_
GGCTGT
20676
CGATGGTC
21486
AGTCCCTTT
22296
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23106


SpRY_
GCTGAC

AGCACAGC

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGCACAGCC



PB17













pU6_A1AT_
GGCTGT
20677
CGATGGTC
21487
AGTCCCTTT
22297
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23107


SpRY_
GCTGAC

AGCACAGC

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGCACAGC



PB16













pU6_A1AT_
GGCTGT
20678
CGATGGTC
21488
AGTCCCTTT
22298
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23108


SpRY_
GCTGAC

AGCACAG

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGCACAG



PB15













pU6_A1AT_
GGCTGT
20679
CGATGGTC
21489
AGTCCCTTT
22299
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23109


SpRY_
GCTGAC

AGCACA

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGCACA



PB14













pU6_A1AT_
GGCTGT
20680
CGATGGTC
21490
AGTCCCTTT
22300
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23110


SpRY_
GCTGAC

AGCAC

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGCAC



PB13













pU6_A1AT_
GGCTGT
20681
CGATGGTC
21491
AGTCCCTTT
22301
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23111


SpRY_
GCTGAC

AGCA

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGCA



PB12













pU6_A1AT_
GGCTGT
20682
CGATGGTC
21492
AGTCCCTTT
22302
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23112


SpRY_
GCTGAC

AGC

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAGC



PB11













pU6_A1AT_
GGCTGT
20683
CGATGGTC
21493
AGTCCCTTT
22303
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23113


SpRY_
GCTGAC

AG

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20684
CGATGGTC

AGTCCCTTT
22304
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23114


SpRY_
GCTGAC

A

CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20685
CGATGGTC

AGTCCCTTT
22305
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23115


SpRY_
GCTGAC



CTCGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_11FE_
CA





AGTCCCTTTCTCGTCGATGGTC



PB8













pU6_A1AT_
GGCTGT
20686
CGATGGTC
21496
TCCCTTTCTC
22306
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23116


SpRY_
GCTGAC

AGCACAGC

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGCACAGCC



PB17













pU6_A1AT_
GGCTGT
20687
CGATGGTC
21497
TCCCTTTCTC
22307
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23117


SpRY_
GCTGAC

AGCACAGC

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGCACAGC



PB16













pU6_A1AT_
GGCTGT
20688
CGATGGTC
21498
TCCCTTTCTC
22308
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23118


SpRY_
GCTGAC

AGCACAG

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGCACAG



PB15













pU6_A1AT_
GGCTGT
20689
CGATGGTC
21499
TCCCTTTCTC
22309
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23119


SpRY_
GCTGAC

AGCACA

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGCACA



PB14













pU6_A1AT_
GGCTGT
20690
CGATGGTC
21500
TCCCTTTCTC
22310
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23120


SpRY_
GCTGAC

AGCAC

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGCAC



PB13













pU6_A1AT_
GGCTGT
20691
CGATGGTC
21501
TCCCTTTCTC
22311
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23121


SpRY_
GCTGAC

AGCA

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGCA



PB12













pU6_A1AT_
GGCTGT
20692
CGATGGTC
21502
TCCCTTTCTC
22312
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23122


SpRY_
GCTGAC

AGC

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAGC



PB11













pU6_A1AT_
GGCTGT
20693
CGATGGTC
21503
TCCCTTTCTC
22313
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23123


SpRY_
GCTGAC

AG

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20694
CGATGGTC

TCCCTTTCTC
22314
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23124


SpRY_
GCTGAC

A

GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20695
CGATGGTC

TCCCTTTCTC
22315
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23125


SpRY_
GCTGAC



GT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_9FE_
CA





TCCCTTTCTCGTCGATGGTC



PB8













pU6_A1AT_
GGCTGT
20696
CGATGGTC
21506
CCTTTCTCG
22316
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23126


SpR
GCTGAC

AGCACAGC

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED2-
CATCGA

C



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGCACAGCC



PB17













pU6_A1AT_
GGCTGT
20697
CGATGGTC
21507
CCTTTCTCG
22317
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23127


SpRY_
GCTGAC

AGCACAGC

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGCACAGC



PB16













pU6_A1AT_
GGCTGT
20698
CGATGGTC
21508
CCTTTCTCG
22318
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23128


SpRY_
GCTGAC

AGCACAG

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGCACAG



PB15













pU6_A1AT_
GGCTGT
20699
CGATGGTC
21509
CCTTTCTCG
22319
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23129


SpRY_
GCTGAC

AGCACA

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGCACA



PB14













pU6_A1AT_
GGCTGT
20700
CGATGGTC
21510
CCTTTCTCG
22320
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23130


SpRY_
GCTGAC

AGCAC

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGCAC



PB13













pU6_A1AT_
GGCTGT
20701
CGATGGTC
21511
CCTTTCTCG
22321
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23131


SpRY_
GCTGAC

AGCA

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGCA



PB12













pU6_A1AT_
GGCTGT
20702
CGATGGTC
21512
CCTTTCTCG
22322
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23132


SpRY_
GCTGAC

AGC

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAGC



PB11













pU6_A1AT_
GGCTGT
20703
CGATGGTC
21513
CCTTTCTCG
22323
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23133


SpRY_
GCTGAC

AG

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20704
CGATGGTC

CCTTTCTCG
22324
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23134


SpRY_
GCTGAC

A

T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20705
CGATGGTC

CCTTTCTCG
22325
GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23135


SpRY_
GCTGAC



T

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_7FE_
CA





CCTTTCTCGTCGATGGTC



PB8













pU6_A1AT_
GGCTGT
20706
CGATGGTC
21516
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23136


SpRY_
GCTGAC

AGCACAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGCACAGCC



PB17













pU6_A1AT_
GGCTGT
20707
CGATGGTC
21517
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23137


SpRY_
GCTGAC

AGCACAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGCACAGC



PB16













pU6_A1AT_
GGCTGT
20708
CGATGGTC
21518
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23138


SpRY_
GCTGAC

AGCACAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGCACAG



PB15













pU6_A1AT_
GGCTGT
20709
CGATGGTC
21519
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23139


SpRY_
GCTGAC

AGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGCACA



PB14













pU6_A1AT_
GGCTGT
20710
CGATGGTC
21520
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23140


SpRY_
GCTGAC

AGCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGCAC



PB13













pU6_A1AT_
GGCTGT
20711
CGATGGTC
21521
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23141


SpRY_
GCTGAC

AGCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGCA



PB12













pU6_A1AT_
GGCTGT
20712
CGATGGTC
21522
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23142


SpRY_
GCTGAC

AGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAGC



PB11













pU6_A1AT_
GGCTGT
20713
CGATGGTC
21523
TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23143


SpRY_
GCTGAC

AG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20714
CGATGGTC

TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23144


SpRY_
GCTGAC

A



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20715
CGATGGTC

TTTCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23145


SpRY_
GCTGAC





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_5FE_
CA





TTTCTCGTCGATGGTC



PB8













pU6_A1AT_
GGCTGT
20716
CGATGGTC
21526
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23146


SpRY_
GCTGAC

AGCACAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA

C



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGCACAGCC



PB17













pU6_A1AT_
GGCTGT
20717
CGATGGTC
21527
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23147


SpRY_
GCTGAC

AGCACAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGCACAGC



PB16













pU6_A1AT_
GGCTGT
20718
CGATGGTC
21528
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23148


SpRY_
GCTGAC

AGCACAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGCACAG



PB15













pU6_A1AT_
GGCTGT
20719
CGATGGTC
21529
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23149


SpRY_
GCTGAC

AGCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGCACA



PB14













pU6_A1AT_
GGCTGT
20720
CGATGGTC
21530
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23150


SpRY_
GCTGAC

AGCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGCAC



PB13













pU6_A1AT_
GGCTGT
20721
CGATGGTC
21531
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23151


SpRY_
GCTGAC

AGCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGCA



PB12













pU6_A1AT_
GGCTGT
20722
CGATGGTC
21532
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23152


SpRY_
GCTGAC

AGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAGC



PB11













pU6_A1AT_
GGCTGT
20723
CGATGGTC
21533
TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23153


SpRY_
GCTGAC

AG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCAG



PB10













pU6_A1AT_
GGCTGT
20724
CGATGGTC

TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23154


SpRY_
GCTGAC

A



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTCA



PB9













pU6_A1AT_
GGCTGT
20725
CGATGGTC

TCTCGT

GGCTGTGCTGACCATCGACAGTTTTAGAGCTA
23155


SpRY_
GCTGAC





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED2-
CATCGA





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_3FE_
CA





TCTCGTCGATGGTC



PB8













pU6_A1AT_
AGGCTG
20726
GATGGTCA
21536
ACATGGCCC
22346
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23156


ScaCas9++_
TGCTGA

GCACAGCC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_ED3-
CCATCG

T

CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_30FE_
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB17






CGATGGTCAGCACAGCCT






pU6_A1AT_
AGGCTG
20727
GATGGTCA
21537
ACATGGCCC
22347
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23157


ScaCas9++_
TGCTGA

GCACAGCC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGCACAGCC



PB16













pU6_A1AT_
AGGCTG
20728
GATGGTCA
21538
ACATGGCCC
22348
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23158


ScaCas9++_
TGCTGA

GCACAGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGCACAGC



PB15













pU6_A1AT_
AGGCTG
20729
GATGGTCA
21539
ACATGGCCC
22349
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23159


ScaCas9++_
TGCTGA

GCACAG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGCACAG



PB14













pU6_A1AT_
AGGCTG
20730
GATGGTCA
21540
ACATGGCCC
22350
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23160


ScaCas9++_
TGCTGA

GCACA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGCACA



PB13













pU6_A1AT_
AGGCTG
20731
GATGGTCA
21541
ACATGGCCC
22351
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23161


ScaCas9++_
TGCTGA

GCAC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGCAC



PB12













pU6_A1AT_
AGGCTG
20732
GATGGTCA
21542
ACATGGCCC
22352
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23162


ScaCas9++_
TGCTGA

GCA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGCA



PB11













pU6_A1AT_
AGGCTG
20733
GATGGTCA
21543
ACATGGCCC
22353
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23163


ScaCas9++_
TGCTGA

GC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAGC



PB10













pU6_A1AT_
AGGCTG
20734
GATGGTCA

ACATGGCCC
22354
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23164


ScaCas9++_
TGCTGA

G

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCAG



PB9













pU6_A1AT_
AGGCTG
20735
GATGGTCA

ACATGGCCC
22355
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23165


ScaCas9++_
TGCTGA



CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



TCTcGTC

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_






CGATGGTCA



PB8













pU6_A1AT_
AGGCTG
20736
GATGGTCA
21546
GCCCCAGCA
22356
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23166


ScaCas9++_
TGCTGA

GCACAGCC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T

CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCACAGCCT



PB17













pU6_A1AT_
AGGCTG
20737
GATGGTCA
21547
GCCCCAGCA
22357
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23167


ScaCas9++_
TGCTGA

GCACAGCC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCACAGCC



PB16













pU6_A1AT_
AGGCTG
20738
GATGGTCA
21548
GCCCCAGCA
22358
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23168


ScaCas9++_
TGCTGA

GCACAGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCACAGC



PB15













pU6_A1AT_
AGGCTG
20739
GATGGTCA
21549
GCCCCAGCA
22359
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23169


ScaCas9++_
TGCTGA

GCACAG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCACAG



PB14













pU6_A1AT_
AGGCTG
20740
GATGGTCA
21550
GCCCCAGCA
22360
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23170


ScaCas9++_
TGCTGA

GCACA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCACA



PB13










pU6_A1AT_
AGGCTG
20741
GATGGTCA
21551
GCCCCAGCA
22361
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23171


ScaCas9++_
TGCTGA

GCAC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCAC



PB12













pU6_A1AT_
AGGCTG
20742
GATGGTCA
21552
GCCCCAGCA
22362
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23172


ScaCas9++_
TGCTGA

GCA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGCA



PB11













pU6_A1AT_
AGGCTG
20743
GATGGTCA
21553
GCCCCAGCA
22363
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23173


ScaCas9++_
TGCTGA

GC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAGC



PB10













pU6_A1AT_
AGGCTG
20744
GATGGTCA

GCCCCAGCA
22364
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23174


ScaCas9++_
TGCTGA

G

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCAG



PB9













pU6_A1AT_
AGGCTG
20745
GATGGTCA

GCCCCAGCA
22365
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23175


ScaCas9++_
TGCTGA



GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC



C

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_






GTCA



PB8













pU6_A1AT_
AGGCTG
20746
GATGGTCA
21556
AGCAGCTTC
22366
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23176


ScaCas9++_
TGCTGA

GCACAGCC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T

CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CACAGCCT



PB17













pU6_A1AT_
AGGCTG
20747
GATGGTCA
21557
AGCAGCTTC
22367
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23177


ScaCas9++_
TGCTGA

GCACAGCC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CACAGCC



PB16













pU6_A1AT_
AGGCTG
20748
GATGGTCA
21558
AGCAGCTTC
22368
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23178


ScaCas9++_
TGCTGA

GCACAGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CACAGC



PB15













pU6_A1AT_
AGGCTG
20749
GATGGTCA
21559
AGCAGCTTC
22369
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23179


ScaCas9++_
TGCTGA

GCACAG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CACAG



PB14













pU6_A1AT_
AGGCTG
20750
GATGGTCA
21560
AGCAGCTTC
22370
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23180


ScaCas9++_
TGCTGA

GCACA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CACA



PB13













pU6_A1AT_
AGGCTG
20751
GATGGTCA
21561
AGCAGCTTC
22371
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23181


ScaCas9++_
TGCTGA

GCAC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CAC



PB12













pU6_A1AT_
AGGCTG
20752
GATGGTCA
21562
AGCAGCTTC
22372
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23182


ScaCas9++_
TGCTGA

GCA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






CA



PB11













pU6_A1AT_
AGGCTG
20753
GATGGTCA
21563
AGCAGCTTC
22373
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23183


ScaCas9++_
TGCTGA

GC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_






C



PB10













pU6_A1AT_
AGGCTG
20754
GATGGTCA

AGCAGCTTC
22374
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23184


ScaCas9++_
TGCTGA

G

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_










PB9













pU6_A1AT_
AGGCTG
20755
GATGGTCA

AGCAGCTTC
22375
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23185


ScaCas9++_
TGCTGA



AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG



CTcGTC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCA



_20FE_










PB8













pU6_A1AT_
AGGCTG
20756
GATGGTCA
21566
TTCAGTCCC
22376
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23186


ScaCas9++_
TGCTGA

GCACAGCC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CT



PB17













pU6_A1AT_
AGGCTG
20757
GATGGTCA
21567
TTCAGTCCC
22377
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23187


ScaCas9++_
TGCTGA

GCACAGCC

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






C



PB16













pU6_A1AT_
AGGCTG
20758
GATGGTCA
21568
TTCAGTCCC
22378
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23188


ScaCas9++_
TGCTGA

GCACAGC

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_










PB15













pU6_A1AT_
AGGCTG
20759
GATGGTCA
21569
TTCAGTCCC
22379
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23189


ScaCas9++_
TGCTGA

GCACAG

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



_14FE_










PB14













pU6_A1AT_
AGGCTG
20760
GATGGTCA
21570
TTCAGTCCC
22380
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23190


ScaCas9++_
TGCTGA

GCACA

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCACA



_14FE_










PB13













pU6_A1AT_
AGGCTG
20761
GATGGTCA
21571
TTCAGTCCC
22381
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23191


ScaCas9++_
TGCTGA

GCAC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCAC



_14FE_










PB12













pU6_A1AT_
AGGCTG
20762
GATGGTCA
21572
TTCAGTCCC
22382
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23192


ScaCas9++_
TGCTGA

GCA

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGCA



_14FE_










PB11













pU6_A1AT_
AGGCTG
20763
GATGGTCA
21573
TTCAGTCCC
22383
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23193


ScaCas9++_
TGCTGA

GC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAGC



_14FE_










PB10













pU6_A1AT_
AGGCTG
20764
GATGGTCA

TTCAGTCCC
22384
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23194


ScaCas9++_
TGCTGA

G

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCAG



_14FE_










PB9













pU6_A1AT_
AGGCTG
20765
GATGGTCA

TTCAGTCCC
22385
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23195


ScaCas9++_
TGCTGA



TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTCAGTCCCTTTCTcGTCGATGGTCA



_14FE_










PB8













pU6_A1AT_
AGGCTG
20766
GATGGTCA
21576
AGTCCCTTT
22386
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23196


ScaCas9++_
TGCTGA

GCACAGCC

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCT



_11FE_










PB17













pU6_A1AT_
AGGCTG
20767
GATGGTCA
21577
AGTCCCTTT
22387
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23197


ScaCas9++_
TGCTGA

GCACAGCC

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCACAGCC



_11FE_










PB16













pU6_A1AT_
AGGCTG
20768
GATGGTCA
21578
AGTCCCTTT
22388
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23198


ScaCas9++_
TGCTGA

GCACAGC

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCACAGC



_11FE_










PB15













pU6_A1AT_
AGGCTG
20769
GATGGTCA
21579
AGTCCCTTT
22389
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23199


ScaCas9++_
TGCTGA

GCACAG

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCACAG



_11FE_










PB14













pU6_A1AT_
AGGCTG
20770
GATGGTCA
21580
AGTCCCTTT
22390
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23200


ScaCas9++_
TGCTGA

GCACA

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCACA



_11FE_










PB13













pU6_A1AT_
AGGCTG
20771
GATGGTCA
21581
AGTCCCTTT
22391
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23201


ScaCas9++_
TGCTGA

GCAC

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCAC



_11FE_










PB12













pU6_A1AT_
AGGCTG
20772
GATGGTCA
21582
AGTCCCTTT
22392
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23202


ScaCas9++_
TGCTGA

GCA

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGCA



_11FE_










PB11













pU6_A1AT_
AGGCTG
20773
GATGGTCA
21583
AGTCCCTTT
22393
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23203


ScaCas9++_
TGCTGA

GC

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAGC



_11FE_










PB10













pU6_A1AT_
AGGCTG
20774
GATGGTCA

AGTCCCTTT
22394
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23204


ScaCas9++_
TGCTGA

G

CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCAG



_11FE_










PB9













pU6_A1AT_
AGGCTG
20775
GATGGTCA

AGTCCCTTT
22395
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23205


ScaCas9++_
TGCTGA



CTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





AGTCCCTTTCTcGTCGATGGTCA



_11FE_










PB8













pU6_A1AT_
AGGCTG
20776
GATGGTCA
21586
TCCCTTTCTc
22396
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23206


ScaCas9++_
TGCTGA

GCACAGCC

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCACAGCCT



_9FE_










PB17













pU6_A1AT_
AGGCTG
20777
GATGGTCA
21587
TCCCTTTCTc
22397
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23207


ScaCas9++_
TGCTGA

GCACAGCC

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCACAGCC



_9FE_










PB16













pU6_A1AT_
AGGCTG
20778
GATGGTCA
21588
TCCCTTTCTc
22398
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23208


ScaCas9++_
TGCTGA

GCACAGC

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCACAGC



_9FE_










PB15













pU6_A1AT_
AGGCTG
20779
GATGGTCA
21589
TCCCTTTCTc
22399
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23209


ScaCas9++_
TGCTGA

GCACAG

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCACAG



_9FE_










PB14













pU6_A1AT_
AGGCTG
20780
GATGGTCA
21590
TCCCTTTCTc
22400
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23210


ScaCas9++_
TGCTGA

GCACA

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCACA



_9FE_










PB13













pU6_A1AT_
AGGCTG
20781
GATGGTCA
21591
TCCCTTTCTc
22401
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23211


ScaCas9++_
TGCTGA

GCAC

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCAC



_9FE_










PB12













pU6_A1AT_
AGGCTG
20782
GATGGTCA
21592
TCCCTTTCTC
22402
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23212


ScaCas9++_
TGCTGA

GCA

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGCA



_9FE_










PB11













pU6_A1AT_
AGGCTG
20783
GATGGTCA
21593
TCCCTTTCTc
22403
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23213


ScaCas9++_
TGCTGA

GC

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAGC



_9FE_










PB10













pU6_A1AT_
AGGCTG
20784
GATGGTCA

TCCCTTTCTc
22404
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23214


ScaCas9++_
TGCTGA

G

GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCAG



_9FE_










PB9













pU6_A1AT_
AGGCTG
20785
GATGGTCA

TCCCTTTCTC
22405
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23215


ScaCas9++_
TGCTGA



GTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCCCTTTCTcGTCGATGGTCA



_9FE_










PB8













pU6_A1AT_
AGGCTG
20786
GATGGTCA
21596
CCTTTCTcGT
22406
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23216


ScaCas9++_
TGCTGA

GCACAGCC

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCACAGCCT



_7FE_










PB17













pU6_A1AT_
AGGCTG
20787
GATGGTCA
21597
CCTTTCTcGT
22407
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23217


ScaCas9++_
TGCTGA

GCACAGCC

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCACAGCC



_7FE_










PB16













pU6_A1AT_
AGGCTG
20788
GATGGTCA
21598
CCTTTCTcGT
22408
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23218


ScaCas9++_
TGCTGA

GCACAGC

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCACAGC



_7FE_










PB15













pU6_A1AT_
AGGCTG
20789
GATGGTCA
21599
CCTTTCTCGT
22409
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23219


ScaCas9++_
TGCTGA

GCACAG

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCACAG



_7FE_










PB14













pU6_A1AT_
AGGCTG
20790
GATGGTCA
21600
CCTTTCTcGT
22410
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23220


ScaCas9++_
TGCTGA

GCACA

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCACA



_7FE_










PB13













pU6_A1AT_
AGGCTG
20791
GATGGTCA
21601
CCTTTCTcGT
22411
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23221


ScaCas9++_
TGCTGA

GCAC

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCAC



_7FE_










PB12













pU6_A1AT_
AGGCTG
20792
GATGGTCA
21602
CCTTTCTcGT
22412
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23222


ScaCas9++_
TGCTGA

GCA

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGCA



_7FE_










PB11













pU6_A1AT_
AGGCTG
20793
GATGGTCA
21603
CCTTTCTcGT
22413
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23223


ScaCas9++_
TGCTGA

GC

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAGC



_7FE_










PB10













pU6_A1AT_
AGGCTG
20794
GATGGTCA

CCTTTCTcGT
22414
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23224


ScaCas9++_
TGCTGA

G

C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCAG



_7FE_










PB9













pU6_A1AT_
AGGCTG
20795
GATGGTCA

CCTTTCTcGT
22415
AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23225


ScaCas9++_
TGCTGA



C

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





CCTTTCTcGTCGATGGTCA



_7FE_










PB8













pU6_A1AT_
AGGCTG
20796
GATGGTCA
21606
TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23226


ScaCas9++_
TGCTGA

GCACAGCC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTCGTCGATGGTCAGCACAGCCT



_5FE_










PB17













pU6_A1AT_
AGGCTG
20797
GATGGTCA
21607
TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23227


ScaCas9++_
TGCTGA

GCACAGCC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTCGTCGATGGTCAGCACAGCC



_5FE_










PB16













pU6_A1AT_
AGGCTG
20798
GATGGTCA
21608
TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23228


ScaCas9++_
TGCTGA

GCACAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTCGTCGATGGTCAGCACAGC



_5FE_










PB15













pU6_A1AT_
AGGCTG
20799
GATGGTCA
21609
TTTCTCGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23229


ScaCas9++_
TGCTGA

GCACAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTCGTCGATGGTCAGCACAG



_5FE_










PB14













pU6_A1AT_
AGGCTG
20800
GATGGTCA
21610
TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23230


ScaCas9++_
TGCTGA

GCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTcGTCGATGGTCAGCACA



_5FE_










PB13













pU6_A1AT_
AGGCTG
20801
GATGGTCA
21611
TTTCTCGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23231


ScaCas9++_
TGCTGA

GCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTCGTCGATGGTCAGCAC



_5FE_










PB12













pU6_A1AT_
AGGCTG
20802
GATGGTCA
21612
TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23232


ScaCas9++_
TGCTGA

GCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_E
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



D3-
AC





TTTCTCGTCGATGGTCAGCA



_5FE_










PB11













pU6_A1AT_
AGGCTG
20803
GATGGTCA
21613
TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23233


ScaCas9++_
TGCTGA

GC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTcGTCGATGGTCAGC



_5FE_










PB10













pU6_A1AT_
AGGCTG
20804
GATGGTCA

TTTCTCGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23234


ScaCas9++_
TGCTGA

G



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTcGTCGATGGTCAG



_5FE_










PB9













pU6_A1AT_
AGGCTG
20805
GATGGTCA

TTTCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23235


ScaCas9++_
TGCTGA





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TTTCTCGTCGATGGTCA



_5FE_










PB8













pU6_A1AT_
AGGCTG
20806
GATGGTCA
21616
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23236


ScaCas9++_
TGCTGA

GCACAGCC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG

T



ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCACAGCCT



_3FE_










PB17













pU6_A1AT_
AGGCTG
20807
GATGGTCA
21617
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23237


ScaCas9++_
TGCTGA

GCACAGCC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCACAGCC



_3FE_










PB16













pU6_A1AT_
AGGCTG
20808
GATGGTCA
21618
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23238


ScaCas9++_
TGCTGA

GCACAGC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCACAGC



_3FE_










PB15













pU6_A1AT_
AGGCTG
20809
GATGGTCA
21619
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23239


ScaCas9++_
TGCTGA

GCACAG



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCACAG



_3FE_










PB14













pU6_A1AT_
AGGCTG
20810
GATGGTCA
21620
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23240


ScaCas9++_
TGCTGA

GCACA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCACA



_3FE_










PB13













pU6_A1AT_
AGGCTG
20811
GATGGTCA
21621
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23241


ScaCas9++_
TGCTGA

GCAC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCAC



_3FE_










PB12













pU6_A1AT_
AGGCTG
20812
GATGGTCA
21622
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23242


ScaCas9++_
TGCTGA

GCA



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGCA



_3FE_










PB11













pU6_A1AT_
AGGCTG
20813
GATGGTCA
21623
TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23243


ScaCas9++_
TGCTGA

GC



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAGC



_3FE_










PB10













pU6_A1AT_
AGGCTG
20814
GATGGTCA

TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23244


ScaCas9++_
TGCTGA

G



GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCAG



_3FE_










PB9













pU6_A1AT_
AGGCTG
20815
GATGGTCA

TCTcGTC

AGGCTGTGCTGACCATCGACGTTTTAGAGCTA
23245


ScaCas9++_
TGCTGA





GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



SpRY_
CCATCG





ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED3-
AC





TCTcGTCGATGGTCA



_3FE_










PB8













pU6_A1AT_
AAGGCT
20816
ATGGTCAG
21626
ACATGGCCC
22436
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23246


St1_
GTGCTG

CACAGCCT

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T

CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB17






TTCTcGTCGATGGTCAGCACAGCCTT






pU6_A1AT_
AAGGCT
20817
ATGGTCAG
21627
ACATGGCCC
22437
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23247


St1_
GTGCTG

CACAGCCT

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB16






TTCTcGTCGATGGTCAGCACAGCCT






pU6_A1AT_
AAGGCT
20818
ATGGTCAG
21628
ACATGGCCC
22438
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23248


St1_
GTGCTG

CACAGCC

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB15






TTCTcGTCGATGGTCAGCACAGCC






pU6_A1AT_
AAGGCT
20819
ATGGTCAG
21629
ACATGGCCC
22439
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23249


St1_
GTGCTG

CACAGC

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB14






TTCTcGTCGATGGTCAGCACAGC






pU6_A1AT_
AAGGCT
20820
ATGGTCAG
21630
ACATGGCCC
22440
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23250


St1_
GTGCTG

CACAG

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB13






TTCTcGTCGATGGTCAGCACAG






pU6_A1AT_
AAGGCT
20821
ATGGTCAG
21631
ACATGGCCC
22441
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23251


St1_
GTGCTG

CACA

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB12






TTCTcGTCGATGGTCAGCACA






pU6_A1AT_
AAGGCT
20822
ATGGTCAG
21632
ACATGGCCC
22442
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23252


St1_
GTGCTG

CAC

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB11






TTCTcGTCGATGGTCAGCAC






pU6_A1AT_
AAGGCT
20823
ATGGTCAG
21633
ACATGGCCC
22443
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23253


St1_
GTGCTG

CA

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30FE_
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



PB10






TTCTcGTCGATGGTCAGCA






pU6_A1
AAGGCT
20824
ATGGTCAG

ACATGGCCC
22444
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23254


AT_St1
GTGCTG

C

CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



G_30F
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



E PB9






TTCTcGTCGATGGTCAGC






pU6_A1
AAGGCT
20825
ATGGTCAG

ACATGGCCC
22445
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23255


AT_St1
GTGCTG



CAGCAGCTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CAGTCCCTT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_30F
GA



TCTcGTCG

GTGTTTTACATGGCCCCAGCAGCTTCAGTCCCT



E PB8






TTCTcGTCGATGGTCAG






pU6_A1AT_
AAGGCT
20826
ATGGTCAG
21636
GCCCCAGCA
22446
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23256


St1_
GTGCTG

CACAGCCT

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T

CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB17






GTCGATGGTCAGCACAGCCTT






pU6_A1AT_
AAGGCT
20827
ATGGTCAG
21637
GCCCCAGCA
22447
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23257


St1_
GTGCTG

CACAGCCT

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB16






GTCGATGGTCAGCACAGCCT






pU6_A1AT_
AAGGCT
20828
ATGGTCAG
21638
GCCCCAGCA
22448
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23258


St1_
GTGCTG

CACAGCC

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTCGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB15






GTCGATGGTCAGCACAGCC






pU6_A1
AAGGCT
20829
ATGGTCAG
21639
GCCCCAGCA
22449
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23259


AT_St1
GTGCTG

CACAGC

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25F
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



E_PB14






GTCGATGGTCAGCACAGC






pU6_A1AT_
AAGGCT
20830
ATGGTCAG
21640
GCCCCAGCA
22450
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23260


_St1_
GTGCTG

CACAG

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB13






GTCGATGGTCAGCACAG






pU6_A1
AAGGCT
20831
ATGGTCAG
21641
GCCCCAGCA
22451
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23261


AT_St1
GTGCTG

CACA

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTCGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB12






GTCGATGGTCAGCACA






pU6_A1AT_
AAGGCT
20832
ATGGTCAG
21642
GCCCCAGCA
22452
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23262


St1_
GTGCTG

CAC

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB11






GTCGATGGTCAGCAC






pU6_A1
AAGGCT
20833
ATGGTCAG
21643
GCCCCAGCA
22453
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23263


AT_St1
GTGCTG

CA

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25F
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



E_PB10






GTCGATGGTCAGCA






pU6_A1AT_
AAGGCT
20834
ATGGTCAG

GCCCCAGCA
22454
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23264


St1_
GTGCTG

C

GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB9






GTCGATGGTCAGC






pU6_A1AT_
AAGGCT
20835
ATGGTCAG

GCCCCAGCA
22455
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23265


St1_
GTGCTG



GCTTCAGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CCTTTCTcGT

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_25FE_
GA



CG

GTGTTTTGCCCCAGCAGCTTCAGTCCCTTTCTc



PB8






GTCGATGGTCAG






pU6_A1AT_
AAGGCT
20836
ATGGTCAG
21646
AGCAGCTTC
22456
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23266


St1_
GTGCTG

CACAGCCT

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T

CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB17






GGTCAGCACAGCCTT






pU6_A1AT_
AAGGCT
20837
ATGGTCAG
21647
AGCAGCTTC
22457
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23267


St1_
GTGCTG

CACAGCCT

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB16






GGTCAGCACAGCCT






pU6_A1AT_
AAGGCT
20838
ATGGTCAG
21648
AGCAGCTTC
22458
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23268


St1_
GTGCTG

CACAGCC

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB15






GGTCAGCACAGCC






pU6_A1AT_
AAGGCT
20839
ATGGTCAG
21649
AGCAGCTTC
22459
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23269


St1_
GTGCTG

CACAGC

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB14






GGTCAGCACAGC






pU6_A1AT_
AAGGCT
20840
ATGGTCAG
21650
AGCAGCTTC
22460
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23270


St1_
GTGCTG

CACAG

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB13






GGTCAGCACAG






pU6_A1AT_
AAGGCT
20841
ATGGTCAG
21651
AGCAGCTTC
22461
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23271


St1_
GTGCTG

CACA

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB12






GGTCAGCACA






pU6_A1AT_
AAGGCT
20842
ATGGTCAG
21652
AGCAGCTTC
22462
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23272


St1_
GTGCTG

CAC

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB11






GGTCAGCAC






pU6_A1AT_
AAGGCT
20843
ATGGTCAG
21653
AGCAGCTTC
22463
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23273


St1_
GTGCTG

CA

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB10






GGTCAGCA






pU6_A1AT_
AAGGCT
20844
ATGGTCAG

AGCAGCTTC
22464
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23274


St1_
GTGCTG

C

AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB9






GGTCAGC






pU6_A1AT_
AAGGCT
20845
ATGGTCAG

AGCAGCTTC
22465
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23275


St1_
GTGCTG



AGTCCCTTT

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



CTcGTCG

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_20FE_
GA





GTGTTTTAGCAGCTTCAGTCCCTTTCTcGTCGAT



PB8






GGTCAG






pU6_A1AT_
AAGGCT
20846
ATGGTCAG
21656
TTCAGTCCC
22466
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23276


St1_
GTGCTG

CACAGCCT

TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T

G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB17






GCACAGCCTT






pU6_A1AT_
AAGGCT
20847
ATGGTCAG
21657
TTCAGTCCC
22467
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23277


St1_
GTGCTG

CACAGCCT

TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB16






GCACAGCCT






pU6_A1AT_
AAGGCT
20848
ATGGTCAG
21658
TTCAGTCCC
22468
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23278


St1_
GTGCTG

CACAGCC

TTTCTcGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB15






GCACAGCC






pU6_A1AT_
AAGGCT
20849
ATGGTCAG
21659
TTCAGTCCC
22469
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23279


St1_
GTGCTG

CACAGC

TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB14






GCACAGC






pU6_A1AT_
AAGGCT
20850
ATGGTCAG
21660
TTCAGTCCC
22470
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23280


St1_
GTGCTG

CACAG

TTTCTcGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB13






GCACAG






pU6_A1AT_
AAGGCT
20851
ATGGTCAG
21661
TTCAGTCCC
22471
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23281


St1
GTGCTG

CACA

TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB12






GCACA






pU6_A1AT_
AAGGCT
20852
ATGGTCAG
21662
TTCAGTCCC
22472
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23282


St1_
GTGCTG

CAC

TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB11






GCAC






pU6_A1AT_
AAGGCT
20853
ATGGTCAG
21663
TTCAGTCCC
22473
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23283


St1_
GTGCTG

CA

TTTCTcGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB10






GCA






pU6_A1AT_
AAGGCT
20854
ATGGTCAG

TTCAGTCCC
22474
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23284


St1_
GTGCTG

C

TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB9






GC






pU6_A1AT_
AAGGCT
20855
ATGGTCAG

TTCAGTCCC
22475
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23285


St1_
GTGCTG



TTTCTCGTC

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC



G

CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_14FE_
GA





GTGTTTTTTCAGTCCCTTTCTcGTCGATGGTCA



PB8






G






pU6_A1AT_
AAGGCT
20856
ATGGTCAG
21666
AGTCCCTTT
22476
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23286


St1_
GTGCTG

CACAGCCT

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T



CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB17






CAGCCTT






pU6_A1AT_
AAGGCT
20857
ATGGTCAG
21667
AGTCCCTTT
22477
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23287


St1_
GTGCTG

CACAGCCT

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB16






CAGCCT






pU6_A1AT_
AAGGCT
20858
ATGGTCAG
21668
AGTCCCTTT
22478
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23288


St1_
GTGCTG

CACAGCC

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB15






CAGCC






pU6_A1AT_
AAGGCT
20859
ATGGTCAG
21669
AGTCCCTTT
22479
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23289


St1_
GTGCTG

CACAGC

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB14






CAGC






pU6_A1AT_
AAGGCT
20860
ATGGTCAG
21670
AGTCCCTTT
22480
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23290


St1_
GTGCTG

CACAG

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB13






CAG






pU6_A1AT_
AAGGCT
20861
ATGGTCAG
21671
AGTCCCTTT
22481
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23291


St1_
GTGCTG

CACA

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB12






CA






pU6_A1AT_
AAGGCT
20862
ATGGTCAG
21672
AGTCCCTTT
22482
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23292


St1_
GTGCTG

CAC

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB11






C






pU6_A1AT_
AAGGCT
20863
ATGGTCAG
21673
AGTCCCTTT
22483
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23293


St1_
GTGCTG

CA

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGCA



PB10













pU6_A1AT_
AAGGCT
20864
ATGGTCAG

AGTCCCTTT
22484
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23294


St1_
GTGCTG

C

CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAGC



PB9













pU6_A1AT_
AAGGCT
20865
ATGGTCAG

AGTCCCTTT
22485
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23295


St1_
GTGCTG



CTcGTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_11FE_
GA





GTGTTTTAGTCCCTTTCTcGTCGATGGTCAG



PB8













pU6_A1AT_
AAGGCT
20866
ATGGTCAG
21676
TCCCTTTCTc
22486
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23296


St1_
GTGCTG

CACAGCCT

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T



CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCACA



PB17






GCCTT






pU6_A1AT_
AAGGCT
20867
ATGGTCAG
21677
TCCCTTTCTc
22487
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23297


St1_
GTGCTG

CACAGCCT

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCACA



PB16






GCCT






pU6_A1AT_
AAGGCT
20868
ATGGTCAG
21678
TCCCTTTCTc
22488
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23298


St1_
GTGCTG

CACAGCC

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCACA



PB15






GCC






pU6_A1AT_
AAGGCT
20869
ATGGTCAG
21679
TCCCTTTCTc
22489
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23299


St1_
GTGCTG

CACAGC

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCACA



PB14






GC






pU6_A1AT_
AAGGCT
20870
ATGGTCAG
21680
TCCCTTTCTc
22490
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23300


St1_
GTGCTG

CACAG

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCACA



PB13






G






pU6_A1AT_
AAGGCT
20871
ATGGTCAG
21681
TCCCTTTCTc
22491
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23301


St1_
GTGCTG

CACA

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCACA



PB12













pU6_A1AT_
AAGGCT
20872
ATGGTCAG
21682
TCCCTTTCTc
22492
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23302


St1_
GTGCTG

CAC

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCAC



PB11













pU6_A1AT_
AAGGCT
20873
ATGGTCAG
21683
TCCCTTTCTc
22493
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23303


St1_
GTGCTG

CA

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGCA



PB10













pU6_A1AT_
AAGGCT
20874
ATGGTCAG

TCCCTTTCTc
22494
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23304


St1_
GTGCTG

C

GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAGC



PB9













pU6_A1AT_
AAGGCT
20875
ATGGTCAG

TCCCTTTCTc
22495
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23305


St1_
GTGCTG



GTCG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_9FE_
GA





GTGTTTTTCCCTTTCTcGTCGATGGTCAG



PB8













pU6_A1AT_
AAGGCT
20876
ATGGTCAG
21686
CCTTTCTcGT
22496
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23306


St1_
GTGCTG

CACAGCCT

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T



CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCACAGC



PB17






CTT






pU6_A1AT_
AAGGCT
20877
ATGGTCAG
21687
CCTTTCTcGT
22497
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23307


St1_
GTGCTG

CACAGCCT

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCACAGC



PB16






CT






pU6_A1AT_
AAGGCT
20878
ATGGTCAG
21688
CCTTTCTcGT
22498
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23308


St1_
GTGCTG

CACAGCC

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCACAGC



PB15






C






pU6_A1AT_
AAGGCT
20879
ATGGTCAG
21689
CCTTTCTcGT
22499
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23309


St1_
GTGCTG

CACAGC

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCACAGC



PB14













pU6_A1AT_
AAGGCT
20880
ATGGTCAG
21690
CCTTTCTcGT
22500
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23310


St1_
GTGCTG

CACAG

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCACAG



PB13













pU6_A1AT_
AAGGCT
20881
ATGGTCAG
21691
CCTTTCTcGT
22501
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23311


St1_
GTGCTG

CACA

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCACA



PB12













pU6_A1AT_
AAGGCT
20882
ATGGTCAG
21692
CCTTTCTcGT
22502
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23312


St1_
GTGCTG

CAC

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCAC



PB11













pU6_A1AT_
AAGGCT
20883
ATGGTCAG
21693
CCTTTCTcGT
22503
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23313


St1_
GTGCTG

CA

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGCA



PB10













pU6_A1AT_
AAGGCT
20884
ATGGTCAG

CCTTTCTcGT
22504
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23314


St1_
GTGCTG

C

CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAGC



PB9













pU6_A1AT_
AAGGCT
20885
ATGGTCAG

CCTTTCTcGT
22505
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23315


St1_
GTGCTG



CG

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_7FE_
GA





GTGTTTTCCTTTCTcGTCGATGGTCAG



PB8













pU6_A1AT_
AAGGCT
20886
ATGGTCAG
21696
TTTCTcGTC
22506
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23316


St1_
GTGCTG

CACAGCCT

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T



CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCACAGCCT



PB17






T






pU6_A1AT_
AAGGCT
20887
ATGGTCAG
21697
TTTCTcGTC
22507
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23317


St1_
GTGCTG

CACAGCCT

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCACAGCCT



PB16













pU6_A1AT_
AAGGCT
20888
ATGGTCAG
21698
TTTCTcGTC
22508
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23318


St1_
GTGCTG

CACAGCC

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCACAGCC



PB15













pU6_A1AT_
AAGGCT
20889
ATGGTCAG
21699
TTTCTcGTC
22509
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23319


St1_
GTGCTG

CACAGC

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCACAGC



PB14













pU6_A1AT_
AAGGCT
20890
ATGGTCAG
21700
TTTCTcGTC
22510
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23320


St1_
GTGCTG

CACAG

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCACAG



PB13













pU6_A1AT_
AAGGCT
20891
ATGGTCAG
21701
TTTCTcGTC
22511
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23321


St1_
GTGCTG

CACA

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCACA



PB12













pU6_A1AT_
AAGGCT
20892
ATGGTCAG
21702
TTTCTcGTC
22512
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23322


St1_
GTGCTG

CAC

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCAC



PB11













pU6_A1AT_
AAGGCT
20893
ATGGTCAG
21703
TTTCTcGTC
22513
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23323


St1_
GTGCTG

CA

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGCA



PB10













pU6_A1AT_
AAGGCT
20894
ATGGTCAG

TTTCTcGTC
22514
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23324


St1_
GTGCTG

C

G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAGC



PB9













pU6_A1AT_
AAGGCT
20895
ATGGTCAG

TTTCTCGTC
22515
AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23325


St1_
GTGCTG



G

GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_5FE_
GA





GTGTTTTTTTCTcGTCGATGGTCAG



PB8













pU6_A1AT_
AAGGCT
20896
ATGGTCAG
21706
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23326


St1_
GTGCTG

CACAGCCT



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC

T



CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCACAGCCTT



PB17













pU6_A1AT_
AAGGCT
20897
ATGGTCAG
21707
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23327


St1_
GTGCTG

CACAGCCT



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCACAGCCT



PB16













pU6_A1AT_
AAGGCT
20898
ATGGTCAG
21708
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23328


St1_
GTGCTG

CACAGCC



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCACAGCC



PB15













pU6_A1AT_
AAGGCT
20899
ATGGTCAG
21709
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23329


St1_
GTGCTG

CACAGC



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCACAGC



PB14













pU6_A1AT_
AAGGCT
20900
ATGGTCAG
21710
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23330


St1_
GTGCTG

CACAG



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCACAG



PB13













pU6_A1AT_
AAGGCT
20901
ATGGTCAG
21711
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23331


St1_
GTGCTG

CACA



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCACA



PB12













pU6_A1AT_
AAGGCT
20902
ATGGTCAG
21712
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23332


St1_
GTGCTG

CAC



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCAC



PB11













pU6_A1AT_
AAGGCT
20903
ATGGTCAG
21713
TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23333


St1_
GTGCTG

CA



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGCA



PB10













pU6_A1AT_
AAGGCT
20904
ATGGTCAG

TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23334


St1_
GTGCTG

C



GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAGC



PB9













pU6_A1AT_
AAGGCT
20905
ATGGTCAG

TCTcGTCG

AAGGCTGTGCTGACCATCGAGTCTTTGTACTCT
23335


St1_
GTGCTG





GGTACCAGAAGCTACAAAGATAAGGCTTCATG



ED4-
ACCATC





CCGAAATCAACACCCTGTCATTTTATGGCAGG



_G_3FE_
GA





GTGTTTTTCTcGTCGATGGTCAG



PB8













pU6_A1AT_
TAAAAA
20906
CTGCTGGG
21716
GCCGTGCAT
22526
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
203336


Nme2_
CATGGC

GCCATGTT

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB17




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCCATGTTT






pU6_A1AT_
TAAAAA
20907
CTGCTGGG
21717
GCCGTGCAT
22527
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
203337


Nme2_
CATGGC

GCCATGTT

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB16




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCCATGTT






pU6_A1AT_
TAAAAA
20908
CTGCTGGG
21718
GCCGTGCAT
22528
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
203338


Nme2_
CATGGC

GCCATGT

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB15




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20909
CTGCTGGG
21719
GCCGTGCAT
22529
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
203339


Nme2_
CATGGC

GCCATG

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB14




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20910
CTGCTGGG
21720
GCCGTGCAT
22530
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
203340


Nme2_
CATGGC

GCCAT

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB13




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20911
CTGCTGGG
21721
GCCGTGCAT
22531
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23341


Nme2_
CATGGC

GCCA

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB12




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20912
CTGCTGGG
21722
GCCGTGCAT
22532
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23342


Nme2_
CATGGC

GCC

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB11




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20913
CTGCTGGG
21723
GCCGTGCAT
22533
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23343


Nme2_
CATGGC

GC

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB10




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20914
CTGCTGGG

GCCGTGCAT
22534
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23344


Nme2_
CATGGC

G

AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB9




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20915
CTGCTGGG

GCCGTGCAT
22535
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23345


Nme2_
CATGGC



AAGGCTGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GCTGACCAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_30FE_
AGCTT



CGACgAGAA

TTCTGCTTTAAGGGGCATCGTTGCCGTGCATAA



PB8




AGGGACTG

GGCTGTGCTGACCATCGACgAGAAAGGGACTG








AAG

AAGCTGCTGGG






pU6_A1AT_
TAAAAA
20916
CTGCTGGG
21726
GCATAAGG
22536
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23346


Nme2_
CATGGC

GCCATGTT

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB17




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGCCATGTTT






pU6_A1AT_
TAAAAA
20917
CTGCTGGG
21727
GCATAAGG
22537
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23347


Nme2_
CATGGC

GCCATGTT

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG



16






CTGCTGGGGCCATGTT






pU6_A1AT_
TAAAAA
20918
CTGCTGGG
21728
GCATAAGG
22538
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23348


Nme2_
CATGGC

GCCATGT

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB15




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20919
CTGCTGGG
21729
GCATAAGG
22539
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23349


Nme2_
CATGGC

GCCATG

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB14




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20920
CTGCTGGG
21730
GCATAAGG
22540
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23350


Nme2_
CATGGC

GCCAT

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB13




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20921
CTGCTGGG
21731
GCATAAGG
22541
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23351


Nme2_
CATGGC

GCCA

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB12




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20922
CTGCTGGG
21732
GCATAAGG
22542
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23352


Nme2_
CATGGC

GCC

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB11




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGCC






pU6_A1AT_
TAAAAA
20923
CTGCTGGG
21733
GCATAAGG
22543
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23353


Nme2_
CATGGC

GC

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB10




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGGC






pU6_A1AT_
TAAAAA
20924
CTGCTGGG

GCATAAGG
22544
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23354


Nme2_
CATGGC

G

CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB9




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGGG






pU6_A1AT_
TAAAAA
20925
CTGCTGGG

GCATAAGG
22545
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23355


Nme2_
CATGGC



CTGTGCTGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



CCATCGACg

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_25FE_
AGCTT



AGAAAGGG

TTCTGCTTTAAGGGGCATCGTTGCATAAGGCT



PB8




ACTGAAG

GTGCTGACCATCGACgAGAAAGGGACTGAAG










CTGCTGGG






pU6_A1AT_
TAAAAA
20926
CTGCTGGG
21736
AGGCTGTGC
22546
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23356


Nme2_
CATGGC

GCCATGTT

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB17




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCCATGTTT






pU6_A1AT_
TAAAAA
20927
CTGCTGGG
21737
AGGCTGTGC
22547
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23357


Nme2_
CATGGC

GCCATGTT

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB16




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCCATGTT






pU6_A1AT_
TAAAAA
20928
CTGCTGGG
21738
AGGCTGTGC
22548
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23358


Nme2_
CATGGC

GCCATGT

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB15




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCCATGT






pU6_A1AT_
TAAAAA
20929
CTGCTGGG
21739
AGGCTGTGC
22549
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23359


Nme2_
CATGGC

GCCATG

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB14




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCCATG






pU6_A1AT_
TAAAAA
20930
CTGCTGGG
21740
AGGCTGTGC
22550
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23360


Nme2_
CATGGC

GCCAT

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB13




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCCAT






pU6_A1AT_
TAAAAA
20931
CTGCTGGG
21741
AGGCTGTGC
22551
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23361


Nme2_
CATGGC

GCCA

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB12




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCCA






pU6_A1AT_
TAAAAA
20932
CTGCTGGG
21742
AGGCTGTGC
22552
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23362


Nme2_
CATGGC

GCC

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB11




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGCC






pU6_A1AT_
TAAAAA
20933
CTGCTGGG
21743
AGGCTGTGC
22553
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23363


Nme2_
CATGGC

GC

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB10




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGGC






pU6_A1AT_
TAAAAA
20934
CTGCTGGG

AGGCTGTGC
22554
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23364


Nme2_
CATGGC

G

TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB9




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGGG






pU6_A1AT_
TAAAAA
20935
CTGCTGGG

AGGCTGTGC
22555
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23365


Nme2_
CATGGC



TGACCATCG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



ACgAGAAA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_20FE_
AGCTT



GGGACTGA

TTCTGCTTTAAGGGGCATCGTTAGGCTGTGCT



PB8




AG

GACCATCGACgAGAAAGGGACTGAAGCTGCT










GGG






pU6_A1AT_
TAAAAA
20936
CTGCTGGG
21746
TGCTGACCA
22556
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23366


Nme2_
CATGGC

GCCATGTT

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB17






GACgAGAAAGGGACTGAAGCTGCTGGGGCCA










TGTTT






pU6_A1AT_
TAAAAA
20937
CTGCTGGG
21747
TGCTGACCA
22557
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23367


Nme2_
CATGGC

GCCATGTT

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB16






GACgAGAAAGGGACTGAAGCTGCTGGGGCCA










TGTT






pU6_A1AT_
TAAAAA
20938
CTGCTGGG
21748
TGCTGACCA
22558
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23368


Nme2_
CATGGC

GCCATGT

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB15






GACgAGAAAGGGACTGAAGCTGCTGGGGCCA










TGT






pU6_A1AT_
TAAAAA
20939
CTGCTGGG
21749
TGCTGACCA
22559
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23369


Nme2_
CATGGC

GCCATG

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB14






GACgAGAAAGGGACTGAAGCTGCTGGGGCCA










TG






pU6_A1AT_
TAAAAA
20940
CTGCTGGG
21750
TGCTGACCA
22560
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23370


Nme2_
CATGGC

GCCAT

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB13






GACgAGAAAGGGACTGAAGCTGCTGGGGCCA










T






pU6_A1AT_
TAAAAA
20941
CTGCTGGG
21751
TGCTGACCA
22561
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23371


Nme2_
CATGGC

GCCA

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB12






GACgAGAAAGGGACTGAAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20942
CTGCTGGG
21752
TGCTGACCA
22562
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23372


Nme2_
CATGGC

GCC

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB11






GACgAGAAAGGGACTGAAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20943
CTGCTGGG
21753
TGCTGACCA
22563
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23373


Nme2_
CATGGC

GC

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB10






GACgAGAAAGGGACTGAAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20944
CTGCTGGG

TGCTGACCA
22564
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23374


Nme2_
CATGGC

G

TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB9






GACgAGAAAGGGACTGAAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20945
CTGCTGGG

TGCTGACCA
22565
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23375


Nme2_
CATGGC



TCGACgAGA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAGGGACT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_14FE_
AGCTT



GAAG

TTCTGCTTTAAGGGGCATCGTTTGCTGACCATC



PB8






GACgAGAAAGGGACTGAAGCTGCTGGG






pU6_A1AT_
TAAAAA
20946
CTGCTGGG
21756
TGACCATCG
22566
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23376


Nme2_
CATGGC

GCCATGTT

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB17






gAGAAAGGGACTGAAGCTGCTGGGGCCATGT










TT






pU6_A1AT_
TAAAAA
20947
CTGCTGGG
21757
TGACCATCG
22567
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23377


Nme2_
CATGGC

GCCATGTT

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB16






gAGAAAGGGACTGAAGCTGCTGGGGCCATGT










T






pU6_A1AT_
TAAAAA
20948
CTGCTGGG
21758
TGACCATCG
22568
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23378


Nme2_
CATGGC

GCCATGT

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB15






gAGAAAGGGACTGAAGCTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20949
CTGCTGGG
21759
TGACCATCG
22569
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23379


Nme2_
CATGGC

GCCATG

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB14






gAGAAAGGGACTGAAGCTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20950
CTGCTGGG
21760
TGACCATCG
22570
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23380


Nme2_
CATGGC

GCCAT

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB13






gAGAAAGGGACTGAAGCTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20951
CTGCTGGG
21761
TGACCATCG
22571
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23381


Nme2_
CATGGC

GCCA

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB12






gAGAAAGGGACTGAAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20952
CTGCTGGG
21762
TGACCATCG
22572
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23382


Nme2_
CATGGC

GCC

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB11






gAGAAAGGGACTGAAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20953
CTGCTGGG
21763
TGACCATCG
22573
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23383


Nme2_
CATGGC

GC

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB10






gAGAAAGGGACTGAAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20954
CTGCTGGG

TGACCATCG
22574
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23384


Nme2_
CATGGC

G

ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB9






gAGAAAGGGACTGAAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20955
CTGCTGGG

TGACCATCG
22575
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23385


Nme2_
CATGGC



ACgAGAAA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GGGACTGA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_11FE_
AGCTT



AG

TTCTGCTTTAAGGGGCATCGTTTGACCATCGAC



PB8






gAGAAAGGGACTGAAGCTGCTGGG






pU6_A1AT_
TAAAAA
20956
CTGCTGGG
21766
ACCATCGAC
22576
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23386


Nme2_
CATGGC

GCCATGTT

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB17






GAAAGGGACTGAAGCTGCTGGGGCCATGTTT






pU6_A1AT_
TAAAAA
20957
CTGCTGGG
21767
ACCATCGAC
22577
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23387


Nme2_
CATGGC

GCCATGTT

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB16






GAAAGGGACTGAAGCTGCTGGGGCCATGTT






pU6_A1AT_
TAAAAA
20958
CTGCTGGG
21768
ACCATCGAC
22578
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23388


Nme2_
CATGGC

GCCATGT

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB15






GAAAGGGACTGAAGCTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20959
CTGCTGGG
21769
ACCATCGAC
22579
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23389


Nme2_
CATGGC

GCCATG

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB14






GAAAGGGACTGAAGCTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20960
CTGCTGGG
21770
ACCATCGAC
22580
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23390


Nme2_
CATGGC

GCCAT

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB13






GAAAGGGACTGAAGCTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20961
CTGCTGGG
21771
ACCATCGAC
22581
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23391


Nme2_
CATGGC

GCCA

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB12






GAAAGGGACTGAAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20962
CTGCTGGG
21772
ACCATCGAC
22582
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23392


Nme2_
CATGGC

GCC

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB11






GAAAGGGACTGAAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20963
CTGCTGGG
21773
ACCATCGAC
22583
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23393


Nme2_
CATGGC

GC

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB10






GAAAGGGACTGAAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20964
CTGCTGGG

ACCATCGAC
22584
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23394


Nme2_
CATGGC

G

gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB9






GAAAGGGACTGAAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20965
CTGCTGGG

ACCATCGAC
22585
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23395


Nme2_
CATGGC



gAGAAAGG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GACTGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_9FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTACCATCGACgA



PB8






GAAAGGGACTGAAGCTGCTGGG






pU6_A1AT_
TAAAAA
20966
CTGCTGGG
21776
ATCGACgAG
22586
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23396


Nme2_
CATGGC

GCCATGTT

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB17






AGGGACTGAAGCTGCTGGGGCCATGTTT






pU6_A1AT_
TAAAAA
20967
CTGCTGGG
21777
ATCGACgAG
22587
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23397


Nme2_
CATGGC

GCCATGTT

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB16






AGGGACTGAAGCTGCTGGGGCCATGTT






pU6_A1AT_
TAAAAA
20968
CTGCTGGG
21778
ATCGACgAG
22588
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23398


Nme2_
CATGGC

GCCATGT

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB15






AGGGACTGAAGCTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20969
CTGCTGGG
21779
ATCGACgAG
22589
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23399


Nme2_
CATGGC

GCCATG

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB14






AGGGACTGAAGCTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20970
CTGCTGGG
21780
ATCGACgAG
22590
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23400


Nme2_
CATGGC

GCCAT

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB13






AGGGACTGAAGCTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20971
CTGCTGGG
21781
ATCGACgAG
22591
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23401


Nme2_
CATGGC

GCCA

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB12






AGGGACTGAAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20972
CTGCTGGG
21782
ATCGACgAG
22592
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23402


Nme2_
CATGGC

GCC

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB11






AGGGACTGAAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20973
CTGCTGGG
21783
ATCGACgAG
22593
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23403


Nme2_
CATGGC

GC

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB10






AGGGACTGAAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20974
CTGCTGGG

ATCGACgAG
22594
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23404


Nme2_
CATGGC

G

AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB9






AGGGACTGAAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20975
CTGCTGGG

ATCGACgAG
22595
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23405


Nme2_
CATGGC



AAAGGGAC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



TGAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_6FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTATCGACgAGAA



PB8






AGGGACTGAAGCTGCTGGG






pU6_A1AT_
TAAAAA
20976
CTGCTGGG
21786
TCGACgAGA
22596
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23406


Nme2_
CATGGC

GCCATGTT

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB17






GGGACTGAAGCTGCTGGGGCCATGTTT






pU6_A1AT_
TAAAAA
20977
CTGCTGGG
21787
TCGACgAGA
22597
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23407


Nme2_
CATGGC

GCCATGTT

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB16






GGGACTGAAGCTGCTGGGGCCATGTT






pU6_A1AT_
TAAAAA
20978
CTGCTGGG
21788
TCGACgAGA
22598
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23408


Nme2_
CATGGC

GCCATGT

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB15






GGGACTGAAGCTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20979
CTGCTGGG
21789
TCGACgAGA
22599
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23409


Nme2_
CATGGC

GCCATG

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB14






GGGACTGAAGCTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20980
CTGCTGGG
21790
TCGACgAGA
22600
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23410


Nme2_
CATGGC

GCCAT

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB13






GGGACTGAAGCTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20981
CTGCTGGG
21791
TCGACgAGA
22601
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23411


Nme2_
CATGGC

GCCA

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB12






GGGACTGAAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20982
CTGCTGGG
21792
TCGACgAGA
22602
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23412


Nme2_
CATGGC

GCC

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB11






GGGACTGAAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20983
CTGCTGGG
21793
TCGACgAGA
22603
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23413


Nme2_
CATGGC

GC

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB10






GGGACTGAAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20984
CTGCTGGG

TCGACgAGA
22604
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23414


Nme2_
CATGGC

G

AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB9






GGGACTGAAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20985
CTGCTGGG

TCGACgAGA

TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23415


Nme2_
CATGGC



AAGGGACT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



GAAG
22605
AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_5FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTTCGACgAGAAA



PB8






GGGACTGAAGCTGCTGGG






pU6_A1AT_
TAAAAA
20986
CTGCTGGG
21796
GACgAGAA
22606
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23416


Nme2_
CATGGC

GCCATGTT

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC

T

AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB17






GGACTGAAGCTGCTGGGGCCATGTTT






pU6_A1AT_
TAAAAA
20987
CTGCTGGG
21797
GACgAGAA
22607
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23417


Nme2_
CATGGC

GCCATGTT

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB16






GGACTGAAGCTGCTGGGGCCATGTT






pU6_A1AT_
TAAAAA
20988
CTGCTGGG
21798
GACgAGAA
22608
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23418


Nme2_
CATGGC

GCCATGT

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB15






GGACTGAAGCTGCTGGGGCCATGT






pU6_A1AT_
TAAAAA
20989
CTGCTGGG
21799
GACgAGAA
22609
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23419


Nme2_
CATGGC

GCCATG

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB14






GGACTGAAGCTGCTGGGGCCATG






pU6_A1AT_
TAAAAA
20990
CTGCTGGG
21800
GACgAGAA
22610
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23420


Nme2_
CATGGC

GCCAT

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB13






GGACTGAAGCTGCTGGGGCCAT






pU6_A1AT_
TAAAAA
20991
CTGCTGGG
21801
GACgAGAA
22611
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23421


Nme2_
CATGGC

GCCA

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB12






GGACTGAAGCTGCTGGGGCCA






pU6_A1AT_
TAAAAA
20992
CTGCTGGG
21802
GACgAGAA
22612
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23422


Nme2_
CATGGC



AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT

GCC



TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB11






GGACTGAAGCTGCTGGGGCC






pU6_A1AT_
TAAAAA
20993
CTGCTGGG
21803
GACgAGAA
22613
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23423


Nme2_
CATGGC

GC

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB10






GGACTGAAGCTGCTGGGGC






pU6_A1AT_
TAAAAA
20994
CTGCTGGG

GACgAGAA
22614
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23424


Nme2_
CATGGC

G

AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB9






GGACTGAAGCTGCTGGGG






pU6_A1AT_
TAAAAA
20995
CTGCTGGG

GACgAGAA
22615
TAAAAACATGGCCCCAGCAGCTTGTTGTAGCT
23425


Nme2_
CATGGC



AGGGACTG

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED15+_
CCCAGC



AAG

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



G_3FE_
AGCTT





TTCTGCTTTAAGGGGCATCGTTGACgAGAAAG



PB8






GGACTGAAGCTGCTGGG






pU6_A1AT_
AGGCC
20996
AGCCTTAT
21806
ACATGGCCC
22616
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23426


SpCas9-
GTGCAT

GCACGGCC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB17




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCT








C








pU6_A1AT_
AGGCC
20997
AGCCTTAT
21807
ACATGGCCC
22617
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23427


SpCas9-
GTGCAT

GCACGGCC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB16




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCC








C








pU6_A1AT_
AGGCC
20998
AGCCTTAT
21808
ACATGGCCC
22618
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23428


SpCas9-
GTGCAT

GCACGGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB15




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGC








C








pU6_A1AT_
AGGCC
20999
AGCCTTAT
21809
ACATGGCCC
22619
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23429


SpCas9-
GTGCAT

GCACGG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB14




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGG








C








pU6_A1AT_
AGGCC
21000
AGCCTTAT
21810
ACATGGCCC
22620
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23430


SpCas9-
GTGCAT

GCACG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB13




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACG








C








pU6_A1AT_
AGGCC
21001
AGCCTTAT
21811
ACATGGCCC
22621
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23431


SpCas9-
GTGCAT

GCAC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



30FE_PB12




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCAC








C








pU6_A1AT_
AGGCC
21002
AGCCTTAT
21812
ACATGGCCC
22622
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23432


SpCas9-
GTGCAT

GCA

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB11




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCA








C








pU6_A1AT_
AGGCC
21003
AGCCTTAT
21813
ACATGGCCC
22623
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23433


SpCas9-
GTGCAT

GC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB10




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGC








C








pU6_A1AT_
AGGCC
21004
AGCCTTAT

ACATGGCCC
22624
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23434


SpCas9-
GTGCAT

G

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB9




TGGTCAGCA

CGATGGTCAGCACAGCCTTATG








C








pU6_A1AT_
AGGCC
21005
AGCCTTAT

ACATGGCCC
22625
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23435


SpCas9-
GTGCAT



CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



30FE_PB8




TGGTCAGCA

CGATGGTCAGCACAGCCTTAT








C








pU6_A1AT_
AGGCC
21006
AGCCTTAT
21816
GCCCCAGCA
22626
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23436


SpCas9-
GTGCAT

GCACGGCC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB17




GCAC

GTCAGCACAGCCTTATGCACGGCCT






pU6_A1AT_
AGGCC
21007
AGCCTTAT
21817
GCCCCAGCA
22627
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23437


SpCas9-
GTGCAT

GCACGGCC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB16




GCAC

GTCAGCACAGCCTTATGCACGGCC






pU6_A1AT_
AGGCC
21008
AGCCTTAT
21818
GCCCCAGCA
22628
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23438


SpCas9-
GTGCAT

GCACGGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB15




GCAC

GTCAGCACAGCCTTATGCACGGC






pU6_A1AT_
AGGCC
21009
AGCCTTAT
21819
GCCCCAGCA
22629
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23439


SpCas9-
GTGCAT

GCACGG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB14




GCAC

GTCAGCACAGCCTTATGCACGG






pU6_A1AT_
AGGCC
21010
AGCCTTAT
21820
GCCCCAGCA
22630
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23440


SpCas9-
GTGCAT

GCACG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB13




GCAC

GTCAGCACAGCCTTATGCACG






pU6_A1AT_
AGGCC
21011
AGCCTTAT
21821
GCCCCAGCA
22631
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23441


SpCas9-
GTGCAT

GCAC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB12




GCAC

GTCAGCACAGCCTTATGCAC






pU6_A1AT_
AGGCC
21012
AGCCTTAT
21822
GCCCCAGCA
22632
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23442


SpCas9-
GTGCAT

GCA

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB11




GCAC

GTCAGCACAGCCTTATGCA






pU6_A1AT_
AGGCC
21013
AGCCTTAT
21823
GCCCCAGCA
22633
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23443


SpCas9-
GTGCAT

GC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB10




GCAC

GTCAGCACAGCCTTATGC






pU6_A1AT_
AGGCC
21014
AGCCTTAT

GCCCCAGCA
22634
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23444


SpCas9-
GTGCAT

G

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB9




GCAC

GTCAGCACAGCCTTATG






pU6_A1AT_
AGGCC
21015
AGCCTTAT

GCCCCAGCA
22635
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23445


SpCas9-
GTGCAT



GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



25FE_PB8




GCAC

GTCAGCACAGCCTTAT






pU6_A1AT_
AGGCC
21016
AGCCTTAT
21826
AGCAGCTTC
22636
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23446


SpCas9-
GTGCAT

GCACGGCC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB17






CACAGCCTTATGCACGGCCT






pU6_A1AT_
AGGCC
21017
AGCCTTAT
21827
AGCAGCTTC
22637
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23447


SpCas9-
GTGCAT

GCACGGCC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB16






CACAGCCTTATGCACGGCC






pU6_A1AT_
AGGCC
21018
AGCCTTAT
21828
AGCAGCTTC
22638
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23448


SpCas9-
GTGCAT

GCACGGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB15






CACAGCCTTATGCACGGC






pU6_A1AT_
AGGCC
21019
AGCCTTAT
21829
AGCAGCTTC
22639
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23449


SpCas9-
GTGCAT

GCACGG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB14






CACAGCCTTATGCACGG






pU6_A1AT_
AGGCC
21020
AGCCTTAT
21830
AGCAGCTTC
22640
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23450


SpCas9-
GTGCAT

GCACG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB13






CACAGCCTTATGCACG






pU6_A1AT_
AGGCC
21021
AGCCTTAT
21831
AGCAGCTTC
22641
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23451


SpCas9-
GTGCAT

GCAC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_






ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
AAGGCT



CTcGTCGAT

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB12
GTG



GGTCAGCAC

CACAGCCTTATGCAC






pU6_A1AT_
AGGCC
21022
AGCCTTAT
21832
AGCAGCTTC
22642
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23452


SpCas9-
GTGCAT

GCA

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB11






CACAGCCTTATGCA






pU6_A1AT_
AGGCC
21023
AGCCTTAT
21833
AGCAGCTTC
22643
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23453


SpCas9-
GTGCAT

GC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB10






CACAGCCTTATGC






pU6_A1AT_
AGGCC
21024
AGCCTTAT

AGCAGCTTC
22644
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23454


SpCas9-
GTGCAT

G

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB9






CACAGCCTTATG






pU6_A1AT_
AGGCC
21025
AGCCTTAT

AGCAGCTTC
22645
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23455


SpCas9-
GTGCAT



AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



20FE_PB8






CACAGCCTTAT






pU6_A1AT_
AGGCC
21026
AGCCTTAT
21836
TTCAGTCCC
22646
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23456


SpCas9-
GTGCAT

GCACGGCC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB17






CTTATGCACGGCCT






pU6_A1AT_
AGGCC
21027
AGCCTTAT
21837
TTCAGTCCC
22647
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23457


SpCas9-
GTGCAT

GCACGGCC

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB16






CTTATGCACGGCC






pU6_A1AT_
AGGCC
21028
AGCCTTAT
21838
TTCAGTCCC
22648
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23458


SpCas9-
GTGCAT

GCACGGC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB15






CTTATGCACGGC






pU6_A1AT_
AGGCC
21029
AGCCTTAT
21839
TTCAGTCCC
22649
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23459


SpCas9-
GTGCAT

GCACGG

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB14






CTTATGCACGG






pU6_A1AT_
AGGCC
21030
AGCCTTAT
21840
TTCAGTCCC
22650
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23460


SpCas9-
GTGCAT

GCACG

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB13






CTTATGCACG






pU6_A1AT_
AGGCC
21031
AGCCTTAT
21841
TTCAGTCCC
22651
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23461


SpCas9-
GTGCAT

GCAC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB12






CTTATGCAC






pU6_A1AT_
AGGCC
21032
AGCCTTAT
21842
TTCAGTCCC
22652
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23462


SpCas9-
GTGCAT

GCA

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB11






CTTATGCA






pU6_A1AT_
AGGCC
21033
AGCCTTAT
21843
TTCAGTCCC
22653
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23463


SpCas9-
GTGCAT

GC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTCGTCGATGGTCAGCACAGC



14FE_PB10






CTTATGC






pU6_A1AT_
AGGCC
21034
AGCCTTAT

TTCAGTCCC
22654
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23464


SpCas9-
GTGCAT

G

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTCGTCGATGGTCAGCACAGC



14FE_PB9






CTTATG






pU6_A1AT_
AGGCC
21035
AGCCTTAT

TTCAGTCCC
22655
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23465


SpCas9-
GTGCAT



TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG



GCAC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



14FE_PB8






CTTAT






pU6_A1AT_
AGGCC
21036
AGCCTTAT
21846
AGTCCCTTT
22656
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23466


SpCas9-
GTGCAT

GCACGGCC

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB17






ATGCACGGCCT






pU6_A1AT_
AGGCC
21037
AGCCTTAT
21847
AGTCCCTTT
22657
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23467


SpCas9-
GTGCAT

GCACGGCC

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB16






ATGCACGGCC






pU6_A1AT_
AGGCC
21038
AGCCTTAT
21848
AGTCCCTTT
22658
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23468


SpCas9-
GTGCAT

GCACGGC

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB15






ATGCACGGC






pU6_A1AT_
AGGCC
21039
AGCCTTAT
21849
AGTCCCTTT
22659
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23469


SpCas9-
GTGCAT

GCACGG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB14






ATGCACGG






pU6_A1AT_
AGGCC
21040
AGCCTTAT
21850
AGTCCCTTT
22660
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23470


SpCas9-
GTGCAT

GCACG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB13






ATGCACG






pU6_A1AT_
AGGCC
21041
AGCCTTAT
21851
AGTCCCTTT
22661
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23471


SpCas9-
GTGCAT

GCAC

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB12






ATGCAC






pU6_A1AT_
AGGCC
21042
AGCCTTAT
21852
AGTCCCTTT
22662
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23472


SpCas9-
GTGCAT

GCA

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB11






ATGCA






pU6_A1AT_
AGGCC
21043
AGCCTTAT
21853
AGTCCCTTT
22663
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23473


SpCas9-
GTGCAT

GC

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB10






ATGC






pU6_A1AT_
AGGCC
21044
AGCCTTAT

AGTCCCTTT
22664
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23474


SpCas9-
GTGCAT

G

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB9






ATG






pU6_A1AT_
AGGCC
21045
AGCCTTAT

AGTCCCTTT
22665
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23475


SpCas9-
GTGCAT



CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



11FE_PB8






AT






pU6_A1AT_
AGGCC
21046
AGCCTTAT
21856
TCCCTTTCTc
22666
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23476


SpCas9-
GTGCAT

GCACGGCC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB17






GCACGGCCT






pU6_A1AT_
AGGCC
21047
AGCCTTAT
21857
TCCCTTTCTc
22667
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23477


SpCas9-
GTGCAT

GCACGGCC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB16






GCACGGCC






pU6_A1AT_
AGGCC
21048
AGCCTTAT
21858
TCCCTTTCTc
22668
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23478


SpCas9-
GTGCAT

GCACGGC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB15






GCACGGC






pU6_A1AT_
AGGCC
21049
AGCCTTAT
21859
TCCCTTTCTc
22669
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23479


SpCas9-
GTGCAT

GCACGG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB14






GCACGG






pU6_A1AT_
AGGCC
21050
AGCCTTAT
21860
TCCCTTTCTc
22670
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23480


SpCas9-
GTGCAT

GCACG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB13






GCACG






pU6_A1AT_
AGGCC
21051
AGCCTTAT
21861
TCCCTTTCTc
22671
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23481


SpCas9-
GTGCAT

GCAC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB12






GCAC






pU6_A1AT_
AGGCC
21052
AGCCTTAT
21862
TCCCTTTCTC
22672
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23482


SpCas9-
GTGCAT

GCA

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB11






GCA






pU6_A1AT_
AGGCC
21053
AGCCTTAT
21863
TCCCTTTCTc
22673
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23483


SpCas9-
GTGCAT

GC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB10






GC






pU6_A1AT_
AGGCC
21054
AGCCTTAT

TCCCTTTCTc
22674
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23484


SpCas9-
GTGCAT

G

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB9






G






pU6_A1AT_
AGGCC
21055
AGCCTTAT

TCCCTTTCTc
22675
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23485


SpCas9-
GTGCAT



GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



CAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



9FE_PB8













pU6_A1AT_
AGGCC
21056
AGCCTTAT
21866
CCTTTCTcGT
22676
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23486


SpCas9-
GTGCAT

GCACGGCC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB17






ACGGCCT






pU6_A1AT_
AGGCC
21057
AGCCTTAT
21867
CCTTTCTCGT
22677
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23487


SpCas9-
GTGCAT

GCACGGCC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB16






ACGGCC






pU6_A1AT_
AGGCC
21058
AGCCTTAT
21868
CCTTTCTcGT
22678
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23488


SpCas9-
GTGCAT

GCACGGC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB15






ACGGC






pU6_A1AT_
AGGCC
21059
AGCCTTAT
21869
CCTTTCTcGT
22679
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23489


SpCas9-
GTGCAT

GCACGG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB14






ACGG






pU6_A1AT_
AGGCC
21060
AGCCTTAT
21870
CCTTTCTcGT
22680
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23490


SpCas9-
GTGCAT

GCACG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB13






ACG






pU6_A1AT_
AGGCC
21061
AGCCTTAT
21871
CCTTTCTcGT
22681
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23491


SpCas9-
GTGCAT

GCAC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB12






AC






pU6_A1AT_
AGGCC
21062
AGCCTTAT
21872
CCTTTCTcGT
22682
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23492


SpCas9-
GTGCAT

GCA

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB11






A






pU6_A1AT_
AGGCC
21063
AGCCTTAT
21873
CCTTTCTcGT
22683
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23493


SpCas9-
GTGCAT

GC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



7FE_PB10













pU6_A1AT_
AGGCC
21064
AGCCTTAT

CCTTTCTcGT
22684
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23494


SpCas9-
GTGCAT

G

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATG



7FE_PB9













pU6_A1AT_
AGGCC
21065
AGCCTTAT

CCTTTCTcGT
22685
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23495


SpCas9-
GTGCAT



CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





CCTTTCTcGTCGATGGTCAGCACAGCCTTAT



7FE_PB8













pU6_A1AT_
AGGCC
21066
AGCCTTAT
21876
TTTCTCGTC
22686
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23496


SpCas9-
GTGCAT

GCACGGCC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



5FE_PB17






GGCCT






pU6_A1AT_
AGGCC
21067
AGCCTTAT
21877
TTTCTcGTC
22687
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23497


SpCas9-
GTGCAT

GCACGGCC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



5FE_PB16






GGCC






pU6_A1AT_
AGGCC
21068
AGCCTTAT
21878
TTTCTcGTC
22688
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23498


SpCas9-
GTGCAT

GCACGGC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



5FE_PB15






GGC






pU6_A1AT_
AGGCC
21069
AGCCTTAT
21879
TTTCTCGTC
22689
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23499


SpCas9-
GTGCAT

GCACGG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



5FE_PB14






GG






pU6_A1AT_
AGGCC
21070
AGCCTTAT
21880
TTTCTCGTC
22690
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23500


SpCas9-
GTGCAT

GCACG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



5FE_PB13






G






pU6_A1AT_
AGGCC
21071
AGCCTTAT
21881
TTTCTcGTC
22691
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23501


SpCas9-
GTGCAT

GCAC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



5FE_PB12













pU6_A1AT_
AGGCC
21072
AGCCTTAT
21882
TTTCTCGTC
22692
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23502


SpCas9-
GTGCAT

GCA

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCA



5FE_PB11













pU6_A1AT_
AGGCC
21073
AGCCTTAT
21883
TTTCTCGTC
22693
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23503


SpCas9-
GTGCAT

GC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTCGTCGATGGTCAGCACAGCCTTATGC



5FE_PB10













pU6_A1AT_
AGGCC
21074
AGCCTTAT

TTTCTCGTC
22694
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23504


SpCas9-
GTGCAT

G

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTcGTCGATGGTCAGCACAGCCTTATG



5FE_PB9













pU6_A1AT_
AGGCC
21075
AGCCTTAT

TTTCTcGTC
22695
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23505


SpCas9-
GTGCAT



GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



GCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TTTCTCGTCGATGGTCAGCACAGCCTTAT



5FE_PB8













pU6_A1AT_
AGGCC
21076
AGCCTTAT
21886
TCTcGTCGA
22696
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23506


SpCas9-
GTGCAT

GCACGGCC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT

T

C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



3FE_PB17






GCCT






pU6_A1AT_
AGGCC
21077
AGCCTTAT
21887
TCTcGTCGA
22697
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23507


SpCas9-
GTGCAT

GCACGGCC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



3FE_PB16






GCC






pU6_A1AT_
AGGCC
21078
AGCCTTAT
21888
TCTcGTCGA
22698
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23508


SpCas9-
GTGCAT

GCACGGC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



3FE_PB15






GC






pU6_A1AT_
AGGCC
21079
AGCCTTAT
21889
TCTcGTCGA
22699
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23509


SpCas9-
GTGCAT

GCACGG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



3FE_PB14






G






pU6_A1AT_
AGGCC
21080
AGCCTTAT
21890
TCTcGTCGA
22700
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23510


SpCas9-
GTGCAT

GCACG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



3FE_PB13













pU6_A1AT_
AGGCC
21081
AGCCTTAT
21891
TCTcGTCGA
22701
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23511


SpCas9-
GTGCAT

GCAC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCAC



3FE_PB12













pU6_A1AT_
AGGCC
21082
AGCCTTAT
21892
TCTcGTCGA
22702
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23512


SpCas9-
GTGCAT

GCA

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGCA



3FE_PB11













pU6_A1AT_
AGGCC
21083
AGCCTTAT
21893
TCTcGTCGA
22703
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23513


SpCas9-
GTGCAT

GC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATGC



3FE_PB10













pU6_A1AT_
AGGCC
21084
AGCCTTAT

TCTcGTCGA
22704
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23514


SpCas9-
GTGCAT

G

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTATG



3FE_PB9













pU6_A1AT_
AGGCC
21085
AGCCTTAT

TCTcGTCGA
22705
AGGCCGTGCATAAGGCTGTGGTTTTAGAGCTA
23515


SpCas9-
GTGCAT



TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_SpRY_
AAGGCT



C

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



ED15-_
GTG





TCTcGTCGATGGTCAGCACAGCCTTAT



3FE_PB8













pU6_A1AT_
TCCAGG
21086
AGCCTTAT
21896
ACATGGCCC
22706
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23516


Nme2_
CCGTGC

GCACGGCC

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB17




C

ACAGCCTTATGCACGGCCT






pU6_A1AT_
TCCAGG
21087
AGCCTTAT
21897
ACATGGCCC
22707
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23517


Nme2_
CCGTGC

GCACGGCC

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB16




C

ACAGCCTTATGCACGGCC






pU6_A1AT_
TCCAGG
21088
AGCCTTAT
21898
ACATGGCCC
22708
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23518


Nme2_
CCGTGC

GCACGGC

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB15




C

ACAGCCTTATGCACGGC






pU6_A1AT_
TCCAGG
21089
AGCCTTAT
21899
ACATGGCCC
22709
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23519


Nme2_
CCGTGC

GCACGG

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB14




C

ACAGCCTTATGCACGG






pU6_A1AT_
TCCAGG
21090
AGCCTTAT
21900
ACATGGCCC
22710
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23520


Nme2_
CCGTGC

GCACG

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB13




C

ACAGCCTTATGCACG






pU6_A1AT_
TCCAGG
21091
AGCCTTAT
21901
ACATGGCCC
22711
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23521


Nme2_
CCGTGC

GCAC

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB12




C

ACAGCCTTATGCAC






pU6_A1AT_
TCCAGG
21092
AGCCTTAT
21902
ACATGGCCC
22712
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23522


Nme2_
CCGTGC

GCA

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB11




C

ACAGCCTTATGCA






pU6_A1
TCCAGG
21093
AGCCTTAT
21903
ACATGGCCC
22713
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23523


AT_Nm
CCGTGC

GC

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



e2_ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



G_30F




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



E_PB10




C

ACAGCCTTATGC






pU6_A1AT_
TCCAGG
21094
AGCCTTAT

ACATGGCCC
22714
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23524


Nme2_
CCGTGC

G

CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB9




C

ACAGCCTTATG






pU6_A1AT_
TCCAGG
21095
AGCCTTAT

ACATGGCCC
22715
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23525


Nme2_
CCGTGC



CAGCAGCTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGTCCCTT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



TCTcGTCGA

TTCTGCTTTAAGGGGCATCGTTACATGGCCCCA



_G_30FE_




TGGTCAGCA

GCAGCTTCAGTCCCTTTCTcGTCGATGGTCAGC



PB8




C

ACAGCCTTAT






pU6_A1
TCCAGG
21096
AGCCTTAT
21906
GCCCCAGCA
22716
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23526


AT_Nme2_
CCGTGC

GCACGGCC

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

CCTTTCTCGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB17






CCTTATGCACGGCCT






pU6_A1AT_
TCCAGG
21097
AGCCTTAT
21907
GCCCCAGCA
22717
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23527


Nme2_
CCGTGC

GCACGGCC

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTCGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB16






CCTTATGCACGGCC






pU6_A1AT_
TCCAGG
21098
AGCCTTAT
21908
GCCCCAGCA
22718
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23528


Nme2_
CCGTGC

GCACGGC

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB15






CCTTATGCACGGC






pU6_A1AT_
TCCAGG
21099
AGCCTTAT
21909
GCCCCAGCA
22719
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23529


Nme2_
CCGTGC

GCACGG

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB14






CCTTATGCACGG






pU6_A1AT_
TCCAGG
21100
AGCCTTAT
21910
GCCCCAGCA
22720
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23530


Nme2_
CCGTGC

GCACG

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB13






CCTTATGCACG






pU6_A1AT_
TCCAGG
21101
AGCCTTAT
21911
GCCCCAGCA
22721
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23531


Nme2_
CCGTGC

GCAC

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB12






CCTTATGCAC






pU6_A1AT_
TCCAGG
21102
AGCCTTAT
21912
GCCCCAGCA
22722
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23532


Nme2_
CCGTGC

GCA

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB11






CCTTATGCA






pU6_A1AT_
TCCAGG
21103
AGCCTTAT
21913
GCCCCAGCA
22723
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23533


Nme2_
CCGTGC

GC

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB10






CCTTATGC






pU6_A1AT_
TCCAGG
21104
AGCCTTAT

GCCCCAGCA
22724
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23534


Nme2_
CCGTGC

G

GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTCGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB9






CCTTATG






pU6_A1AT_
TCCAGG
21105
AGCCTTAT

GCCCCAGCA
22725
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23535


Nme2_
CCGTGC



GCTTCAGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CCTTTCTcGT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



CGATGGTCA

TTCTGCTTTAAGGGGCATCGTTGCCCCAGCAG



_G_25FE_




GCAC

CTTCAGTCCCTTTCTcGTCGATGGTCAGCACAG



PB8






CCTTAT






pU6_A1AT_
TCCAGG
21106
AGCCTTAT
21916
AGCAGCTTC
22726
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23536


Nme2_
CCGTGC

GCACGGCC

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB17






GCACGGCCT






pU6_A1AT_
TCCAGG
21107
AGCCTTAT
21917
AGCAGCTTC
22727
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23537


Nme2_
CCGTGC

GCACGGCC

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB16






GCACGGCC






pU6_A1AT_
TCCAGG
21108
AGCCTTAT
21918
AGCAGCTTC
22728
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23538


Nme2_
CCGTGC

GCACGGC

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB15






GCACGGC






pU6_A1AT_
TCCAGG
21109
AGCCTTAT
21919
AGCAGCTTC
22729
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23539


Nme2_
CCGTGC

GCACGG

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTCGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB14






GCACGG






pU6_A1AT_
TCCAGG
21110
AGCCTTAT
21920
AGCAGCTTC
22730
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23540


Nme2_
CCGTGC

GCACG

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB13






GCACG






pU6_A1AT_
TCCAGG
21111
AGCCTTAT
21921
AGCAGCTTC
22731
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23541


Nme2_
CCGTGC

GCAC

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB12






GCAC






pU6_A1AT_
TCCAGG
21112
AGCCTTAT
21922
AGCAGCTTC
22732
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23542


Nme2_
CCGTGC

GCA

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTCGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB11






GCA






pU6_A1AT_
TCCAGG
21113
AGCCTTAT
21923
AGCAGCTTC
22733
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23543


Nme2_
CCGTGC

GC

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB10






GC






pU6_A1AT_
TCCAGG
21114
AGCCTTAT

AGCAGCTTC
22734
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23544


Nme2_
CCGTGC

G

AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTcGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB9






G






pU6_A1AT_
TCCAGG
21115
AGCCTTAT

AGCAGCTTC
22735
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23545


Nme2_
CCGTGC



AGTCCCTTT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CTCGTCGAT

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GGTCAGCAC

TTCTGCTTTAAGGGGCATCGTTAGCAGCTTCAG



_G_20FE_






TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB8













pU6_A1AT_
TCCAGG
21116
AGCCTTAT
21926
TTCAGTCCC
22736
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23546


Nme2_
CCGTGC

GCACGGCC

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB17






GCCT






pU6_A1AT_
TCCAGG
21117
AGCCTTAT
21927
TTCAGTCCC
22737
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23547


Nme2_
CCGTGC

GCACGGCC

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB16






GCC






pU6_A1AT_
TCCAGG
21118
AGCCTTAT
21928
TTCAGTCCC
22738
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23548


Nme2_
CCGTGC

GCACGGC

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB15






GC






pU6_A1AT_
TCCAGG
21119
AGCCTTAT
21929
TTCAGTCCC
22739
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23549


Nme2_
CCGTGC

GCACGG

TTTCTcGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB14






G






pU6_A1AT_
TCCAGG
21120
AGCCTTAT
21930
TTCAGTCCC
22740
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23550


Nme2_
CCGTGC

GCACG

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB13













pU6_A1AT_
TCCAGG
21121
AGCCTTAT
21931
TTCAGTCCC
22741
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23551


Nme2_
CCGTGC

GCAC

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCAC



PB12













pU6_A1AT_
TCCAGG
21122
AGCCTTAT
21932
TTCAGTCCC
22742
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23552


Nme2_
CCGTGC

GCA

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGCA



PB11













pU6_A1AT_
TCCAGG
21123
AGCCTTAT
21933
TTCAGTCCC
22743
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23553


Nme2_
CCGTGC

GC

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATGC



PB10













pU6_A1AT_
TCCAGG
21124
AGCCTTAT

TTCAGTCCC
22744
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23554


Nme2_
CCGTGC

G

TTTCTCGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTATG



PB9













pU6_A1AT_
TCCAGG
21125
AGCCTTAT

TTCAGTCCC
22745
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23555


Nme2_
CCGTGC



TTTCTcGTC

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GATGGTCA

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG



GCAC

TTCTGCTTTAAGGGGCATCGTTTTCAGTCCCTT



_G_14FE_






TCTcGTCGATGGTCAGCACAGCCTTAT



PB8













pU6_A1AT_
TCCAGG
21126
AGCCTTAT
21936
AGTCCCTTT
22746
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23556


Nme2_
CCGTGC

GCACGGCC

CTCGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTc



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCACGGCCT



PB17













pU6_A1AT_
TCCAGG
21127
AGCCTTAT
21937
AGTCCCTTT
22747
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23557


Nme2_
CCGTGC

GCACGGCC

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTC



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCACGGCC



PB16













pU6_A1AT_
TCCAGG
21128
AGCCTTAT
21938
AGTCCCTTT
22748
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23558


Nme2_
CCGTGC

GCACGGC

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTC



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCACGGC



PB15













pU6_A1AT_
TCCAGG
21129
AGCCTTAT
21939
AGTCCCTTT
22749
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23559


Nme2_
CCGTGC

GCACGG

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTc



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCACGG



PB14













pU6_A1AT_
TCCAGG
21130
AGCCTTAT
21940
AGTCCCTTT
22750
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23560


Nme2_
CCGTGC

GCACG

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTc



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCACG



PB13













pU6_A1AT_
TCCAGG
21131
AGCCTTAT
21941
AGTCCCTTT
22751
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23561


Nme2_
CCGTGC

GCAC

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTc



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCAC



PB12













pU6_A1AT_
TCCAGG
21132
AGCCTTAT
21942
AGTCCCTTT
22752
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23562


Nme2_
CCGTGC

GCA

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTC



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGCA



PB11













pU6_A1AT_
TCCAGG
21133
AGCCTTAT
21943
AGTCCCTTT
22753
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23563


Nme2_
CCGTGC

GC

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTC



_G_11FE_






GTCGATGGTCAGCACAGCCTTATGC



PB10













pU6_A1AT_
TCCAGG
21134
AGCCTTAT

AGTCCCTTT
22754
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23564


Nme2_
CCGTGC

G

CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTC



_G_11FE_






GTCGATGGTCAGCACAGCCTTATG



PB9













pU6_A1AT_
TCCAGG
21135
AGCCTTAT

AGTCCCTTT
22755
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23565


Nme2_
CCGTGC



CTcGTCGAT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GGTCAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTAGTCCCTTTCTc



_G_11FE_






GTCGATGGTCAGCACAGCCTTAT



PB8













pU6_A1AT_
TCCAGG
21136
AGCCTTAT
21946
TCCCTTTCTc
22756
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23566


Nme2_
CCGTGC

GCACGGCC

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCACGGCCT



PB17













pU6_A1AT_
TCCAGG
21137
AGCCTTAT
21947
TCCCTTTCTc
22757
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23567


Nme2_
CCGTGC

GCACGGCC

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCACGGCC



PB16













pU6_A1AT_
TCCAGG
21138
AGCCTTAT
21948
TCCCTTTCTc
22758
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23568


Nme2_
CCGTGC

GCACGGC

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCACGGC



PB15













pU6_A1AT_
TCCAGG
21139
AGCCTTAT
21949
TCCCTTTCTc
22759
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23569


Nme2_
CCGTGC

GCACGG

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCACGG



PB14













pU6_A1AT_
TCCAGG
21140
AGCCTTAT
21950
TCCCTTTCTc
22760
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23570


Nme2_
CCGTGC

GCACG

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCACG



PB13













pU6_A1AT_
TCCAGG
21141
AGCCTTAT
21951
TCCCTTTCTc
22761
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23571


Nme2_
CCGTGC

GCAC

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCAC



PB12













pU6_A1AT_
TCCAGG
21142
AGCCTTAT
21952
TCCCTTTCTc
22762
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23572


Nme2_
CCGTGC

GCA

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGCA



PB11













pU6_A1AT_
TCCAGG
21143
AGCCTTAT
21953
TCCCTTTCTc
22763
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23573


Nme2_
CCGTGC

GC

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATGC



PB10













pU6_A1AT_
TCCAGG
21144
AGCCTTAT

TCCCTTTCTc
22764
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23574


Nme2_
CCGTGC

G

GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTATG



PB9













pU6_A1AT_
TCCAGG
21145
AGCCTTAT

TCCCTTTCTc
22765
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23575


Nme2_
CCGTGC



GTCGATGGT

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



CAGCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCCCTTTCTcGT



_G_9FE_






CGATGGTCAGCACAGCCTTAT



PB8













pU6_A1AT_
TCCAGG
21146
AGCCTTAT
21956
CCTTTCTcGT
22766
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23576


Nme2_
CCGTGC

GCACGGCC

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCACGGCCT



PB17













pU6_A1AT_
TCCAGG
21147
AGCCTTAT
21957
CCTTTCTcGT
22767
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23577


Nme2_
CCGTGC

GCACGGCC

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCACGGCC



PB16













pU6_A1AT_
TCCAGG
21148
AGCCTTAT
21958
CCTTTCTcGT
22768
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23578


Nme2_
CCGTGC

GCACGGC

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCACGGC



PB15













pU6_A1AT_
TCCAGG
21149
AGCCTTAT
21959
CCTTTCTcGT
22769
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23579


Nme2_
CCGTGC

GCACGG

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCACGG



PB14













pU6_A1AT_
TCCAGG
21150
AGCCTTAT
21960
CCTTTCTcGT
22770
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23580


Nme2_
CCGTGC

GCACG

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCACG



PB13













pU6_A1AT_
TCCAGG
21151
AGCCTTAT
21961
CCTTTCTcGT
22771
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23581


Nme2_
CCGTGC

GCAC

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCAC



PB12













pU6_A1AT_
TCCAGG
21152
AGCCTTAT
21962
CCTTTCTcGT
22772
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23582


Nme2_
CCGTGC

GCA

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGCA



PB11













pU6_A1AT_
TCCAGG
21153
AGCCTTAT
21963
CCTTTCTCGT
22773
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23583


Nme2_
CCGTGC

GC

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATGC



PB10













pU6_A1AT_
TCCAGG
21154
AGCCTTAT

CCTTTCTcGT
22774
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23584


Nme2_
CCGTGC

G

CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTATG



PB9













pU6_A1AT_
TCCAGG
21155
AGCCTTAT

CCTTTCTCGT
22775
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23585


Nme2_
CCGTGC



CGATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTCCTTTCTcGTC



_G_7FE_






GATGGTCAGCACAGCCTTAT



PB8













pU6_A1AT_
TCCAGG
21156
AGCCTTAT
21966
TTTCTCGTC
22776
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23586


Nme2_
CCGTGC

GCACGGCC

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCACGGCCT



PB17













pU6_A1AT_
TCCAGG
21157
AGCCTTAT
21967
TTTCTCGTC
22777
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23587


Nme2_
CCGTGC

GCACGGCC

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCACGGCC



PB16













pU6_A1AT_
TCCAGG
21158
AGCCTTAT
21968
TTTCTcGTC
22778
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23588


Nme2_
CCGTGC

GCACGGC

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCACGGC



PB15













pU6_A1AT_
TCCAGG
21159
AGCCTTAT
21969
TTTCTcGTC
22779
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23589


Nme2_
CCGTGC

GCACGG

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCACGG



PB14













pU6_A1AT_
TCCAGG
21160
AGCCTTAT
21970
TTTCTCGTC
22780
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23590


Nme2_
CCGTGC

GCACG

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTCGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCACG



PB13













pU6_A1AT_
TCCAGG
21161
AGCCTTAT
21971
TTTCTCGTC
22781
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23591


Nme2_
CCGTGC

GCAC

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCAC



PB12













pU6_A1AT_
TCCAGG
21162
AGCCTTAT
21972
TTTCTCGTC
22782
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23592


Nme2_
CCGTGC

GCA

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGCA



PB11













pU6_A1AT_
TCCAGG
21163
AGCCTTAT
21973
TTTCTCGTC
22783
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23593


Nme2_
CCGTGC

GC

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATGC



PB10













pU6_A1AT_
TCCAGG
21164
AGCCTTAT

TTTCTcGTC
22784
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23594


Nme2_
CCGTGC

G

GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTCGTCGA



_G_5FE_






TGGTCAGCACAGCCTTATG



PB9













pU6_A1AT_
TCCAGG
21165
AGCCTTAT

TTTCTCGTC
22785
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23595


Nme2_
CCGTGC



GATGGTCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



GCAC

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTTTCTcGTCGA



_G_5FE_






TGGTCAGCACAGCCTTAT



PB8













pU6_A1AT_
TCCAGG
21166
AGCCTTAT
21976
TCTcGTCGA
22786
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23596


Nme2_
CCGTGC

GCACGGCC

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG

T

C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCACGGCCT



PB17













pU6_A1AT_
TCCAGG
21167
AGCCTTAT
21977
TCTcGTCGA
22787
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23597


Nme2_
CCGTGC

GCACGGCC

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCACGGCC



PB16













pU6_A1AT_
TCCAGG
21168
AGCCTTAT
21978
TCTcGTCGA
22788
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23598


Nme2_
CCGTGC

GCACGGC

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCACGGC



PB15













pU6_A1AT_
TCCAGG
21169
AGCCTTAT
21979
TCTcGTCGA
22789
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23599


Nme2_
CCGTGC

GCACGG

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCACGG



PB14













pU6_A1AT_
TCCAGG
21170
AGCCTTAT
21980
TCTcGTCGA
22790
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23600


Nme2_
CCGTGC

GCACG

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCACG



PB13













pU6_A1AT_
TCCAGG
21171
AGCCTTAT
21981
TCTcGTCGA
22791
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23601


Nme2_
CCGTGC

GCAC

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCAC



PB12













pU6_A1AT_
TCCAGG
21172
AGCCTTAT
21982
TCTcGTCGA
22792
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23602


Nme2_
CCGTGC

GCA

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGCA



PB11













pU6_A1AT_
TCCAGG
21173
AGCCTTAT
21983
TCTcGTCGA
22793
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23603


Nme2_
CCGTGC

GC

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATGC



PB10













pU6_A1AT_
TCCAGG
21174
AGCCTTAT

TCTcGTCGA
22794
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23604


Nme2_
CCGTGC

G

TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTATG



PB9













pU6_A1AT_
TCCAGG
21175
AGCCTTAT

TCTcGTCGA
22795
TCCAGGCCGTGCATAAGGCTGTGGTTGTAGCT
23605


Nme2_
CCGTGC



TGGTCAGCA

CCCGAAACGTTGCTACAATAAGGCCGTCTGAA



ED1
ATAAGG



C

AAGATGTGCCGCAACGCTCTGCCCCTTAAAGC



5-
CTGTG





TTCTGCTTTAAGGGGCATCGTTTCTcGTCGATG



_G_3FE_






GTCAGCACAGCCTTAT



PB8













pU6_A1AT_
CCAGGC
21176
CCTTATGC
21986
ACATGGCCC
22796
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23606


SpCas9-
CGTGCA

ACGGCCTG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCTGG



PB17




CAG








pU6_A1AT_
CCAGGC
21177
CCTTATGC
21987
ACATGGCCC
22797
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23607


SpCas9-
CGTGCA

ACGGCCTG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCTG



PB16




CAG








pU6_A1AT_
CCAGGC
21178
CCTTATGC
21988
ACATGGCCC
22798
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23608


SpCas9-
CGTGCA

ACGGCCT

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCT



PB15




CAG








pU6_A1AT_
CCAGGC
21179
CCTTATGC
21989
ACATGGCCC
22799
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23609


SpCas9-
CGTGCA

ACGGCC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCC



PB14




CAG








pU6_A1AT_
CCAGGC
21180
CCTTATGC
21990
ACATGGCCC
22800
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23610


SpCas9-
CGTGCA

ACGGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT




TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



NG_ED1
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTCGT



7-




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGC



_30FE_




CAG





PB13













pU6_A1AT_
CCAGGC
21181
CCTTATGC
21991
ACATGGCCC
22801
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23611


SpCas9-
CGTGCA

ACGG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT




TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



NG_ED1
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



7-




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGG



_30FE_




CAG





PB12













pU6_A1AT_
CCAGGC
21182
CCTTATGC
21992
ACATGGCCC
22802
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23612


SpCas9-
CGTGCA

ACG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACG



PB11




CAG








pU6_A1AT_
CCAGGC
21183
CCTTATGC
21993
ACATGGCCC
22803
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23613


SpCas9-
CGTGCA

AC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCAC



PB10




CAG








pU6_A1AT_
CCAGGC
21184
CCTTATGC

ACATGGCCC
22804
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23614


SpCas9-
CGTGCA

A

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCA



PB9




CAG








pU6_A1AT_
CCAGGC
21185
CCTTATGC

ACATGGCCC
22805
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23615


SpCas9-
CGTGCA



CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



_30FE_




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGC



PB8




CAG








pU6_A1AT_
CCAGGC
21186
CCTTATGC
21996
GCCCCAGCA
22806
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23616


SpCas9-
CGTGCA

ACGGCCTG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACGGCCTGG



PB17













pU6_A1AT_
CCAGGC
21187
CCTTATGC
21997
GCCCCAGCA
22807
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23617


SpCas9-
CGTGCA

ACGGCCTG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACGGCCTG



PB16













pU6_A1AT_
CCAGGC
21188
CCTTATGC
21998
GCCCCAGCA
22808
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23618


SpCas9-
CGTGCA

ACGGCCT

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACGGCCT



PB15













pU6_A1AT_
CCAGGC
21189
CCTTATGC
21999
GCCCCAGCA
22809
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23619


SpCas9-
CGTGCA

ACGGCC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACGGCC



PB14













pU6_A1AT_
CCAGGC
21190
CCTTATGC
22000
GCCCCAGCA
22810
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23620


SpCas9-
CGTGCA

ACGGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACGGC



PB13













pU6_A1AT_
CCAGGC
21191
CCTTATGC
22001
GCCCCAGCA
22811
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23621


SpCas9-
CGTGCA

ACGG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACGG



PB12













pU6_A1AT_
CCAGGC
21192
CCTTATGC
22002
GCCCCAGCA
22812
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23622


SpCas9-
CGTGCA

ACG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCACG



PB11













pU6_A1AT_
CCAGGC
21193
CCTTATGC
22003
GCCCCAGCA
22813
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23623


SpCas9-
CGTGCA

AC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCAC



PB10













pU6_A1AT_
CCAGGC
21194
CCTTATGC

GCCCCAGCA
22814
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23624


SpCas9-
CGTGCA

A

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGCA



PB9













pU6_A1AT_
CCAGGC
21195
CCTTATGC

GCCCCAGCA
22815
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23625


SpCas9-
CGTGCA



GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



_25FE_




GCACAG

GTCAGCACAGCCTTATGC



PB8













pU6_A1AT_
CCAGGC
21196
CCTTATGC
22006
AGCAGCTTC
22816
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23626


SpCas9-
CGTGCA

ACGGCCTG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACGGCCTGG



PB17













pU6_A1AT_
CCAGGC
21197
CCTTATGC
22007
AGCAGCTTC
22817
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23627


SpCas9-
CGTGCA

ACGGCCTG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACGGCCTG



PB16













pU6_A1AT_
CCAGGC
21198
CCTTATGC
22008
AGCAGCTTC
22818
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23628


SpCas9-
CGTGCA

ACGGCCT

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACGGCCT



PB15













pU6_A1AT_
CCAGGC
21199
CCTTATGC
22009
AGCAGCTTC
22819
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23629


SpCas9-
CGTGCA

ACGGCC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACGGCC



PB14













pU6_A1AT_
CCAGGC
21200
CCTTATGC
22010
AGCAGCTTC
22820
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23630


SpCas9-
CGTGCA

ACGGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACGGC



PB13













pU6_A1AT_
CCAGGC
21201
CCTTATGC
22011
AGCAGCTTC
22821
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23631


SpCas9-
CGTGCA

ACGG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACGG



PB12













pU6_A1AT_
CCAGGC
21202
CCTTATGC
22012
AGCAGCTTC
22822
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23632


SpCas9-
CGTGCA

ACG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCACG



PB11













pU6_A1AT_
CCAGGC
21203
CCTTATGC
22013
AGCAGCTTC
22823
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23633


SpCas9-
CGTGCA

AC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCAC



PB10













pU6_A1AT_
CCAGGC
21204
CCTTATGC

AGCAGCTTC
22824
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23634


SpCas9-
CGTGCA

A

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGCA



PB9













pU6_A1AT_
CCAGGC
21205
CCTTATGC

AGCAGCTTC
22825
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23635


SpCas9-
CGTGCA



AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



_20FE_




AG

CACAGCCTTATGC



PB8













pU6_A1AT_
CCAGGC
21206
CCTTATGC
22016
TTCAGTCCC
22826
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23636


SpCas9-
CGTGCA

ACGGCCTG

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACGGCCTGG



PB17













pU6_A1AT_
CCAGGC
21207
CCTTATGC
22017
TTCAGTCCC
22827
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23637


SpCas9-
CGTGCA

ACGGCCTG

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACGGCCTG



PB16










pU6_A1AT_
CCAGGC
21208
CCTTATGC
22018
TTCAGTCCC
22828
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23638


SpCas9-
CGTGCA

ACGGCCT

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACGGCCT



PB15













pU6_A1AT_
CCAGGC
21209
CCTTATGC
22019
TTCAGTCCC
22829
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23639


SpCas9-
CGTGCA

ACGGCC

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACGGCC



PB14













pU6_A1AT_
CCAGGC
21210
CCTTATGC
22020
TTCAGTCCC
22830
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23640


SpCas9-
CGTGCA

ACGGC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACGGC



PB13













pU6_A1AT_
CCAGGC
21211
CCTTATGC
22021
TTCAGTCCC
22831
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23641


SpCas9-
CGTGCA

ACGG

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACGG



PB12













pU6_A1AT_
CCAGGC
21212
CCTTATGC
22022
TTCAGTCCC
22832
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23642


SpCas9-
CGTGCA

ACG

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCACG



PB11













pU6_A1AT_
CCAGGC
21213
CCTTATGC
22023
TTCAGTCCC
22833
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23643


SpCas9-
CGTGCA

AC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCAC



PB10













pU6_A1AT_
CCAGGC
21214
CCTTATGC

TTCAGTCCC
22834
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23644


SpCas9-
CGTGCA

A

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGCA



PB9













pU6_A1AT_
CCAGGC
21215
CCTTATGC

TTCAGTCCC
22835
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23645


SpCas9-
CGTGCA



TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



GCACAG

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



_14FE_






CTTATGC



PB8













pU6_A1AT_
CCAGGC
21216
CCTTATGC
22026
AGTCCCTTT
22836
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23646


SpCas9-
CGTGCA

ACGGCCTG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACGGCCTGG



PB17













pU6_A1AT_
CCAGGC
21217
CCTTATGC
22027
AGTCCCTTT
22837
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23647


SpCas9-
CGTGCA

ACGGCCTG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACGGCCTG



PB16













pU6_A1AT_
CCAGGC
21218
CCTTATGC
22028
AGTCCCTTT
22838
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23648


SpCas9-
CGTGCA

ACGGCCT

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACGGCCT



PB15













pU6_A1AT_
CCAGGC
21219
CCTTATGC
22029
AGTCCCTTT
22839
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23649


SpCas9-
CGTGCA

ACGGCC

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACGGCC



PB14













pU6_A1AT_
CCAGGC
21220
CCTTATGC
22030
AGTCCCTTT
22840
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23650


SpCas9-
CGTGCA

ACGGC

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACGGC



PB13













pU6_A1AT_
CCAGGC
21221
CCTTATGC
22031
AGTCCCTTT
22841
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23651


SpCas9-
CGTGCA

ACGG

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACGG



PB12













pU6_A1AT_
CCAGGC
21222
CCTTATGC
22032
AGTCCCTTT
22842
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23652


SpCas9-
CGTGCA

ACG

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCACG



PB11













pU6_A1AT_
CCAGGC
21223
CCTTATGC
22033
AGTCCCTTT
22843
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23653


SpCas9-
CGTGCA

AC

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCAC



PB10













pU6_A1AT_
CCAGGC
21224
CCTTATGC

AGTCCCTTT
22844
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23654


SpCas9-
CGTGCA

A

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGCA



PB9













pU6_A1AT_
CCAGGC
21225
CCTTATGC

AGTCCCTTT
22845
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23655


SpCas9-
CGTGCA



CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG



AG

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



_11FE_






ATGC



PB8













pU6_A1AT_
CCAGGC
21226
CCTTATGC
22036
TCCCTTTCTc
22846
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23656


SpCas9-
CGTGCA

ACGGCCTG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACGGCCTGG



PB17













pU6_A1AT_
CCAGGC
21227
CCTTATGC
22037
TCCCTTTCTc
22847
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23657


SpCas9-
CGTGCA

ACGGCCTG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACGGCCTG



PB16













pU6_A1AT_
CCAGGC
21228
CCTTATGC
22038
TCCCTTTCTc
22848
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23658


SpCas9-
CGTGCA

ACGGCCT

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACGGCCT



PB15













pU6_A1AT_
CCAGGC
21229
CCTTATGC
22039
TCCCTTTCTc
22849
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23659


SpCas9-
CGTGCA

ACGGCC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACGGCC



PB14













pU6_A1AT_
CCAGGC
21230
CCTTATGC
22040
TCCCTTTCTc
22850
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23660


SpCas9-
CGTGCA

ACGGC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACGGC



PB13













pU6_A1AT_
CCAGGC
21231
CCTTATGC
22041
TCCCTTTCTc
22851
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23661


SpCas9-
CGTGCA

ACGG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACGG



PB12













pU6_A1AT_
CCAGGC
21232
CCTTATGC
22042
TCCCTTTCTc
22852
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23662


SpCas9-
CGTGCA

ACG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCACG



PB11













pU6_A1AT_
CCAGGC
21233
CCTTATGC
22043
TCCCTTTCTc
22853
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23663


SpCas9-
CGTGCA

AC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCAC



PB10













pU6_A1AT_
CCAGGC
21234
CCTTATGC

TCCCTTTCTc
22854
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23664


SpCas9-
CGTGCA

A

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GCA



PB9













pU6_A1AT_
CCAGGC
21235
CCTTATGC

TCCCTTTCTc
22855
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23665


SpCas9-
CGTGCA



GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAGCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



_9FE_






GC



PB8













pU6_A1AT_
CCAGGC
21236
CCTTATGC
22046
CCTTTCTcGT
22856
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23666


SpCas9-
CGTGCA

ACGGCCTG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACGGCCTGG



PB17













pU6_A1AT_
CCAGGC
21237
CCTTATGC
22047
CCTTTCTcGT
22857
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23667


SpCas9-
CGTGCA

ACGGCCTG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACGGCCTG



PB16













pU6_A1AT_
CCAGGC
21238
CCTTATGC
22048
CCTTTCTcGT
22858
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23668


SpCas9-
CGTGCA

ACGGCCT

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACGGCCT



PB15













pU6_A1AT_
CCAGGC
21239
CCTTATGC
22049
CCTTTCTcGT
22859
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23669


SpCas9-
CGTGCA

ACGGCC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACGGCC



PB14













pU6_A1AT_
CCAGGC
21240
CCTTATGC
22050
CCTTTCTcGT
22860
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23670


SpCas9-
CGTGCA

ACGGC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACGGC



PB13













pU6_A1AT_
CCAGGC
21241
CCTTATGC
22051
CCTTTCTcGT
22861
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23671


SpCas9-
CGTGCA

ACGG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACGG



PB12













pU6_A1AT_
CCAGGC
21242
CCTTATGC
22052
CCTTTCTCGT
22862
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23672


SpCas9-
CGTGCA

ACG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






ACG



PB11













pU6_A1AT_
CCAGGC
21243
CCTTATGC
22053
CCTTTCTcGT
22863
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23673


SpCas9-
CGTGCA

AC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






AC



PB10













pU6_A1AT_
CCAGGC
21244
CCTTATGC

CCTTTCTcGT
22864
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23674


SpCas9-
CGTGCA

A

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_






A



PB9













pU6_A1AT_
CCAGGC
21245
CCTTATGC

CCTTTCTCGT
22865
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23675


SpCas9-
CGTGCA



CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



_7FE_










PB8













pU6_A1AT_
CCAGGC
21246
CCTTATGC
22056
TTTCTCGTC
22866
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23676


SpCas9-
CGTGCA

ACGGCCTG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






GGCCTGG



PB17













pU6_A1AT_
CCAGGC
21247
CCTTATGC
22057
TTTCTcGTC
22867
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23677


SpCas9-
CGTGCA

ACGGCCTG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






GGCCTG



PB16













pU6_A1AT_
CCAGGC
21248
CCTTATGC
22058
TTTCTCGTC
22868
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23678


SpCas9-
CGTGCA

ACGGCCT

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






GGCCT



PB15













pU6_A1AT_
CCAGGC
21249
CCTTATGC
22059
TTTCTCGTC
22869
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23679


SpCas9-
CGTGCA

ACGGCC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






GGCC



PB14













pU6_A1AT_
CCAGGC
21250
CCTTATGC
22060
TTTCTCGTC
22870
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23680


SpCas9-
CGTGCA

ACGGC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






GGC



PB13













pU6_A1AT_
CCAGGC
21251
CCTTATGC
22061
TTTCTCGTC
22871
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23681


SpCas9-
CGTGCA

ACGG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






GG



PB12













pU6_A1AT_
CCAGGC
21252
CCTTATGC
22062
TTTCTcGTC
22872
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23682


SpCas9-
CGTGCA

ACG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_






G



PB11













pU6_A1AT_
CCAGGC
21253
CCTTATGC
22063
TTTCTcGTC
22873
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23683


SpCas9-
CGTGCA

AC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



_5FE_










PB10













pU6_A1AT_
CCAGGC
21254
CCTTATGC

TTTCTCGTC
22874
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23684


SpCas9-
CGTGCA

A

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTCGTCGATGGTCAGCACAGCCTTATGCA



_5FE_










PB9













pU6_A1AT_
CCAGGC
21255
CCTTATGC

TTTCTCGTC
22875
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23685


SpCas9-
CGTGCA



GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



GCACAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TTTCTCGTCGATGGTCAGCACAGCCTTATGC



_5FE_










PB8













pU6_A1AT_
CCAGGC
21256
CCTTATGC
22066
TCTcGTCGA
22876
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23686


SpCas9-
CGTGCA

ACGGCCTG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC

G

CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_






GCCTGG



PB17













pU6_A1AT_
CCAGGC
21257
CCTTATGC
22067
TCTcGTCGA
22877
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23687


SpCas9-
CGTGCA

ACGGCCTG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_






GCCTG



PB16













pU6_A1AT_
CCAGGC
21258
CCTTATGC
22068
TCTcGTCGA
22878
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23688


SpCas9-
CGTGCA

ACGGCCT

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_






GCCT



PB15













pU6_A1AT_
CCAGGC
21259
CCTTATGC
22069
TCTcGTCGA
22879
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23689


SpCas9-
CGTGCA

ACGGCC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_






GCC



PB14













pU6_A1AT_
CCAGGC
21260
CCTTATGC
22070
TCTcGTCGA
22880
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23690


SpCas9-
CGTGCA

ACGGC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_






GC



PB13













pU6_A1AT_
CCAGGC
21261
CCTTATGC
22071
TCTcGTCGA
22881
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23691


SpCas9-
CGTGCA

ACGG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_






G



PB12













pU6_A1AT_
CCAGGC
21262
CCTTATGC
22072
TCTcGTCGA
22882
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23692


SpCas9-
CGTGCA

ACG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



_3FE_










PB11













pU6_A1AT_
CCAGGC
21263
CCTTATGC
22073
TCTcGTCGA
22883
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23693


SpCas9-
CGTGCA

AC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCAC



_3FE_










PB10













pU6_A1AT_
CCAGGC
21264
CCTTATGC

TCTcGTCGA
22884
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23694


SpCas9-
CGTGCA

A

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGCA



_3FE_










PB9













pU6_A1AT_
CCAGGC
21265
CCTTATGC

TCTcGTCGA
22885
CCAGGCCGTGCATAAGGCTGGTTTTAGAGCTA
23695


SpCas9-
CGTGCA



TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



NG_ED1
TAAGGC



CAG

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



7-
TG





TCTcGTCGATGGTCAGCACAGCCTTATGC



_3FE_










PB8













pU6_A1AT_
TCCAGG
21266
CTTATGCA
22076
ACATGGCCC
22886
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23696


SpRY_
CCGTGC

CGGCCTGG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB17




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCTGG








CAGC

A






pU6_A1AT_
TCCAGG
21267
CTTATGCA
22077
ACATGGCCC
22887
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23697


SpRY_
CCGTGC

CGGCCTGG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB16




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCTGG








CAGC








pU6_A1
TCCAGG
21268
CTTATGCA
22078
ACATGGCCC
22888
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23698


AT_SpR
CCGTGC

CGGCCTG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



Y_ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



G_30F
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



E_PB15




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCTG








CAGC








pU6_A1AT_
TCCAGG
21269
CTTATGCA
22079
ACATGGCCC
22889
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23699


SpRY_
CCGTGC

CGGCCT

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB14




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCCT








CAGC








pU6_A1AT_
TCCAGG
21270
CTTATGCA
22080
ACATGGCCC
22890
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23700


SpRY_
CCGTGC

CGGCC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB13




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGCC








CAGC








pU6_A1AT_
TCCAGG
21271
CTTATGCA
22081
ACATGGCCC
22891
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23701


SpRY_
CCGTGC

CGGC

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB12




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGGC








CAGC








pU6_A1AT_
TCCAGG
21272
CTTATGCA
22082
ACATGGCCC
22892
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23702


SpRY_
CCGTGC

CGG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB11




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACGG








CAGC








pU6_A1AT_
TCCAGG
21273
CTTATGCA
22083
ACATGGCCC
22893
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23703


SpRY_
CCGTGC

CG

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB10




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCACG








CAGC








pU6_A1AT_
TCCAGG
21274
CTTATGCA

ACATGGCCC
22894
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23704


SpRY_
CCGTGC

C

CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB9




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCAC








CAGC








pU6_A1AT_
TCCAGG
21275
CTTATGCA

ACATGGCCC
22895
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23705


SpRY_
CCGTGC



CAGCAGCTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGTCCCTT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_30FE_
CT



TCTcGTCGA

ACATGGCCCCAGCAGCTTCAGTCCCTTTCTcGT



PB8




TGGTCAGCA

CGATGGTCAGCACAGCCTTATGCA








CAGC








pU6_A1AT_
TCCAGG
21276
CTTATGCA
22086
GCCCCAGCA
22896
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23706


SpRY_
CCGTGC

CGGCCTGG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB17




GCACAGC

GTCAGCACAGCCTTATGCACGGCCTGGA






pU6_A1AT_
TCCAGG
21277
CTTATGCA
22087
GCCCCAGCA
22897
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23707


SpRY_
CCGTGC

CGGCCTGG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB16




GCACAGC

GTCAGCACAGCCTTATGCACGGCCTGG






pU6_A1AT_
TCCAGG
21278
CTTATGCA
22088
GCCCCAGCA
22898
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23708


SpRY_
CCGTGC

CGGCCTG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB15




GCACAGC

GTCAGCACAGCCTTATGCACGGCCTG






pU6_A1AT_
TCCAGG
21279
CTTATGCA
22089
GCCCCAGCA
22899
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23709


SpRY_
CCGTGC

CGGCCT

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB14




GCACAGC

GTCAGCACAGCCTTATGCACGGCCT






pU6_A1AT_
TCCAGG
21280
CTTATGCA
22090
GCCCCAGCA
22900
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23710


SpRY_
CCGTGC

CGGCC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB13




GCACAGC

GTCAGCACAGCCTTATGCACGGCC






pU6_A1AT_
TCCAGG
21281
CTTATGCA
22091
GCCCCAGCA
22901
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23711


SpRY_
CCGTGC

CGGC

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB12




GCACAGC

GTCAGCACAGCCTTATGCACGGC






pU6_A1AT_
TCCAGG
21282
CTTATGCA
22092
GCCCCAGCA
22902
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23712


SpRY_
CCGTGC

CGG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTCGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB11




GCACAGC

GTCAGCACAGCCTTATGCACGG






pU6_A1AT_
TCCAGG
21283
CTTATGCA
22093
GCCCCAGCA
22903
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23713


SpRY_
CCGTGC

CG

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB10




GCACAGC

GTCAGCACAGCCTTATGCACG






pU6_A1AT_
TCCAGG
21284
CTTATGCA

GCCCCAGCA
22904
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23714


SpRY_
CCGTGC

C

GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB9




GCACAGC

GTCAGCACAGCCTTATGCAC






pU6_A1AT_
TCCAGG
21285
CTTATGCA

GCCCCAGCA
22905
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23715


SpRY_
CCGTGC



GCTTCAGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CCTTTCTcGT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_25FE_
CT



CGATGGTCA

GCCCCAGCAGCTTCAGTCCCTTTCTcGTCGATG



PB8




GCACAGC

GTCAGCACAGCCTTATGCA






pU6_A1AT_
TCCAGG
21286
CTTATGCA
22096
AGCAGCTTC
22906
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23716


SpRY_
CCGTGC

CGGCCTGG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB17




AGC

CACAGCCTTATGCACGGCCTGGA






pU6_A1AT_
TCCAGG
21287
CTTATGCA
22097
AGCAGCTTC
22907
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23717


SpRY_
CCGTGC

CGGCCTGG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB16




AGC

CACAGCCTTATGCACGGCCTGG






pU6_A1AT_
TCCAGG
21288
CTTATGCA
22098
AGCAGCTTC
22908
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23718


SpRY_
CCGTGC

CGGCCTG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB15




AGC

CACAGCCTTATGCACGGCCTG






pU6_A1AT_
TCCAGG
21289
CTTATGCA
22099
AGCAGCTTC
22909
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23719


SpRY_
CCGTGC

CGGCCT

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTCGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB14




AGC

CACAGCCTTATGCACGGCCT






pU6_A1AT_
TCCAGG
21290
CTTATGCA
22100
AGCAGCTTC
22910
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23720


SpRY_
CCGTGC

CGGCC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB13




AGC

CACAGCCTTATGCACGGCC






pU6_A1AT_
TCCAGG
21291
CTTATGCA
22101
AGCAGCTTC
22911
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23721


SpRY_
CCGTGC

CGGC

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB12




AGC

CACAGCCTTATGCACGGC






pU6_A1AT_
TCCAGG
21292
CTTATGCA
22102
AGCAGCTTC
22912
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23722


SpRY_
CCGTGC

CGG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB11




AGC

CACAGCCTTATGCACGG






pU6_A1AT_
TCCAGG
21293
CTTATGCA
22103
AGCAGCTTC
22913
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23723


SpRY_
CCGTGC

CG

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB10




AGC

CACAGCCTTATGCACG






pU6_A1AT_
TCCAGG
21294
CTTATGCA

AGCAGCTTC
22914
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23724


SpRY_
CCGTGC

C

AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB9




AGC

CACAGCCTTATGCAC






pU6_A1AT_
TCCAGG
21295
CTTATGCA

AGCAGCTTC
22915
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23725


SpRY_
CCGTGC



AGTCCCTTT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CTcGTCGAT

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_20FE_
CT



GGTCAGCAC

AGCAGCTTCAGTCCCTTTCTcGTCGATGGTCAG



PB8




AGC

CACAGCCTTATGCA






pU6_A1AT_
TCCAGG
21296
CTTATGCA
22106
TTCAGTCCC
22916
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23726


SpRY_
CCGTGC

CGGCCTGG

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB17






CTTATGCACGGCCTGGA






pU6_A1AT_
TCCAGG
21297
CTTATGCA
22107
TTCAGTCCC
22917
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23727


SpRY_
CCGTGC

CGGCCTGG

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB16






CTTATGCACGGCCTGG






pU6_A1AT_
TCCAGG
21298
CTTATGCA
22108
TTCAGTCCC
22918
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23728


SpRY_
CCGTGC

CGGCCTG

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB15






CTTATGCACGGCCTG






pU6_A1AT_
TCCAGG
21299
CTTATGCA
22109
TTCAGTCCC
22919
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23729


SpRY_
CCGTGC

CGGCCT

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTCGTCGATGGTCAGCACAGC



PB14






CTTATGCACGGCCT






pU6_A1AT_
TCCAGG
21300
CTTATGCA
22110
TTCAGTCCC
22920
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23730


SpRY_
CCGTGC

CGGCC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB13






CTTATGCACGGCC






pU6_A1AT_
TCCAGG
21301
CTTATGCA
22111
TTCAGTCCC
22921
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23731


SpRY_
CCGTGC

CGGC

TTTCTcGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTCGTCGATGGTCAGCACAGC



PB12






CTTATGCACGGC






pU6_A1AT_
TCCAGG
21302
CTTATGCA
22112
TTCAGTCCC
22922
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23732


SpRY_
CCGTGC

CGG

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB11






CTTATGCACGG






pU6_A1AT_
TCCAGG
21303
CTTATGCA
22113
TTCAGTCCC
22923
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23733


SpRY_
CCGTGC

CG

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB10






CTTATGCACG






pU6_A1AT_
TCCAGG
21304
CTTATGCA

TTCAGTCCC
22924
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23734


SpRY_
CCGTGC

C

TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB9






CTTATGCAC






pU6_A1AT_
TCCAGG
21305
CTTATGCA

TTCAGTCCC
22925
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23735


SpRY_
CCGTGC



TTTCTCGTC

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GATGGTCA

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_14FE_
CT



GCACAGC

TTCAGTCCCTTTCTcGTCGATGGTCAGCACAGC



PB8






CTTATGCA






pU6_A1AT_
TCCAGG
21306
CTTATGCA
22116
AGTCCCTTT
22926
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23736


SpRY_
CCGTGC

CGGCCTGG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB17






ATGCACGGCCTGGA






pU6_A1AT_
TCCAGG
21307
CTTATGCA
22117
AGTCCCTTT
22927
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23737


SpRY_
CCGTGC

CGGCCTGG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB16






ATGCACGGCCTGG






pU6_A1AT_
TCCAGG
21308
CTTATGCA
22118
AGTCCCTTT
22928
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23738


SpRY_
CCGTGC

CGGCCTG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB15






ATGCACGGCCTG






pU6_A1AT_
TCCAGG
21309
CTTATGCA
22119
AGTCCCTTT
22929
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23739


SpRY_
CCGTGC

CGGCCT

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB14






ATGCACGGCCT






pU6_A1AT_
TCCAGG
21310
CTTATGCA
22120
AGTCCCTTT
22930
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23740


SpRY_
CCGTGC

CGGCC

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB13






ATGCACGGCC






pU6_A1AT_
TCCAGG
21311
CTTATGCA
22121
AGTCCCTTT
22931
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23741


SpRY_
CCGTGC

CGGC

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB12






ATGCACGGC






pU6_A1AT_
TCCAGG
21312
CTTATGCA
22122
AGTCCCTTT
22932
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23742


SpRY_
CCGTGC

CGG

CTCGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB11






ATGCACGG






pU6_A1AT_
TCCAGG
21313
CTTATGCA
22123
AGTCCCTTT
22933
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23743


SpRY_
CCGTGC

CG

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB10






ATGCACG






pU6_A1AT_
TCCAGG
21314
CTTATGCA

AGTCCCTTT
22934
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23744


SpRY_
CCGTGC

C

CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB9






ATGCAC






pU6_A1AT_
TCCAGG
21315
CTTATGCA

AGTCCCTTT
22935
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23745


SpRY_
CCGTGC



CTcGTCGAT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GGTCAGCAC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_11FE_
CT



AGC

AGTCCCTTTCTcGTCGATGGTCAGCACAGCCTT



PB8






ATGCA






pU6_A1AT_
TCCAGG
21316
CTTATGCA
22126
TCCCTTTCTc
22936
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23746


SpRY_
CCGTGC

CGGCCTGG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB17






GCACGGCCTGGA






pU6_A1AT_
TCCAGG
21317
CTTATGCA
22127
TCCCTTTCTc
22937
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23747


SpRY_
CCGTGC

CGGCCTGG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB16






GCACGGCCTGG






pU6_A1AT_
TCCAGG
21318
CTTATGCA
22128
TCCCTTTCTc
22938
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23748


SpRY_
CCGTGC

CGGCCTG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB15






GCACGGCCTG






pU6_A1AT_
TCCAGG
21319
CTTATGCA
22129
TCCCTTTCTc
22939
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23749


SpRY_
CCGTGC

CGGCCT

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB14






GCACGGCCT






pU6_A1AT_
TCCAGG
21320
CTTATGCA
22130
TCCCTTTCTc
22940
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23750


SpRY_
CCGTGC

CGGCC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB13






GCACGGCC






pU6_A1AT_
TCCAGG
21321
CTTATGCA
22131
TCCCTTTCTc
22941
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23751


SpRY_
CCGTGC

CGGC

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB12






GCACGGC






pU6_A1AT_
TCCAGG
21322
CTTATGCA
22132
TCCCTTTCTc
22942
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23752


SpRY_
CCGTGC

CGG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB11






GCACGG






pU6_A1AT_
TCCAGG
21323
CTTATGCA
22133
TCCCTTTCTc
22943
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23753


SpRY_
CCGTGC

CG

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB10






GCACG






pU6_A1AT_
TCCAGG
21324
CTTATGCA

TCCCTTTCTc
22944
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23754


SpRY_
CCGTGC

C

GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB9






GCAC






pU6_A1AT_
TCCAGG
21325
CTTATGCA

TCCCTTTCTc
22945
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23755


SpRY_
CCGTGC



GTCGATGGT

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_9FE_
CT





TCCCTTTCTcGTCGATGGTCAGCACAGCCTTAT



PB8






GCA






pU6_A1AT_
TCCAGG
21326
CTTATGCA
22136
CCTTTCTCGT
22946
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23756


SpRY_
CCGTGC

CGGCCTGG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB17






ACGGCCTGGA






pU6_A1AT_
TCCAGG
21327
CTTATGCA
22137
CCTTTCTcGT
22947
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23757


SpRY_
CCGTGC

CGGCCTGG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB16






ACGGCCTGG






pU6_A1AT_
TCCAGG
21328
CTTATGCA
22138
CCTTTCTcGT
22948
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23758


SpRY_
CCGTGC

CGGCCTG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB15






ACGGCCTG






pU6_A1AT_
TCCAGG
21329
CTTATGCA
22139
CCTTTCTCGT
22949
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23759


SpRY_
CCGTGC

CGGCCT

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB14






ACGGCCT






pU6_A1AT_
TCCAGG
21330
CTTATGCA
22140
CCTTTCTcGT
22950
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23760


SpRY_
CCGTGC

CGGCC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB13






ACGGCC






pU6_A1AT_
TCCAGG
21331
CTTATGCA
22141
CCTTTCTcGT
22951
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23761


SpRY_
CCGTGC

CGGC

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB12






ACGGC






pU6_A1AT_
TCCAGG
21332
CTTATGCA
22142
CCTTTCTcGT
22952
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23762


SpRY_
CCGTGC

CGG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB11






ACGG






pU6_A1AT_
TCCAGG
21333
CTTATGCA
22143
CCTTTCTcGT
22953
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23763


SpRY_
CCGTGC

CG

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB10






ACG






pU6_A1AT_
TCCAGG
21334
CTTATGCA

CCTTTCTcGT
22954
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23764


SpRY_
CCGTGC

C

CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB9






AC






pU6_A1AT_
TCCAGG
21335
CTTATGCA

CCTTTCTcGT
22955
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23765


SpRY_
CCGTGC



CGATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_7FE_
CT





CCTTTCTcGTCGATGGTCAGCACAGCCTTATGC



PB8






A






pU6_A1AT_
TCCAGG
21336
CTTATGCA
22146
TTTCTCGTC
22956
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23766


SpRY_
CCGTGC

CGGCCTGG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



PB17






GGCCTGGA






pU6_A1AT_
TCCAGG
21337
CTTATGCA
22147
TTTCTCGTC
22957
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23767


SpRY_
CCGTGC

CGGCCTGG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



PB16






GGCCTGG






pU6_A1AT_
TCCAGG
21338
CTTATGCA
22148
TTTCTCGTC
22958
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23768


SpRY_
CCGTGC

CGGCCTG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



PB15






GGCCTG






pU6_A1AT_
TCCAGG
21339
CTTATGCA
22149
TTTCTcGTC
22959
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23769


SpRY_
CCGTGC

CGGCCT

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



PB14






GGCCT






pU6_A1AT_
TCCAGG
21340
CTTATGCA
22150
TTTCTcGTC
22960
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23770


SpRY_
CCGTGC

CGGCC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



PB13






GGCC






pU6_A1AT_
TCCAGG
21341
CTTATGCA
22151
TTTCTCGTC
22961
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23771


SpRY_
CCGTGC

CGGC

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



PB12






GGC






pU6_A1AT_
TCCAGG
21342
CTTATGCA
22152
TTTCTcGTC
22962
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23772


SpRY_
CCGTGC

CGG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



PB11






GG






pU6_A1AT_
TCCAGG
21343
CTTATGCA
22153
TTTCTCGTC
22963
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23773


SpRY_
CCGTGC

CG

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTcGTCGATGGTCAGCACAGCCTTATGCAC



PB10






G






pU6_A1AT_
TCCAGG
21344
CTTATGCA

TTTCTcGTC
22964
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23774


SpRY_
CCGTGC

C

GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTCGTCGATGGTCAGCACAGCCTTATGCAC



PB9













pU6_A1AT_
TCCAGG
21345
CTTATGCA

TTTCTcGTC
22965
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23775


SpRY_
CCGTGC



GATGGTCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



GCACAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_5FE_
CT





TTTCTCGTCGATGGTCAGCACAGCCTTATGCA



PB8













pU6_A1AT_
TCCAGG
21346
CTTATGCA
22156
TCTcGTCGA
22966
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23776


SpRY_
CCGTGC

CGGCCTGG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG

A

CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB17






GCCTGGA






pU6_A1AT_
TCCAGG
21347
CTTATGCA
22157
TCTcGTCGA
22967
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23777


SpRY_
CCGTGC

CGGCCTGG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB16






GCCTGG






pU6_A1AT_
TCCAGG
21348
CTTATGCA
22158
TCTcGTCGA
22968
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23778


SpRY_
CCGTGC

CGGCCTG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB15






GCCTG






pU6_A1AT_
TCCAGG
21349
CTTATGCA
22159
TCTcGTCGA
22969
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23779


SpRY_
CCGTGC

CGGCCT

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB14






GCCT






pU6_A1AT_
TCCAGG
21350
CTTATGCA
22160
TCTcGTCGA
22970
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23780


SpRY_
CCGTGC

CGGCC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB13






GCC






pU6_A1AT_
TCCAGG
21351
CTTATGCA
22161
TCTcGTCGA
22971
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23781


SpRY_
CCGTGC

CGGC

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB12






GC






pU6_A1AT_
TCCAGG
21352
CTTATGCA
22162
TCTcGTCGA
22972
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23782


SpRY_
CCGTGC

CGG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB11






G






pU6_A1AT_
TCCAGG
21353
CTTATGCA
22163
TCTcGTCGA
22973
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23783


SpRY_
CCGTGC

CG

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCACG



PB10













pU6_A1AT_
TCCAGG
21354
CTTATGCA

TCTcGTCGA
22974
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23784


SpRY_
CCGTGC

C

TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCAC



PB9













pU6_A1AT_
TCCAGG
21355
CTTATGCA

TCTcGTCGA
22975
TCCAGGCCGTGCATAAGGCTGTTTTAGAGCTA
23785


SpRY_
CCGTGC



TGGTCAGCA

GAAATAGCAAGTTAAAATAAGGCTAGTCCGTT



ED18-
ATAAGG



CAGC

ATCAACTTGAAAAAGTGGCACCGAGTCGGTGC



_G_3FE_
CT





TCTcGTCGATGGTCAGCACAGCCTTATGCA



PB8









The template RNA sequences shown in Tables 1-4, 5, 6A, and 6B may be customized depending on the cell being targeted. For example, in some embodiments it is desired to inactivate a PAM sequence upon editing (e.g., using a “PAM-kill” modification) to decrease the potential for further gene editing (e.g., by Cas retargeting) following the initial edit. Consequently, certain template RNAs described herein are designed to write a mutation (e.g., a substitution) into the PAM of the target site, such that upon editing, the PAM site will be mutated to a sequence no longer recognized by the gene modifying polypeptide. Thus, a mutation region within the heterologous object sequence of the template RNA may comprise a PAM-kill sequence. Without wishing to be bound by theory, in some embodiments, a PAM-kill sequence prevents re-engagement of the gene modifying polypeptide upon completion of a gene modification, or decreases re-engagement relative to a template RNA lacking a PAM-kill sequence. In some embodiments, a PAM-kill sequence does not alter the amino acid sequence encoded by a gene, e.g., the PAM-kill sequence results in a silent mutation. In other embodiments, it is desired to leave the PAM sequence intact (no PAM-kill).


Similarly, in some embodiments, to decrease the potential for further gene editing (e.g., by Cas retargeting) following the initial edit, it may be desirable to alter the first three nucleotides of the RT template sequence via a “seed-kill” motif. Consequently, certain template RNAs described herein are designed to write a mutation (e.g., a substitution) into the portion of the target site corresponding to the first three nucleotides of the RT template sequence, such that upon editing, the target site will be mutated to a sequence with lower homology to the RT template sequence. Thus, a mutation region within the heterologous object sequence of the template RNA may comprise a seed-kill sequence. Without wishing to be bound by theory, in some embodiments, a seed-kill sequence prevents re-engagement of the gene modifying polypeptide upon completion of genetic modification, or decreases re-engagement relative to an otherwise similar template RNA lacking a seed-kill sequence. In some embodiments, a seed-kill sequence does not alter the amino acid sequence encoded by a gene, e.g., the seed-kill sequence results in a silent mutation. In other embodiments, it is desired to leave the seed region intact, and a seed-kill sequence is not used.


In further embodiments, to optimize or improve gene editing efficiency, it may be desirable to evade the target cell's mismatch repair or nucleotide repair pathways or to bias the target cell's repair pathways toward preservation of the edited strand. In some embodiments, multiple silent mutations (for example, silent substitutions) may be introduced within the RT template sequence to evade the target cell's mismatch repair or nucleotide repair pathways or to bias the target cell's repair pathways toward preservation of the edited strand.


Table 7B provides exemplary silent mutations for various positions within the SERPINA1 gene.









TABLE 7B







Exemplary Silent Mutation Codons for the SERPINA1 Gene















Amino Acid










Position
WT









(counting
Amino
WT
















initial Met)
Acid
Codon
ALL_CODONS


















356
A
GCC
GCT
GCC
GCA
GCG




357
V
GTG
GTT
GTC
GTA
GTG




358
H
CAT
CAT
CAC






359
K
AAG
AAA
AAG






360
A
GCT
GCT
GCC
GCA
GCG




361
V
GTG
GTT
GTC
GTA
GTG




362
L
CTG
TTA
TTG
CTT
CTC
CTA
CTG


363
T
ACC
ACT
ACC
ACA
ACG




364
I
ATC
ATA
ATT
ATC





365
D
GAC
GAT
GAC






367
K
AAA
AAA
AAG






368
G
GGG
GGT
GGC
GGA
GGG




369
T
ACT
ACT
ACC
ACA
ACG




370
E
GAA
GAA
GAG






371
A
GCT
GCT
GCC
GCA
GCG




372
A
GCT
GCT
GCC
GCA
GCG




373
G
GGG
GGT
GGC
GGA
GGG




374
A
GCC
GCT
GCC
GCA
GCG




375
M
ATG
ATG







376
F
TTT
TTT
TTC






377
L
TTA
TTA
TTG
CTT
CTC
CTA
CTG


378
E
GAG
GAA
GAG






379
A
GCC
GCT
GCC
GCA
GCG




380
I
ATA
ATA
ATT
ATC





381
P
CCC
CCT
CCC
CCA
CCG




382
M
ATG
ATG







383
S
TCT
TCT
TCC
TCA
TCG
AGT
AGC


384
I
ATC
ATA
ATT
ATC





385
P
CCC
CCT
CCC
CCA
CCG




386
P
CCC
CCT
CCC
CCA
CCG




387
E
GAG
GAA
GAG






388
V
GTC
GTT
GTC
GTA
GTG




389
K
AAG
AAA
AAG






390
F
TTC
TTT
TTC






391
N
AAC
AAT
AAC






392
K
AAA
AAA
AAG






393
P
CCC
CCT
CCC
CCA
CCG




394
F
TTT
TTT
TTC






395
V
GTC
GTT
GTC
GTA
GTG




396
F
TTC
TTT
TTC






397
L
TTA
TTA
TTG
CTT
CTC
CTA
CTG


398
M
ATG
ATG







399
I
ATT
ATA
ATT
ATC





400
E
GAA
GAA
GAG






401
Q
CAA
CAA
CAG






402
N
AAT
AAT
AAC






403
T
ACC
ACT
ACC
ACA
ACG




404
K
AAG
AAA
AAG






405
S
TCT
TCT
TCC
TCA
TCG
AGT
AGC


406
P
CCC
CCT
CCC
CCA
CCG




407
L
CTC
TTA
TTG
CTT
CTC
CTA
CTG


408
F
TTC
TTT
TTC






409
M
ATG
ATG







410
G
GGA
GGT
GGC
GGA
GGG




411
K
AAA
AAA
AAG






412
V
GTG
GTT
GTC
GTA
GTG




413
V
GTG
GTT
GTC
GTA
GTG




414
N
AAT
AAT
AAC






415
P
CCC
CCT
CCC
CCA
CCG




416
T
ACC
ACT
ACC
ACA
ACG




417
Q
CAA
CAA
CAG






418
K
AAA
AAA
AAG






419
*
TAA
TAA
TAG
TGA









In some embodiments, the template RNA comprises one or more silent mutations.


It should be understood that the silent mutations illustrated in Table 7B may be used individually or combined in any manner in a template RNA sequence described herein.


gRNAs with Inducible Activity


In some embodiments, a gRNA described herein (e.g., a gRNA that is part of a template RNA or a gRNA used for second strand nicking) has inducible activity. Inducible activity may be achieved by the template nucleic acid, e.g., template RNA, further comprising (in addition to the gRNA) a blocking domain, wherein the sequence of a portion of or all of the blocking domain is at least partially complementary to a portion or all of the gRNA. The blocking domain is thus capable of hybridizing or substantially hybridizing to a portion of or all of the gRNA. In some embodiments, the blocking domain and inducibly active gRNA are disposed on the template nucleic acid, e.g., template RNA, such that the gRNA can adopt a first conformation where the blocking domain is hybridized or substantially hybridized to the gRNA, and a second conformation where the blocking domain is not hybridized or not substantially hybridized to the gRNA. In some embodiments, in the first conformation the gRNA is unable to bind to the gene modifying polypeptide (e.g., the template nucleic acid binding domain, DNA binding domain, or endonuclease domain (e.g., a CRISPR/Cas protein)) or binds with substantially decreased affinity compared to an otherwise similar template RNA lacking the blocking domain. In some embodiments, in the second conformation the gRNA is able to bind to the gene modifying polypeptide (e.g., the template nucleic acid binding domain, DNA binding domain, or endonuclease domain (e.g., a CRISPR/Cas protein)). In some embodiments, whether the gRNA is in the first or second conformation can influence whether the DNA binding or endonuclease activities of the gene modifying polypeptide (e.g., of the CRISPR/Cas protein the gene modifying polypeptide comprises) are active.


In some embodiments, the gRNA that coordinates the second nick has inducible activity. In some embodiments, the gRNA that coordinates the second nick is induced after the template is reverse transcribed. In some embodiments, hybridization of the gRNA to the blocking domain can be disrupted using an opener molecule. In some embodiments, an opener molecule comprises an agent that binds to a portion or all of the gRNA or blocking domain and inhibits hybridization of the gRNA to the blocking domain. In some embodiments, the opener molecule comprises a nucleic acid, e.g., comprising a sequence that is partially or wholly complementary to the gRNA, blocking domain, or both. By choosing or designing an appropriate opener molecule, providing the opener molecule can promote a change in the conformation of the gRNA such that it can associate with a CRISPR/Cas protein and provide the associated functions of the CRISPR/Cas protein (e.g., DNA binding and/or endonuclease activity). Without wishing to be bound by theory, providing the opener molecule at a selected time and/or location may allow for spatial and temporal control of the activity of the gRNA, CRISPR/Cas protein, or gene modifying system comprising the same. In some embodiments, the opener molecule is exogenous to the cell comprising the gene modifying polypeptide and or template nucleic acid. In some embodiments, the opener molecule comprises an endogenous agent (e.g., endogenous to the cell comprising the gene modifying polypeptide and or template nucleic acid comprising the gRNA and blocking domain). For example, an inducible gRNA, blocking domain, and opener molecule may be chosen such that the opener molecule is an endogenous agent expressed in a target cell or tissue, e.g., thereby ensuring activity of a gene modifying system in the target cell or tissue. As a further example, an inducible gRNA, blocking domain, and opener molecule may be chosen such that the opener molecule is absent or not substantially expressed in one or more non-target cells or tissues, e.g., thereby ensuring that activity of a gene modifying system does not occur or substantially occur in the one or more non-target cells or tissues, or occurs at a reduced level compared to a target cell or tissue. Exemplary blocking domains, opener molecules, and uses thereof are described in PCT App. Publication WO2020044039A1, which is incorporated herein by reference in its entirety. In some embodiments, the template nucleic acid, e.g., template RNA, may comprise one or more sequences or structures for binding by one or more components of a gene modifying polypeptide, e.g., by a reverse transcriptase or RNA binding domain, and a gRNA. In some embodiments, the gRNA facilitates interaction with the template nucleic acid binding domain (e.g., RNA binding domain) of the gene modifying polypeptide. In some embodiments, the gRNA directs the gene modifying polypeptide to the matching target sequence, e.g., in a target cell genome.


Circular RNAs and Ribozymes in Gene Modifying Systems

It is contemplated that it may be useful to employ circular and/or linear RNA states during the formulation, delivery, or gene modifying reaction within the target cell. Thus, in some embodiments of any of the aspects described herein, a gene modifying system comprises one or more circular RNAs (circRNAs). In some embodiments of any of the aspects described herein, a gene modifying system comprises one or more linear RNAs. In some embodiments, a nucleic acid as described herein (e.g., a template nucleic acid, a nucleic acid molecule encoding a gene modifying polypeptide, or both) is a circRNA. In some embodiments, a circular RNA molecule encodes the gene modifying polypeptide. In some embodiments, the circRNA molecule encoding the gene modifying polypeptide is delivered to a host cell. In some embodiments, a circular RNA molecule encodes a recombinase, e.g., as described herein. In some embodiments, the circRNA molecule encoding the recombinase is delivered to a host cell. In some embodiments, the circRNA molecule encoding the gene modifying polypeptide is linearized (e.g., in the host cell, e.g., in the nucleus of the host cell) prior to translation.


Circular RNAs (circRNAs) have been found to occur naturally in cells and have been found to have diverse functions, including both non-coding and protein coding roles in human cells. It has been shown that a circRNA can be engineered by incorporating a self-splicing intron into an RNA molecule (or DNA encoding the RNA molecule) that results in circularization of the RNA, and that an engineered circRNA can have enhanced protein production and stability (Wesselhoeft et al. Nature Communications 2018). In some embodiments, the gene modifying polypeptide is encoded as circRNA. In certain embodiments, the template nucleic acid is a DNA, such as a dsDNA or ssDNA. In certain embodiments, the circDNA comprises a template RNA.


In some embodiments, the circRNA comprises one or more ribozyme sequences. In some embodiments, the ribozyme sequence is activated for autocleavage, e.g., in a host cell, e.g., thereby resulting in linearization of the circRNA. In some embodiments, the ribozyme is activated when the concentration of magnesium reaches a sufficient level for cleavage, e.g., in a host cell. In some embodiments the circRNA is maintained in a low magnesium environment prior to delivery to the host cell. In some embodiments, the ribozyme is a protein-responsive ribozyme. In some embodiments, the ribozyme is a nucleic acid-responsive ribozyme. In some embodiments, the circRNA comprises a cleavage site. In some embodiments, the circRNA comprises a second cleavage site.


In some embodiments, the circRNA is linearized in the nucleus of a target cell. In some embodiments, linearization of a circRNA in the nucleus of a cell involves components present in the nucleus of the cell, e.g., to activate a cleavage event. In some embodiments, a ribozyme, e.g., a ribozyme from a B2 or ALU element, that is responsive to a nuclear element, e.g., a nuclear protein, e.g., a genome-interacting protein, e.g., an epigenetic modifier, e.g., EZH2, is incorporated into a circRNA, e.g., of a gene modifying system. In some embodiments, nuclear localization of the circRNA results in an increase in autocatalytic activity of the ribozyme and linearization of the circRNA.


In some embodiments, the ribozyme is heterologous to one or more of the other components of the gene modifying system. In some embodiments, an inducible ribozyme (e.g., in a circRNA as described herein) is created synthetically, for example, by utilizing a protein ligand-responsive aptamer design. A system for utilizing the satellite RNA of tobacco ringspot virus hammerhead ribozyme with an MS2 coat protein aptamer has been described (Kennedy et al. Nucleic Acids Res 42(19):12306-12321 (2014), incorporated herein by reference in its entirety) that results in activation of the ribozyme activity in the presence of the MS2 coat protein. In embodiments, such a system responds to protein ligand localized to the cytoplasm or the nucleus. In some embodiments the protein ligand is not MS2. Methods for generating RNA aptamers to target ligands have been described, for example, based on the systematic evolution of ligands by exponential enrichment (SELEX) (Tuerk and Gold, Science 249(4968):505-510 (1990); Ellington and Szostak, Nature 346(6287):818-822 (1990); the methods of each of which are incorporated herein by reference) and have, in some instances, been aided by in silico design (Bell et al. PNAS 117(15):8486-8493, the methods of which are incorporated herein by reference). Thus, in some embodiments, an aptamer for a target ligand is generated and incorporated into a synthetic ribozyme system, e.g., to trigger ribozyme-mediated cleavage and circRNA linearization, e.g., in the presence of the protein ligand. In some embodiments, circRNA linearization is triggered in the cytoplasm, e.g., using an aptamer that associates with a ligand in the cytoplasm. In some embodiments, circRNA linearization is triggered in the nucleus, e.g., using an aptamer that associates with a ligand in the nucleus. In embodiments, the ligand in the nucleus comprises an epigenetic modifier or a transcription factor. In some embodiments the ligand that triggers linearization is present at higher levels in on-target cells than off-target cells.


It is further contemplated that a nucleic acid-responsive ribozyme system can be employed for circRNA linearization. For example, biosensors that sense defined target nucleic acid molecules to trigger ribozyme activation are described, e.g., in Penchovsky (Biotechnology Advances 32(5):1015-1027 (2014), incorporated herein by reference). By these methods, a ribozyme naturally folds into an inactive state and is only activated in the presence of a defined target nucleic acid molecule (e.g., an RNA molecule). In some embodiments, a circRNA of a gene modifying system comprises a nucleic acid-responsive ribozyme that is activated in the presence of a defined target nucleic acid, e.g., an RNA, e.g., an mRNA, miRNA, guide RNA, gRNA, sgRNA, ncRNA, lncRNA, tRNA, snRNA, or mtRNA. In some embodiments the nucleic acid that triggers linearization is present at higher levels in on-target cells than off-target cells.


In some embodiments of any of the aspects herein, a gene modifying system incorporates one or more ribozymes with inducible specificity to a target tissue or target cell of interest, e.g., a ribozyme that is activated by a ligand or nucleic acid present at higher levels in a target tissue or target cell of interest. In some embodiments, the gene modifying system incorporates a ribozyme with inducible specificity to a subcellular compartment, e.g., the nucleus, nucleolus, cytoplasm, or mitochondria. In some embodiments, the ribozyme that is activated by a ligand or nucleic acid present at higher levels in the target subcellular compartment. In some embodiments, an RNA component of a gene modifying system is provided as circRNA, e.g., that is activated by linearization. In some embodiments, linearization of a circRNA encoding a gene modifying polypeptide activates the molecule for translation. In some embodiments, a signal that activates a circRNA component of a gene modifying system is present at higher levels in on-target cells or tissues, e.g., such that the system is specifically activated in these cells.


In some embodiments, an RNA component of a gene modifying system is provided as a circRNA that is inactivated by linearization. In some embodiments, a circRNA encoding the gene modifying polypeptide is inactivated by cleavage and degradation. In some embodiments, a circRNA encoding the gene modifying polypeptide is inactivated by cleavage that separates a translation signal from the coding sequence of the polypeptide. In some embodiments, a signal that inactivates a circRNA component of a gene modifying system is present at higher levels in off-target cells or tissues, such that the system is specifically inactivated in these cells.


Target Nucleic Acid Site

In some embodiments, after gene modification, the target site surrounding the edited sequence contains a limited number of insertions or deletions, for example, in less than about 50% or 10% of editing events, e.g., as determined by long-read amplicon sequencing of the target site, e.g., as described in Karst et al. (2020) bioRxiv doi.org/10.1101/645903 (incorporated by reference herein in its entirety). In some embodiments, the target site does not show multiple consecutive editing events, e.g., head-to-tail or head-to-head duplications, e.g., as determined by long-read amplicon sequencing of the target site, e.g., as described in Karst et al. bioRxiv doi.org/10.1101/645903 (2020) (incorporated herein by reference in its entirety). In some embodiments, the target site contains an integrated sequence corresponding to the template RNA. In some embodiments, the target site does not contain insertions resulting from endogenous RNA in more than about 1% or 10% of events, e.g., as determined by long-read amplicon sequencing of the target site, e.g., as described in Karst et al. bioRxiv doi.org/10.1101/645903 (2020) (incorporated herein by reference in its entirety). In some embodiments, the target site contains the integrated sequence corresponding to the template RNA.


In certain aspects of the present invention, the host DNA-binding site integrated into by the gene modifying system can be in a gene, in an intron, in an exon, an ORF, outside of a coding region of any gene, in a regulatory region of a gene, or outside of a regulatory region of a gene. In other aspects, the polypeptide may bind to one or more than one host DNA sequence.


In some embodiments, a gene modifying system is used to edit a target locus in multiple alleles. In some embodiments, a gene modifying system is designed to edit a specific allele. For example, a gene modifying polypeptide may be directed to a specific sequence that is only present on one allele, e.g., comprises a template RNA with homology to a target allele, e.g., a gRNA or annealing domain, but not to a second cognate allele. In some embodiments, a gene modifying system can alter a haplotype-specific allele. In some embodiments, a gene modifying system that targets a specific allele preferentially targets that allele, e.g., has at least a 2, 4, 6, 8, or 10-fold preference for a target allele.


Second Strand Nicking

In some embodiments, a gene modifying system described herein comprises a nickase activity (e.g., in the gene modifying polypeptide) that nicks the first strand, and a nickase activity (e.g., in a polypeptide separate from the gene modifying polypeptide) that nicks the second strand of target DNA. As discussed herein, without wishing to be bound by theory, nicking of the first strand of the target site DNA is thought to provide a 3′ OH that can be used by an RT domain to reverse transcribe a sequence of a template RNA, e.g., a heterologous object sequence. Without wishing to be bound by theory, it is thought that introducing an additional nick to the second strand may bias the cellular DNA repair machinery to adopt the heterologous object sequence-based sequence more frequently than the original genomic sequence. In some embodiments, the additional nick to the second strand is made by the same endonuclease domain (e.g., nickase domain) as the nick to the first strand. In some embodiments, the same gene modifying polypeptide performs both the nick to the first strand and the nick to the second strand. In some embodiments, the gene modifying polypeptide comprises a CRISPR/Cas domain and the additional nick to the second strand is directed by an additional nucleic acid, e.g., comprising a second gRNA directing the CRISPR/Cas domain to nick the second strand. In other embodiments, the additional second strand nick is made by a different endonuclease domain (e.g., nickase domain) than the nick to the first strand. In some embodiments, that different endonuclease domain is situated in an additional polypeptide (e.g., a system of the invention further comprises the additional polypeptide), separate from the gene modifying polypeptide. In some embodiments, the additional polypeptide comprises an endonuclease domain (e.g., nickase domain) described herein. In some embodiments, the additional polypeptide comprises a DNA binding domain, e.g., described herein.


It is contemplated herein that the position at which the second strand nick occurs relative to the first strand nick may influence the extent to which one or more of: desired gene modifying DNA modifications are obtained, undesired double-strand breaks (DSBs) occur, undesired insertions occur, or undesired deletions occur. Without wishing to be bound by theory, second strand nicking may occur in two general orientations: inward nicks and outward nicks.


In some embodiments, in the inward nick orientation, the RT domain polymerizes (e.g., using the template RNA (e.g., the heterologous object sequence)) away from the second strand nick. In some embodiments, in the inward nick orientation, the location of the nick to the first strand and the location of the nick to the second strand are positioned between the first PAM site and second PAM site (e.g., in a scenario wherein both nicks are made by a polypeptide (e.g., a gene modifying polypeptide) comprising a CRISPR/Cas domain). When there are two PAMs on the outside and two nicks on the inside, this inward nick orientation can also be referred to as “PAM-out”. In some embodiments, in the inward nick orientation, the location of the nick to the first strand and the location of the nick to the second strand are between the sites where the polypeptide and the additional polypeptide bind to the target DNA. In some embodiments, in the inward nick orientation, the location of the nick to the second strand is positioned between the binding sites of the polypeptide and additional polypeptide, and the nick to the first strand is also located between the binding sites of the polypeptide and additional polypeptide. In some embodiments, in the inward nick orientation, the location of the nick to the first strand and the location of the nick to the second strand are positioned between the PAM site and the binding site of the second polypeptide which is at a distance from the target site. An example of a gene modifying system that provides an inward nick orientation comprises a gene modifying polypeptide comprising a CRISPR/Cas domain, a template RNA comprising a gRNA that directs nicking of the target site DNA on the first strand, and an additional nucleic acid comprising an additional gRNA that directs nicking at a site a distance from the location of the first nick, wherein the location of the first nick and the location of the second nick are between the PAM sites of the sites to which the two gRNAs direct the gene modifying polypeptide. As a further example, another gene modifying system that provides an inward nick orientation comprises a gene modifying polypeptide comprising a zinc finger molecule and a first nickase domain wherein the zinc finger molecule binds to the target DNA in a manner that directs the first nickase domain to nick the first strand of the target site; an additional polypeptide comprising a CRISPR/Cas domain, and an additional nucleic acid comprising a gRNA that directs the additional polypeptide to nick a site a distance from the target site DNA on the second strand, wherein the location of the first nick and the location of the second nick are between the PAM site and the site to which the zinc finger molecule binds. As a further example, another gene modifying system that provides an inward nick orientation comprises a gene modifying polypeptide comprising a zinc finger molecule and a first nickase domain wherein the zinc finger molecule binds to the target DNA in a manner that directs the first nickase domain to nick the first strand of the target site; an additional polypeptide comprising a TAL effector molecule and a second nickase domain wherein the TAL effector molecule binds to a site a distance from the target site in a manner that directs the additional polypeptide to nick the second strand, wherein the location of the first nick and the location of the second nick are between the site to which the TAL effector molecule binds and the site to which the zinc finger molecule binds.


In some embodiments, in the outward nick orientation, the RT domain polymerizes (e.g., using the template RNA (e.g., the heterologous object sequence)) toward the second strand nick. In some embodiments, in the outward nick orientation when both the first and second nicks are made by a polypeptide comprising a CRISPR/Cas domain (e.g., a gene modifying polypeptide), the first PAM site and second PAM site are positioned between the location of the nick to the first strand and the location of the nick to the second strand. When there are two PAMs on the inside and two nicks on the outside, this outward nick orientation also can be referred to as “PAM-in”. In some embodiments, in the outward nick orientation, the polypeptide (e.g., the gene modifying polypeptide) and the additional polypeptide bind to sites on the target DNA between the location of the nick to the first strand and the location of the nick to the second. In some embodiments, in the outward nick orientation, the location of the nick to the second strand is positioned on the opposite side of the binding sites of the polypeptide and additional polypeptide relative to the location of the nick to the first strand. In some embodiments, in the outward orientation, the PAM site and the binding site of the second polypeptide which is at a distance from the target site are positioned between the location of the nick to the first strand and the location of the nick to the second strand.


An example of a gene modifying system that provides an outward nick orientation comprises a gene modifying polypeptide comprising a CRISPR/Cas domain, a template RNA comprising a gRNA that directs nicking of the target site DNA on the first strand, and an additional nucleic acid comprising an additional gRNA that directs nicking at a site a distance from the location of the first nick, wherein the location of the first nick and the location of the second nick are outside of the PAM sites of the sites to which the two gRNAs direct the gene modifying polypeptide (i.e., the PAM sites are between the location of the first nick and the location of the second nick). As a further example, another gene modifying system that provides an outward nick orientation comprises a gene modifying polypeptide comprising a zinc finger molecule and a first nickase domain wherein the zinc finger molecule binds to the target DNA in a manner that directs the first nickase domain to nick the first strand of the target site; an additional polypeptide comprising a CRISPR/Cas domain, and an additional nucleic acid comprising a gRNA that directs the additional polypeptide to nick a site a distance from the target site DNA on the second strand, wherein the location of the first nick and the location of the second nick are outside the PAM site and the site to which the zinc finger molecule binds (i.e., the PAM site and the site to which the zinc finger molecule binds are between the location of the first nick and the location of the second nick). As a further example, another gene modifying system that provides an outward nick orientation comprises a gene modifying polypeptide comprising a zinc finger molecule and a first nickase domain wherein the zinc finger molecule binds to the target DNA in a manner that directs the first nickase domain to nick the first strand of the target site; an additional polypeptide comprising a TAL effector molecule and a second nickase domain wherein the TAL effector molecule binds to a site a distance from the target site in a manner that directs the additional polypeptide to nick the second strand, wherein the location of the first nick and the location of the second nick are outside the site to which the TAL effector molecule binds and the site to which the zinc finger molecule binds (i.e., the site to which the TAL effector molecule binds and the site to which the zinc finger molecule binds are between the location of the first nick and the location of the second nick).


Without wishing to be bound by theory, it is thought that, for gene modifying systems where a second strand nick is provided, an outward nick orientation is preferred in some embodiments. As is described herein, an inward nick may produce a higher number of double-strand breaks (DSBs) than an outward nick orientation. DSBs may be recognized by the DSB repair pathways in the nucleus of a cell, which can result in undesired insertions and deletions. An outward nick orientation may provide a decreased risk of DSB formation, and a corresponding lower amount of undesired insertions and deletions. In some embodiments, undesired insertions and deletions are insertions and deletions not encoded by the heterologous object sequence, e.g., an insertion or deletion produced by the double-strand break repair pathway unrelated to the modification encoded by the heterologous object sequence. In some embodiments, a desired gene modification comprises a change to the target DNA (e.g., a substitution, insertion, or deletion) encoded by the heterologous object sequence (e.g., and achieved by the gene modifying writing the heterologous object sequence into the target site). In some embodiments, the first strand nick and the second strand nick are in an outward orientation.


In addition, the distance between the first strand nick and second strand nick may influence the extent to which one or more of: desired gene modifying system DNA modifications are obtained, undesired double-strand breaks (DSBs) occur, undesired insertions occur, or undesired deletions occur. Without wishing to be bound by theory, it is thought the second strand nick benefit, the biasing of DNA repair toward incorporation of the heterologous object sequence into the target DNA, increases as the distance between the first strand nick and second strand nick decreases. However, it is thought that the risk of DSB formation also increases as the distance between the first strand nick and second strand nick decreases. Correspondingly, it is thought that the number of undesired insertions and/or deletions may increase as the distance between the first strand nick and second strand nick decreases. In some embodiments, the distance between the first strand nick and second strand nick is chosen to balance the benefit of biasing DNA repair toward incorporation of the heterologous object sequence into the target DNA and the risk of DSB formation and of undesired deletions and/or insertions. In some embodiments, a system where the first strand nick and the second strand nick are at least a threshold distance apart has an increased level of desired gene modifying system modification outcomes, a decreased level of undesired deletions, and/or a decreased level of undesired insertions relative to an otherwise similar inward nick orientation system where the first nick and the second nick are less than the a threshold distance apart. In some embodiments the threshold distance(s) is given below.


In some embodiments, the first nick and the second nick are at least 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200 nucleotides apart. In some embodiments, the first nick and the second nick are no more than 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, or 250 nucleotides apart. In some embodiments, the first nick and the second nick are 20-200, 30-200, 40-200, 50-200, 60-200, 70-200, 80-200, 90-200, 100-200, 110-200, 120-200, 130-200, 140-200, 150-200, 160-200, 170-200, 180-200, 190-200, 20-190, 30-190, 40-190, 50-190, 60-190, 70-190, 80-190, 90-190, 100-190, 110-190, 120-190, 130-190, 140-190, 150-190, 160-190, 170-190, 180-190, 20-180, 30-180, 40-180, 50-180, 60-180, 70-180, 80-180, 90-180, 100-180, 110-180, 120-180, 130-180, 140-180, 150-180, 160-180, 170-180, 20-170, 30-170, 40-170, 50-170, 60-170, 70-170, 80-170, 90-170, 100-170, 110-170, 120-170, 130-170, 140-170, 150-170, 160-170, 20-160, 30-160, 40-160, 50-160, 60-160, 70-160, 80-160, 90-160, 100-160, 110-160, 120-160, 130-160, 140-160, 150-160, 20-150, 30-150, 40-150, 50-150, 60-150, 70-150, 80-150, 90-150, 100-150, 110-150, 120-150, 130-150, 140-150, 20-140, 30-140, 40-140, 50-140, 60-140, 70-140, 80-140, 90-140, 100-140, 110-140, 120-140, 130-140, 20-130, 30-130, 40-130, 50-130, 60-130, 70-130, 80-130, 90-130, 100-130, 110-130, 120-130, 20-120, 30-120, 40-120, 50-120, 60-120, 70-120, 80-120, 90-120, 100-120, 110-120, 20-110, 30-110, 40-110, 50-110, 60-110, 70-110, 80-110, 90-110, 100-110, 20-100, 30-100, 40-100, 50-100, 60-100, 70-100, 80-100, 90-100, 20-90, 30-90, 40-90, 50-90, 60-90, 70-90, 80-90, 20-80, 30-80, 40-80, 50-80, 60-80, 70-80, 20-70, 30-70, 40-70, 50-70, 60-70, 20-60, 30-60, 40-60, 50-60, 20-50, 30-50, 40-50, 20-40, 30-40, or 20-30 nucleotides apart. In some embodiments, the first nick and the second nick are 40-100 nucleotides apart.


Without wishing to be bound by theory, it is thought that, for gene modifying systems where a second strand nick is provided and an inward nick orientation is selected, increasing the distance between the first strand nick and second strand nick may be preferred. As is described herein, an inward nick orientation may produce a higher number of DSBs than an outward nick orientation, and may result in a higher amount of undesired insertions and deletions than an outward nick orientation, but increasing the distance between the nicks may mitigate that increase in DSBs, undesired deletions, and/or undesired insertions. In some embodiments, an inward nick orientation wherein the first nick and the second nick are at least a threshold distance apart has an increased level of desired gene modifying system modification outcomes, a decreased level of undesired deletions, and/or a decreased level of undesired insertions relative to an otherwise similar inward nick orientation system where the first nick and the second nick are less than the a threshold distance apart. In some embodiments the threshold distance is given below.


In some embodiments, the first strand nick and the second strand nick are in an inward orientation. In some embodiments, the first strand nick and the second strand nick are in an inward orientation and the first strand nick and second strand nick are at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 220, 240, 260, 280, 300, 350, 400, 450, or 500 nucleotides apart, e.g., at least 100 nucleotides apart, (and optionally no more than 500, 400, 300, 200, 190, 180, 170, 160, 150, 140, 130, or 120 nucleotides apart). In some embodiments, the first strand nick and the second strand nick are in an inward orientation and the first strand nick and second strand nick are 100-200, 110-200, 120-200, 130-200, 140-200, 150-200, 160-200, 170-200, 180-200, 190-200, 100-190, 110-190, 120-190, 130-190, 140-190, 150-190, 160-190, 170-190, 180-190, 100-180, 110-180, 120-180, 130-180, 140-180, 150-180, 160-180, 170-180, 100-170, 110-170, 120-170, 130-170, 140-170, 150-170, 160-170, 100-160, 110-160, 120-160, 130-160, 140-160, 150-160, 100-150, 110-150, 120-150, 130-150, 140-150, 100-140, 110-140, 120-140, 130-140, 100-130, 110-130, 120-130, 100-120, 110-120, or 100-110 nucleotides apart.


Chemically Modified Nucleic Acids and Nucleic Acid End Features

A nucleic acid described herein (e.g., a template nucleic acid, e.g., a template RNA; or a nucleic acid (e.g., mRNA) encoding a gene modifying polypeptide; or a gRNA) can comprise unmodified or modified nucleobases. Naturally occurring RNAs are synthesized from four basic ribonucleotides: ATP, CTP, UTP and GTP, but may contain post-transcriptionally modified nucleotides. Further, approximately one hundred different nucleoside modifications have been identified in RNA (Rozenski, J, Crain, P, and McCloskey, J. (1999). The RNA Modification Database: 1999 update. Nucl Acids Res 27: 196-197). An RNA can also comprise wholly synthetic nucleotides that do not occur in nature.


In some embodiments, the chemical modification is one provided in WO/2016/183482, US Pat. Pub. No. 20090286852, of International Application No. WO/2012/019168, WO/2012/045075, WO/2012/135805, WO/2012/158736, WO/2013/039857, WO/2013/039861, WO/2013/052523, WO/2013/090648, WO/2013/096709, WO/2013/101690, WO/2013/106496, WO/2013/130161, WO/2013/151669, WO/2013/151736, WO/2013/151672, WO/2013/151664, WO/2013/151665, WO/2013/151668, WO/2013/151671, WO/2013/151667, WO/2013/151670, WO/2013/151666, WO/2013/151663, WO/2014/028429, WO/2014/081507, WO/2014/093924, WO/2014/093574, WO/2014/113089, WO/2014/144711, WO/2014/144767, WO/2014/144039, WO/2014/152540, WO/2014/152030, WO/2014/152031, WO/2014/152027, WO/2014/152211, WO/2014/158795, WO/2014/159813, WO/2014/164253, WO/2015/006747, WO/2015/034928, WO/2015/034925, WO/2015/038892, WO/2015/048744, WO/2015/051214, WO/2015/051173, WO/2015/051169, WO/2015/058069, WO/2015/085318, WO/2015/089511, WO/2015/105926, WO/2015/164674, WO/2015/196130, WO/2015/196128, WO/2015/196118, WO/2016/011226, WO/2016/011222, WO/2016/011306, WO/2016/014846, WO/2016/022914, WO/2016/036902, WO/2016/077125, or WO/2016/077123, each of which is herein incorporated by reference in its entirety. It is understood that incorporation of a chemically modified nucleotide into a polynucleotide can result in the modification being incorporated into a nucleobase, the backbone, or both, depending on the location of the modification in the nucleotide. In some embodiments, the backbone modification is one provided in EP 2813570, which is herein incorporated by reference in its entirety. In some embodiments, the modified cap is one provided in US Pat. Pub. No. 20050287539, which is herein incorporated by reference in its entirety.


In some embodiments, the chemically modified nucleic acid (e.g., RNA, e.g., mRNA) comprises one or more of ARCA: anti-reverse cap analog (m27.3′-OGP3G), GP3G (Unmethylated Cap Analog), m7GP3G (Monomethylated Cap Analog), m32.2.7GP3G (Trimethylated Cap Analog), m5CTP (5′-methyl-cytidine triphosphate), m6ATP (N6-methyl-adenosine-5″-triphosphate), s2UTP (2-thio-uridine triphosphate), and Ψ (pseudouridine triphosphate).


In some embodiments, the chemically modified nucleic acid comprises a 5′ cap, e.g.: a 7-methylguanosine cap (e.g., a O-Me-m7G cap); a hypermethylated cap analog; an NAD+-derived cap analog (e.g., as described in Kiledjian, Trends in Cell Biology 28, 454-464 (2018)); or a modified, e.g., biotinylated, cap analog (e.g., as described in Bednarek et al., Phil Trans R Soc B 373, 20180167 (2018)).


In some embodiments, the chemically modified nucleic acid comprises a 3′ feature selected from one or more of: a polyA tail; a 16-nucleotide long stem-loop structure flanked by unpaired 5 nucleotides (e.g., as described by Mannironi et al., Nucleic Acid Research 17, 9113-9126 (1989)); a triple-helical structure (e.g., as described by Brown et al., PNAS 109, 19202-19207 (2012)); a tRNA, Y RNA, or vault RNA structure (e.g., as described by Labno et al., Biochemica et Biophysica Acta 1863, 3125-3147 (2016)); incorporation of one or more deoxyribonucleotide triphosphates (dNTPs), 2′O-Methylated NTPs, or phosphorothioate-NTPs; a single nucleotide chemical modification (e.g., oxidation of the 3′ terminal ribose to a reactive aldehyde followed by conjugation of the aldehyde-reactive modified nucleotide); or chemical ligation to another nucleic acid molecule.


In some embodiments, the nucleic acid (e.g., template nucleic acid) comprises one or more modified nucleotides, e.g., selected from dihydrouridine, inosine, 7-methylguanosine, 5-methylcytidine (5mC), 5′ Phosphate ribothymidine, 2′-O-methyl ribothymidine, 2′-O-ethyl ribothymidine, 2′-fluoro ribothymidine, C-5 propynyl-deoxycytidine (pdC), C-5 propynyl-deoxyuridine (pdU), C-5 propynyl-cytidine (pC), C-5 propynyl-uridine (pU), 5-methyl cytidine, 5-methyl uridine, 5-methyl deoxycytidine, 5-methyl deoxyuridine methoxy, 2,6-diaminopurine, 5′-Dimethoxytrityl-N4-ethyl-2′-deoxycytidine, C-5 propynyl-f-cytidine (pfC), C-5 propynyl-f-uridine (pfU), 5-methyl f-cytidine, 5-methyl f-uridine, C-5 propynyl-m-cytidine (pmC), C-5 propynyl-f-uridine (pmU), 5-methyl m-cytidine, 5-methyl m-uridine, LNA (locked nucleic acid), MGB (minor groove binder) pseudouridine (T), 1-N-methylpseudouridine (1-Me-′P), or 5-methoxyuridine (S-MO-U).


In some embodiments, the nucleic acid comprises a backbone modification, e.g., a modification to a sugar or phosphate group in the backbone. In some embodiments, the nucleic acid comprises a nucleobase modification.


In some embodiments, the nucleic acid comprises one or more chemically modified nucleotides of Table 13, one or more chemical backbone modifications of Table 14, one or more chemically modified caps of Table 15. For instance, in some embodiments, the nucleic acid comprises two or more (e.g., 3, 4, 5, 6, 7, 8, 9, or 10 or more) different types of chemical modifications. As an example, the nucleic acid may comprise two or more (e.g., 3, 4, 5, 6, 7, 8, 9, or 10 or more) different types of modified nucleobases, e.g., as described herein, e.g., in Table 13. Alternatively or in combination, the nucleic acid may comprise two or more (e.g., 3, 4, 5, 6, 7, 8, 9, or 10 or more) different types of backbone modifications, e.g., as described herein, e.g., in Table 14. Alternatively or in combination, the nucleic acid may comprise one or more modified cap, e.g., as described herein, e.g., in Table 15. For instance, in some embodiments, the nucleic acid comprises one or more type of modified nucleobase and one or more type of backbone modification; one or more type of modified nucleobase and one or more modified cap; one or more type of modified cap and one or more type of backbone modification; or one or more type of modified nucleobase, one or more type of backbone modification, and one or more type of modified cap.


In some embodiments, the nucleic acid comprises one or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1000, or more) modified nucleobases. In some embodiments, all nucleobases of the nucleic acid are modified. In some embodiments, the nucleic acid is modified at one or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1000, or more) positions in the backbone. In some embodiments, all backbone positions of the nucleic acid are modified.









TABLE 13





Modified nucleotides
















5-aza-uridine
N2-methyl-6-thio-guanosine


2-thio-5-aza-midine
N2,N2-dimethyl-6-thio-guanosine


2-thiouridine
pyridin-4-one ribonucleoside


4-thio-pseudouridine
2-thio-5-aza-uridine


2-thio-pseudouridine
2-thiomidine


5-hydroxyuridine
4-thio-pseudomidine


3-methyluridine
2-thio-pseudowidine


5-carboxymethyl-uridine
3-methylmidine


1-carboxymethyl-pseudouridine
1-propynyl-pseudomidine


5-propynyl-uridine
1-methyl-1-deaza-pseudomidine


1-propynyl-pseudouridine
2-thio-1-methyl-1-deaza-pseudouridine


5-taurinomethyluridine
4-methoxy-pseudomidine


1-taurinomethyl-pseudouridine
5′-O-(1-Thiophosphate)-Adenosine


5-taurinomethyl-2-thio-uridine
5′-O-(1-Thiophosphate)-Cytidine


1-taurinomethyl-4-thio-uridine
5′-O-(1-thiophosphate)-Guanosine


5-methyl-uridine
5′-O-(1-Thiophophate)-Uridine


1-methyl-pseudouridine
5′-O-(1-Thiophosphate)-Pseudouridine


4-thio-1-methyl-pseudouridine
2′-O-methyl-Adenosine


2-thio-1-methyl-pseudouridine
2′-O-methyl-Cytidine


1-methyl-1-deaza-pseudouridine
2′-O-methyl-Guanosine


2-thio-1-methyl-1-deaza-pseudomidine
2′-O-methyl-Uridine


dihydrouridine
2′-O-methyl-Pseudouridine


dihydropseudouridine
2′-O-methyl-Inosine


2-thio-dihydromidine
2-methyladenosine


2-thio-dihydropseudouridine
2-methylthio-N6-methyladenosine


2-methoxyuridine
2-methylthio-N6 isopentenyladenosine


2-methoxy-4-thio-uridine
2-methylthio-N6-(cis-


4-methoxy-pseudouridine
hydroxyisopentenyl)adenosine


4-methoxy-2-thio-pseudouridine
N6-methyl-N6-threonylcarbamoyladenosine


5-aza-cytidine
N6-hydroxynorvalylcarbamoyladenosine


pseudoisocytidine
2-methylthio-N6-hydroxynorvalyl


3-methyl-cytidine
carbamoyladenosine


N4-acetylcytidine
2′-O-ribosyladenosine (phosphate)


5-formylcytidine
1,2′-O-dimethylinosine


N4-methylcytidine
5,2′-O-dimethylcytidine


5-hydroxymethylcytidine
N4-acetyl-2′-O-methylcytidine


1-methyl-pseudoisocytidine
Lysidine


pyrrolo-cytidine
7-methylguanosine


pyrrolo-pseudoisocytidine
N2,2′-O-dimethylguanosine


2-thio-cytidine
N2,N2,2′-O-trimethylguanosine


2-thio-5-methyl-cytidine
2′-O-ribosylguanosine (phosphate)


4-thio-pseudoisocytidine
Wybutosine


4-thio-1-methyl-pseudoisocytidine
Peroxywybutosine


4-thio-1-methyl-1-deaza-pseudoisocytidine
Hydroxywybutosine


1-methyl-1-deaza-pseudoisocytidine
undermodified hydroxywybutosine


zebularine
methylwyosine


5-aza-zebularine
queuosine


5-methyl-zebularine
epoxyqueuosine


5-aza-2-thio-zebularine
galactosyl-queuosine


2-thio-zebularine
mannosyl-queuosine


2-methoxy-cytidine
7-cyano-7-deazaguanosine


2-methoxy-5-methyl-cytidine
7-aminomethyl-7-deazaguanosine


4-methoxy-pseudoisocytidine
archaeosine


4-methoxy-1-methyl-pseudoisocytidine
5,2′-O-dimethyluridine


2-aminopurine
4-thiouridine


2,6-diaminopurine
5-methyl-2-thiouridine


7-deaza-adenine
2-thio-2′-O-methyluridine


7-deaza-8-aza-adenine
3-(3-amino-3-carboxypropyl)uridine


7-deaza-2-aminopurine
5-methoxyuridine


7-deaza-8-aza-2-aminopurine
uridine 5-oxyacetic acid


7-deaza-2,6-diaminopurine
uridine 5-oxyacetic acid methyl ester


7-deaza-8-aza-2,6-diarninopurine
5-(carboxyhydroxymethyl)uridine)


1-methyladenosine
5-(carboxyhydroxymethyl)uridine methyl ester


N6-isopentenyladenosine
5-methoxycarbonylmethyluridine


N6-(cis-hydroxyisopentenyl)adenosine
5-methoxycarbonylmethyl-2′-O-methyluridine


2-methylthio-N6-(cis-hydroxyisopentenyl)
5-methoxycarbonylmethyl-2-thiouridine


adenosine
5-aminomethyl-2-thiouridine


N6-glycinylcarbamoyladenosine
5-methylaminomethyluridine


N6-threonylcarbamoyladenosine
5-methylaminomethyl-2-thiouridine


2-methylthio-N6-threonyl
5-methylaminomethyl-2-selenouridine


carbamoyladenosine
5-carbamoylmethyluridine


N6,N6-dimethyladenosine
5-carbamoylmethyl-2′-O-methyluridine


7-methyladenine
5-carboxymethylaminomethyluridine


2-methylthio-adenine
5-carboxymethylaminomethyl-2′-O-


2-methoxy-adenine
methyluridine


inosine
5-carboxymethylaminomethyl-2-thiouridine


1-methyl-inosine
N4,2′-O-dimethylcytidine


wyosine
5-carboxymethyluridine


wybutosine
N6,2′-O-dimethyladenosine


7-deaza-guanosine
N,N6,O-2′-trimethyladenosine


7-deaza-8-aza-guanosine
N2,7-dimethylguanosine


6-thio-guanosine
N2,N2,7-trimethylguanosine


6-thio-7-deaza-guanosine
3,2′-O-dimethyluridine


6-thio-7-deaza-8-aza-guanosine
5-methyldihydrouridine


7-methyl-guanosine
5-formy1-2′-O-methylcytidine


6-thio-7-methyl-guanosine
1,2′-O-dimethylguanosine


7-methylinosine
4-demethylwyosine


6-methoxy-guanosine
Isowyosine


1-methylguanosine
N6-acetyladenosine


N2-methylguanosine



N2,N2-dimethylguanosine



8-oxo-guanosine



7-methyl-8-oxo-guanosine



1-methyl-6-thio-guanosine
















TABLE 14





Backbone modifications







2′-O-Methyl backbone


Peptide Nucleic Acid (PNA) backbone


phosphorothioate backbone


morpholino backbone


carbamate backbone


siloxane backbone


sulfide backbone


sulfoxide backbone


sulfone backbone


formacetyl backbone


thioformacetyl backbone


methyleneformacetyl backbone


riboacetyl backbone


alkene containing backbone


sulfamate backbone


sulfonate backbone


sulfonamide backbone


methyleneimino backbone


methylenehydrazino backbone


amide backbone
















TABLE 15





Modified caps







m7GpppA


m7GpppC


m2,7GpppG


m2,2,7GpppG


m7Gpppm7G


m7,2′OmeGpppG


m72′dGpppG


m7,3′OmeGpppG


m7,3′dGpppG


GppppG


m7GppppG


m7GppppA


m7GppppC


m2,7GppppG


m2,2,7GppppG


m7Gppppm7G


m7,2′OmeGppppG


m72′dGppppG


m7,3′OmeGppppG


m7,3′dGppppG









The nucleotides comprising the template of the gene modifying system can be natural or modified bases, or a combination thereof. For example, the template may contain pseudouridine, dihydrouridine, inosine, 7-methylguanosine, or other modified bases. In some embodiments, the template may contain locked nucleic acid nucleotides. In some embodiments, the modified bases used in the template do not inhibit the reverse transcription of the template. In some embodiments, the modified bases used in the template may improve reverse transcription, e.g., specificity or fidelity.


In some embodiments, an RNA component of the system (e.g., a template RNA or a gRNA) comprises one or more nucleotide modifications. In some embodiments, the modification pattern of a gRNA can significantly affect in vivo activity compared to unmodified or end-modified guides (e.g., as shown in FIG. 1D from Finn et al. Cell Rep 22(9):2227-2235 (2018); incorporated herein by reference in its entirety). Without wishing to be bound by theory, this process may be due, at least in part, to a stabilization of the RNA conferred by the modifications. Non-limiting examples of such modifications may include 2′-O-methyl (2′-O-Me), 2′-O-(2-methoxyethyl) (2′-O-M0E), 2′-fluoro (2′-F), phosphorothioate (PS) bond between nucleotides, G-C substitutions, and inverted abasic linkages between nucleotides and equivalents thereof.


In some embodiments, the template RNA (e.g., at the portion thereof that binds a target site) or the guide RNA comprises a 5′ terminus region. In some embodiments, the template RNA or the guide RNA does not comprise a 5′ terminus region. In some embodiments, the 5′ terminus region comprises a gRNA spacer region, e.g., as described with respect to sgRNA in Briner AE et al, Molecular Cell 56: 333-339 (2014) (incorporated herein by reference in its entirety; applicable herein, e.g., to all guide RNAs). In some embodiments, the 5′ terminus region comprises a 5′ end modification. In some embodiments, a 5′ terminus region with or without a spacer region may be associated with a crRNA, trRNA, sgRNA and/or dgRNA. The gRNA spacer region can, in some instances, comprise a guide region, guide domain, or targeting domain.


In some embodiments, the template RNAs (e.g., at the portion thereof that binds a target site) or guide RNAs described herein comprises any of the sequences shown in Table 4 of WO2018107028A1, incorporated herein by reference in its entirety. In some embodiments, where a sequence shows a guide and/or spacer region, the composition may comprise this region or not. In some embodiments, a guide RNA comprises one or more of the modifications of any of the sequences shown in Table 4 of WO2018107028A1, e.g., as identified therein by a SEQ ID NO. In embodiments, the nucleotides may be the same or different, and/or the modification pattern shown may be the same or similar to a modification pattern of a guide sequence as shown in Table 4 of WO2018107028A1. In some embodiments, a modification pattern includes the relative position and identity of modifications of the gRNA or a region of the gRNA (e.g. 5′ terminus region, lower stem region, bulge region, upper stem region, nexus region, hairpin 1 region, hairpin 2 region, 3′ terminus region). In some embodiments, the modification pattern contains at least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% of the modifications of any one of the sequences shown in the sequence column of Table 4 of WO2018107028A1, and/or over one or more regions of the sequence. In some embodiments, the modification pattern is at least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the modification pattern of any one of the sequences shown in the sequence column of Table 4 of WO2018107028A1. In some embodiments, the modification pattern is at least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over one or more regions of the sequence shown in Table 4 of WO2018107028A1, e.g., in a 5′ terminus region, lower stem region, bulge region, upper stem region, nexus region, hairpin 1 region, hairpin 2 region, and/or 3′ terminus region. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the modification pattern of a sequence over the 5′ terminus region. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the lower stem. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the bulge. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the upper stem. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the nexus. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the hairpin 1. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the hairpin 2. In some embodiments, the modification pattern is least 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical over the 3′ terminus. In some embodiments, the modification pattern differs from the modification pattern of a sequence of Table 4 of WO2018107028A1, or a region (e.g. 5′ terminus, lower stem, bulge, upper stem, nexus, hairpin 1, hairpin 2, 3′ terminus) of such a sequence, e.g., at 0, 1, 2, 3, 4, 5, 6, or more nucleotides. In some embodiments, the gRNA comprises modifications that differ from the modifications of a sequence of Table 4 of WO2018107028A1, e.g., at 0, 1, 2, 3, 4, 5, 6, or more nucleotides. In some embodiments, the gRNA comprises modifications that differ from modifications of a region (e.g. 5′ terminus, lower stem, bulge, upper stem, nexus, hairpin 1, hairpin 2, 3′ terminus) of a sequence of Table 4 of WO2018107028A1, e.g., at 0, 1, 2, 3, 4, 5, 6, or more nucleotides.


In some embodiments, the template RNAs (e.g., at the portion thereof that binds a target site) or the gRNA comprises a 2′-O-methyl (2′-O-Me) modified nucleotide. In some embodiments, the gRNA comprises a 2′-O-(2-methoxy ethyl) (2′-O-moe) modified nucleotide. In some embodiments, the gRNA comprises a 2′-fluoro (2′-F) modified nucleotide. In some embodiments, the gRNA comprises a phosphorothioate (PS) bond between nucleotides. In some embodiments, the gRNA comprises a 5′ end modification, a 3′ end modification, or 5′ and 3′ end modifications. In some embodiments, the 5′ end modification comprises a phosphorothioate (PS) bond between nucleotides. In some embodiments, the 5′ end modification comprises a 2′-O-methyl (2′-O-Me), 2′-O-(2-methoxy ethyl) (2′-O-M0E), and/or 2′-fluoro (2′-F) modified nucleotide. In some embodiments, the 5′ end modification comprises at least one phosphorothioate (PS) bond and one or more of a 2′-O-methyl (2′-O-Me), 2′-O-(2-methoxyethyl) (2′-O-M0E), and/or 2′-fluoro (2′-F) modified nucleotide. The end modification may comprise a phosphorothioate (PS), 2′-O-methyl (2′-O-Me), 2′-O-(2-methoxyethyl) (2′-O-MOE), and/or 2′-fluoro (2′-F) modification. Equivalent end modifications are also encompassed by embodiments described herein. In some embodiments, the template RNA or gRNA comprises an end modification in combination with a modification of one or more regions of the template RNA or gRNA. Additional exemplary modifications and methods for protecting RNA, e.g., gRNA, and formulae thereof, are described in WO2018126176A1, which is incorporated herein by reference in its entirety.


In some embodiments, a template RNA described herein comprises three phosphorothioate linkages at the 5′ end and three phosphorothioate linkages at the 3′ end. In some embodiments, a template RNA described herein comprises three 2′-O-methyl ribonucleotides at the 5′ end and three 2′-O-methyl ribonucleotides at the 3′ end. In some embodiments, the 5′ most three nucleotides of the template RNA are 2′-O-methyl ribonucleotides, the 5′ most three internucleotide linkages of the template RNA are phosphorothioate linkages, the 3′ most three nucleotides of the template RNA are 2′-O-methyl ribonucleotides, and the 3′ most three internucleotide linkages of the template RNA are phosphorothioate linkages. In some embodiments, the template RNA comprises alternating blocks of ribonucleotides and 2′-O-methyl ribonucleotides, for instance, blocks of between 12 and 28 nucleotides in length. In some embodiments, the central portion of the template RNA comprises the alternating blocks and the 5′ and 3′ ends each comprise three 2′-O-methyl ribonucleotides and three phosphorothioate linkages.


In some embodiments, structure-guided and systematic approaches are used to introduce modifications (e.g., 2′-OMe-RNA, 2′-F-RNA, and PS modifications) to a template RNA or guide RNA, for example, as described in Mir et al. Nat Commun 9:2641 (2018) (incorporated by reference herein in its entirety). In some embodiments, the incorporation of 2′-F-RNAs increases thermal and nuclease stability of RNA:RNA or RNA:DNA duplexes, e.g., while minimally interfering with C3′-endo sugar puckering. In some embodiments, 2′-F may be better tolerated than 2′-OMe at positions where the 2′-OH is important for RNA:DNA duplex stability. In some embodiments, a crRNA comprises one or more modifications that do not reduce Cas9 activity, e.g., C10, C20, or C21 (fully modified), e.g., as described in Supplementary Table 1 of Mir et al. Nat Commun 9:2641 (2018), incorporated herein by reference in its entirety. In some embodiments, a tracrRNA comprises one or more modifications that do not reduce Cas9 activity, e.g., T2, T6, T7, or T8 (fully modified) of Supplementary Table 1 of Mir et al. Nat Commun 9:2641 (2018). In some embodiments, a crRNA comprises one or more modifications (e.g., as described herein) may be paired with a tracrRNA comprising one or more modifications, e.g., C20 and T2. In some embodiments, a gRNA comprises a chimera, e.g., of a crRNA and a tracrRNA (e.g., Jinek et al. Science 337(6096):816-821 (2012)). In embodiments, modifications from the crRNA and tracrRNA are mapped onto the single-guide chimera, e.g., to produce a modified gRNA with enhanced stability.


In some embodiments, gRNA molecules may be modified by the addition or subtraction of the naturally occurring structural components, e.g., hairpins. In some embodiments, a gRNA may comprise a gRNA with one or more 3′ hairpin elements deleted, e.g., as described in WO2018106727, incorporated herein by reference in its entirety. In some embodiments, a gRNA may contain an added hairpin structure, e.g., an added hairpin structure in the spacer region, which was shown to increase specificity of a CRISPR-Cas system in the teachings of Kocak et al. Nat Biotechnol 37(6):657-666 (2019). Additional modifications, including examples of shortened gRNA and specific modifications improving in vivo activity, can be found in US20190316121, incorporated herein by reference in its entirety.


In some embodiments, structure-guided and systematic approaches (e.g., as described in Mir et al. Nat Commun 9:2641 (2018); incorporated herein by reference in its entirety) are employed to find modifications for the template RNA. In embodiments, the modifications are identified with the inclusion or exclusion of a guide region of the template RNA. In some embodiments, a structure of polypeptide bound to template RNA is used to determine non-protein-contacted nucleotides of the RNA that may then be selected for modifications, e.g., with lower risk of disrupting the association of the RNA with the polypeptide. Secondary structures in a template RNA can also be predicted in silico by software tools, e.g., the RNAstructure tool available at rna.urmc.rochester.edu/RNAstructureWeb (Bellaousov et al. Nucleic Acids Res 41:W471-W474 (2013); incorporated by reference herein in its entirety), e.g., to determine secondary structures for selecting modifications, e.g., hairpins, stems, and/or bulges.


Production of Compositions and Systems

As will be appreciated by one of skill, methods of designing and constructing nucleic acid constructs and proteins or polypeptides (such as the systems, constructs and polypeptides described herein) are routine in the art. Generally, recombinant methods may be used. See, in general, Smales & James (Eds.), Therapeutic Proteins: Methods and Protocols (Methods in Molecular Biology), Humana Press (2005); and Crommelin, Sindelar & Meibohm (Eds.), Pharmaceutical Biotechnology: Fundamentals and Applications, Springer (2013). Methods of designing, preparing, evaluating, purifying and manipulating nucleic acid compositions are described in Green and Sambrook (Eds.), Molecular Cloning: A Laboratory Manual (Fourth Edition), Cold Spring Harbor Laboratory Press (2012).


The disclosure provides, in part, a nucleic acid, e.g., vector, encoding a gene modifying polypeptide described herein, a template nucleic acid described herein, or both. In some embodiments, a vector comprises a selective marker, e.g., an antibiotic resistance marker. In some embodiments, the antibiotic resistance marker is a kanamycin resistance marker. In some embodiments, the antibiotic resistance marker does not confer resistance to beta-lactam antibiotics. In some embodiments, the vector does not comprise an ampicillin resistance marker. In some embodiments, the vector comprises a kanamycin resistance marker and does not comprise an ampicillin resistance marker. In some embodiments, a vector encoding a gene modifying polypeptide is integrated into a target cell genome (e.g., upon administration to a target cell, tissue, organ, or subject). In some embodiments, a vector encoding a gene modifying polypeptide is not integrated into a target cell genome (e.g., upon administration to a target cell, tissue, organ, or subject). In some embodiments, a vector encoding a template nucleic acid (e.g., template RNA) is not integrated into a target cell genome (e.g., upon administration to a target cell, tissue, organ, or subject). In some embodiments, if a vector is integrated into a target site in a target cell genome, the selective marker is not integrated into the genome. In some embodiments, if a vector is integrated into a target site in a target cell genome, genes or sequences involved in vector maintenance (e.g., plasmid maintenance genes) are not integrated into the genome. In some embodiments, if a vector is integrated into a target site in a target cell genome, transfer regulating sequences (e.g., inverted terminal repeats, e.g., from an AAV) are not integrated into the genome. In some embodiments, administration of a vector (e.g., encoding a gene modifying polypeptide described herein, a template nucleic acid described herein, or both) to a target cell, tissue, organ, or subject results in integration of a portion of the vector into one or more target sites in the genome(s) of said target cell, tissue, organ, or subject. In some embodiments, less than 99, 95, 90, 80, 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2, or 1% of target sites (e.g., no target sites) comprising integrated material comprise a selective marker (e.g., an antibiotic resistance gene), a transfer regulating sequence (e.g., an inverted terminal repeat, e.g., from an AAV), or both from the vector.


Exemplary methods for producing a therapeutic pharmaceutical protein or polypeptide described herein involve expression in mammalian cells, although recombinant proteins can also be produced using insect cells, yeast, bacteria, or other cells under control of appropriate promoters. Mammalian expression vectors may comprise non-transcribed elements such as an origin of replication, a suitable promoter, and other 5′ or 3′ flanking non-transcribed sequences, and 5′ or 3′ non-translated sequences such as necessary ribosome binding sites, a polyadenylation site, splice donor and acceptor sites, and termination sequences. DNA sequences derived from the SV40 viral genome, for example, SV40 origin, early promoter, splice, and polyadenylation sites may be used to provide other genetic elements required for expression of a heterologous DNA sequence. Appropriate cloning and expression vectors for use with bacterial, fungal, yeast, and mammalian cellular hosts are described in Green & Sambrook, Molecular Cloning: A Laboratory Manual (Fourth Edition), Cold Spring Harbor Laboratory Press (2012).


Various mammalian cell culture systems can be employed to express and manufacture recombinant protein. Examples of mammalian expression systems include CHO, COS, HEK293, HeLA, and BHK cell lines. Processes of host cell culture for production of protein therapeutics are described in Zhou and Kantardjieff (Eds.), Mammalian Cell Cultures for Biologics Manufacturing (Advances in Biochemical Engineering/Biotechnology), Springer (2014). Compositions described herein may include a vector, such as a viral vector, e.g., a lentiviral vector, encoding a recombinant protein. In some embodiments, a vector, e.g., a viral vector, may comprise a nucleic acid encoding a recombinant protein.


Purification of protein therapeutics is described in Franks, Protein Biotechnology: Isolation, Characterization, and Stabilization, Humana Press (2013); and in Cutler, Protein Purification Protocols (Methods in Molecular Biology), Humana Press (2010).


The disclosure also provides compositions and methods for the production of template nucleic acid molecules (e.g., template RNAs) with specificity for a gene modifying polypeptide and/or a genomic target site. In an aspect, the method comprises production of RNA segments including an upstream homology segment, a heterologous object sequence segment, a gene modifying polypeptide binding motif, and a gRNA segment.


Therapeutic Applications

In some embodiments, a gene modifying system as described herein can be used to modify a cell (e.g., an animal cell, plant cell, or fungal cell). In some embodiments, a gene modifying system as described herein can be used to modify a mammalian cell (e.g., a human cell). In some embodiments, a gene modifying system as described herein can be used to modify a cell from a livestock animal (e.g., a cow, horse, sheep, goat, pig, llama, alpaca, camel, yak, chicken, duck, goose, or ostrich). In some embodiments, a gene modifying system as described herein can be used as a laboratory tool or a research tool, or used in a laboratory method or research method, e.g., to modify an animal cell, e.g., a mammalian cell (e.g., a human cell), a plant cell, or a fungal cell.


By integrating coding genes into a RNA sequence template, the gene modifying system can address therapeutic needs, for example, by providing expression of a therapeutic transgene in individuals with loss-of-function mutations, by replacing gain-of-function mutations with normal transgenes, by providing regulatory sequences to eliminate gain-of-function mutation expression, and/or by controlling the expression of operably linked genes, transgenes and systems thereof. In certain embodiments, the RNA sequence template encodes a promotor region specific to the therapeutic needs of the host cell, for example a tissue specific promotor or enhancer. In still other embodiments, a promotor can be operably linked to a coding sequence.


Accordingly, provided herein are methods for treating alpha-1 antitrypsin deficiency (AATD) in a subject in need thereof. In some embodiments, treatment results in amelioration of one or more symptoms associated with AATD.


In some embodiments, a system herein is used to treat a subject having a mutation in E342 (e.g., E342K).


In some embodiments, treatment with a system disclosed herein results in correction of the E342K mutation in between about 30-100% (e.g., about 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, or about 50%) of cells. In some embodiments, treatment with a system disclosed herein results in correction of the E342K mutation in between about 30-60% (e.g., about 30-40%, 40-50%, 50-60%, or about 50%) of DNA isolated from the treated cells.


In some embodiments, treatment with a gene modifying system described herein results in one or more of:

    • (a) an increase in alpha-1 antitrypsin (AAT) activity and/or function;
    • (b) an increase in levels of circulating AAT;
    • (c) a reduction in protease-induced lung damage and/or inflammation (e.g., a reduction in protease digestion of connective tissue in the lower airway, e.g., alveoli linings));
    • (d) a reduction in accumulated, polymerized Z-AAT protein within hepatocytes;
    • (e) a reduction in AAT-induced hepatocyte toxicity;
    • (f) a reduction of cellular stress, inflammation, fibrosis, cirrhosis, hepatocellular carcinoma (HCC), and/or neonatal liver disease;
    • (g) an increase in pulmonary function (e.g., an increase in lung elasticity); and/or
    • (h) a reduction of symptoms associated with emphysema, as compared to a subject having AATD that has not been treated with a gene modifying system described herein.


Administration and Delivery

The compositions and systems described herein may be used in vitro or in vivo. In some embodiments the system or components of the system are delivered to cells (e.g., mammalian cells, e.g., human cells), e.g., in vitro or in vivo. In some embodiments, the cells are eukaryotic cells, e.g., cells of a multicellular organism, e.g., an animal, e.g., a mammal (e.g., human, swine, bovine), a bird (e.g., poultry, such as chicken, turkey, or duck), or a fish. In some embodiments, the cells are non-human animal cells (e.g., a laboratory animal, a livestock animal, or a companion animal). In some embodiments, the cell is a stem cell (e.g., a hematopoietic stem cell), a fibroblast, or a T cell. In some embodiments, the cell is an immune cell, e.g., a T cell (e.g., a Treg, CD4, CD8, γδ, or memory T cell), B cell (e.g., memory B cell or plasma cell), or NK cell. In some embodiments, the cell is a non-dividing cell, e.g., a non-dividing fibroblast or non-dividing T cell. In some embodiments, the cell is an HSC and p53 is not upregulated or is upregulated by less than 10%, 5%, 2%, or 1%, e.g., as determined according to the method described in Example 30 of PCT/US2019/048607. The skilled artisan will understand that the components of the gene modifying system may be delivered in the form of polypeptide, nucleic acid (e.g., DNA, RNA), and combinations thereof.


In one embodiment the system and/or components of the system are delivered as nucleic acid. For example, the gene modifying polypeptide may be delivered in the form of a DNA or RNA encoding the polypeptide, and the template RNA may be delivered in the form of RNA or its complementary DNA to be transcribed into RNA. In some embodiments the system or components of the system are delivered on 1, 2, 3, 4, or more distinct nucleic acid molecules. In some embodiments the system or components of the system are delivered as a combination of DNA and RNA. In some embodiments the system or components of the system are delivered as a combination of DNA and protein. In some embodiments the system or components of the system are delivered as a combination of RNA and protein. In some embodiments the gene modifying polypeptide is delivered as a protein.


In some embodiments the system or components of the system are delivered to cells, e.g. mammalian cells or human cells, using a vector. The vector may be, e.g., a plasmid or a virus. In some embodiments, delivery is in vivo, in vitro, ex vivo, or in situ. In some embodiments the virus is an adeno associated virus (AAV), a lentivirus, or an adenovirus. In some embodiments the system or components of the system are delivered to cells with a viral-like particle or a virosome. In some embodiments the delivery uses more than one virus, viral-like particle or virosome.


In one embodiment, the compositions and systems described herein can be formulated in liposomes or other similar vesicles. Liposomes are spherical vesicle structures composed of a uni- or multilamellar lipid bilayer surrounding internal aqueous compartments and a relatively impermeable outer lipophilic phospholipid bilayer. Liposomes may be anionic, neutral or cationic. Liposomes are biocompatible, nontoxic, can deliver both hydrophilic and lipophilic drug molecules, protect their cargo from degradation by plasma enzymes, and transport their load across biological membranes and the blood brain barrier (BBB) (see, e.g., Spuch and Navarro, Journal of Drug Delivery, vol. 2011, Article ID 469679, 12 pages, 2011. doi:10.1155/2011/469679 for review).


Vesicles can be made from several different types of lipids; however, phospholipids are most commonly used to generate liposomes as drug carriers. Methods for preparation of multilamellar vesicle lipids are known in the art (see for example U.S. Pat. No. 6,693,086, the teachings of which relating to multilamellar vesicle lipid preparation are incorporated herein by reference). Although vesicle formation can be spontaneous when a lipid film is mixed with an aqueous solution, it can also be expedited by applying force in the form of shaking by using a homogenizer, sonicator, or an extrusion apparatus (see, e.g., Spuch and Navarro, Journal of Drug Delivery, vol. 2011, Article ID 469679, 12 pages, 2011. doi:10.1155/2011/469679 for review). Extruded lipids can be prepared by extruding through filters of decreasing size, as described in Templeton et al., Nature Biotech, 15:647-652, 1997, the teachings of which relating to extruded lipid preparation are incorporated herein by reference.


A variety of nanoparticles can be used for delivery, such as a liposome, a lipid nanoparticle, a cationic lipid nanoparticle, an ionizable lipid nanoparticle, a polymeric nanoparticle, a gold nanoparticle, a dendrimer, a cyclodextrin nanoparticle, a micelle, or a combination of the foregoing.


Lipid nanoparticles are an example of a carrier that provides a biocompatible and biodegradable delivery system for the pharmaceutical compositions described herein. Nanostructured lipid carriers (NLCs) are modified solid lipid nanoparticles (SLNs) that retain the characteristics of the SLN, improve drug stability and loading capacity, and prevent drug leakage. Polymer nanoparticles (PNPs) are an important component of drug delivery. These nanoparticles can effectively direct drug delivery to specific targets and improve drug stability and controlled drug release. Lipid—polymer nanoparticles (PLNs), a type of carrier that combines liposomes and polymers, may also be employed. These nanoparticles possess the complementary advantages of PNPs and liposomes. A PLN is composed of a core—shell structure; the polymer core provides a stable structure, and the phospholipid shell offers good biocompatibility. As such, the two components increase the drug encapsulation efficiency rate, facilitate surface modification, and prevent leakage of water-soluble drugs. For a review, see, e.g., Li et al. 2017, Nanomaterials 7, 122; doi:10.3390/nano7060122.


Exosomes can also be used as drug delivery vehicles for the compositions and systems described herein. For a review, see Ha et al. July 2016. Acta Pharmaceutica Sinica B. Volume 6, Issue 4, Pages 287-296; doi.org/10.1016/j.apsb.2016.02.001.


Fusosomes interact and fuse with target cells, and thus can be used as delivery vehicles for a variety of molecules. They generally consist of a bilayer of amphipathic lipids enclosing a lumen or cavity and a fusogen that interacts with the amphipathic lipid bilayer. The fusogen component has been shown to be engineerable in order to confer target cell specificity for the fusion and payload delivery, allowing the creation of delivery vehicles with programmable cell specificity (see for example Patent Application WO2020014209, the teachings of which relating to fusosome design, preparation, and usage are incorporated herein by reference).


In some embodiments, the protein component(s) of the gene modifying system may be pre-associated with the template nucleic acid (e.g., template RNA). For example, in some embodiments, the gene modifying polypeptide may be first combined with the template nucleic acid (e.g., template RNA) to form a ribonucleoprotein (RNP) complex. In some embodiments, the RNP may be delivered to cells via, e.g., transfection, nucleofection, virus, vesicle, LNP, exosome, fusosome.


A gene modifying system can be introduced into cells, tissues and multicellular organisms. In some embodiments the system or components of the system are delivered to the cells via mechanical means or physical means.


Formulation of protein therapeutics is described in Meyer (Ed.), Therapeutic Protein Drug Products: Practical Approaches to formulation in the Laboratory, Manufacturing, and the Clinic, Woodhead Publishing Series (2012).


Tissue Specific Activity/Administration

In some embodiments, a system described herein can make use of one or more feature (e.g., a promoter or microRNA binding site) to limit activity in off-target cells or tissues.


In some embodiments, a nucleic acid described herein (e.g., a template RNA or a DNA encoding a template RNA) comprises a promoter sequence, e.g., a tissue specific promoter sequence. In some embodiments, the tissue-specific promoter is used to increase the target-cell specificity of a gene modifying system. For instance, the promoter can be chosen on the basis that it is active in a target cell type but not active in (or active at a lower level in) a non-target cell type. Thus, even if the promoter integrated into the genome of a non-target cell, it would not drive expression (or only drive low level expression) of an integrated gene. A system having a tissue-specific promoter sequence in the template RNA may also be used in combination with a microRNA binding site, e.g., in the template RNA or a nucleic acid encoding a gene modifying protein, e.g., as described herein. A system having a tissue-specific promoter sequence in the template RNA may also be used in combination with a DNA encoding a gene modifying polypeptide, driven by a tissue-specific promoter, e.g., to achieve higher levels of gene modifying protein in target cells than in non-target cells. In some embodiments, e.g., for liver indications, a tissue-specific promoter is selected from Table 3 of WO2020014209, incorporated herein by reference.


In some embodiments, a nucleic acid described herein (e.g., a template RNA or a DNA encoding a template RNA) comprises a microRNA binding site. In some embodiments, the microRNA binding site is used to increase the target-cell specificity of a gene modifying system. For instance, the microRNA binding site can be chosen on the basis that is recognized by a miRNA that is present in a non-target cell type, but that is not present (or is present at a reduced level relative to the non-target cell) in a target cell type. Thus, when the template RNA is present in a non-target cell, it would be bound by the miRNA, and when the template RNA is present in a target cell, it would not be bound by the miRNA (or bound but at reduced levels relative to the non-target cell). While not wishing to be bound by theory, binding of the miRNA to the template RNA may interfere with its activity, e.g., may interfere with insertion of the heterologous object sequence into the genome. Accordingly, the system would edit the genome of target cells more efficiently than it edits the genome of non-target cells, e.g., the heterologous object sequence would be inserted into the genome of target cells more efficiently than into the genome of non-target cells, or an insertion or deletion is produced more efficiently in target cells than in non-target cells. A system having a microRNA binding site in the template RNA (or DNA encoding it) may also be used in combination with a nucleic acid encoding a gene modifying polypeptide, wherein expression of the gene modifying polypeptide is regulated by a second microRNA binding site, e.g., as described herein. In some embodiments, e.g., for liver indications, a miRNA is selected from Table 4 of WO2020014209, incorporated herein by reference.


In some embodiments, the template RNA comprises a microRNA sequence, an siRNA sequence, a guide RNA sequence, or a piwi RNA sequence.


Promoters


In some embodiments, one or more promoter or enhancer elements are operably linked to a nucleic acid encoding a gene modifying protein or a template nucleic acid, e.g., that controls expression of the heterologous object sequence. In certain embodiments, the one or more promoter or enhancer elements comprise cell-type or tissue specific elements. In some embodiments, the promoter or enhancer is the same or derived from the promoter or enhancer that naturally controls expression of the heterologous object sequence. For example, the ornithine transcarbomylase promoter and enhancer may be used to control expression of the ornithine transcarbomylase gene in a system or method provided by the invention for correcting ornithine transcarbomylase deficiencies. In some embodiments, the promoter is a promoter of Table 16 or 17 or a functional fragment or variant thereof.


Exemplary tissue specific promoters that are commercially available can be found, for example, at a uniform resource locator (e.g., invivogen.com/tissue-specific-promoters). In some embodiments, a promoter is a native promoter or a minimal promoter, e.g., which consists of a single fragment from the 5′ region of a given gene. In some embodiments, a native promoter comprises a core promoter and its natural 5′ UTR., in some embodiments, the 5′ UTR comprises an intron. in other embodiments, these include composite promoters, which combine promoter elements of different origins or were generated by assembling a distal enhancer with a minimal promoter of the same origin.


Exemplary cell or tissue specific promoters are provided in the tAles, below, and exemplary nucleic acid sequences encoding them are known in the art and can be readily accessed using a variety of resources, such as the INCM database, including RefSeq, as well as the Eukaryotic Promoter Database (//epd.epfl.ch//index.php).









TABLE 16







Exemplary cell or tissue-specific promoters










Promoter
Target cells







B29 Promoter
B cells



CD14 Promoter
Monocytic Cells



CD43 Promoter
Leukocytes and platelets



CD45 Promoter
Hematopoeitic cells



CD68 promoter
macrophages



Desmin promoter
muscle cells



Elastase-1
pancreatic acinar cells



promoter




Endoglin promoter
endothelial cells



fibronectin
differentiating cells, healing



promoter
tissue



Flt-1 promoter
endothelial cells



GFAP promoter
Astrocytes



GPIIB promoter
megakaryocytes



ICAM-2 Promoter
Endothelial cells



INF-Beta promoter
Hematopoeitic cells



Mb promoter
muscle cells



Nphs1 promoter
podocytes



OG-2 promoter
Osteoblasts, Odonblasts



SP-B promoter
Lung



Syn1 promoter
Neurons



WASP promoter
Hematopoeitic cells



SV40/bAlb
Liver



promoter




SV40/bAlb
Liver



promoter




SV40/Cd3
Leukocytes and platelets



promoter




SV40/CD45
hematopoeitic cells



promoter




NSE/RU5′
Mature Neurons



promoter

















TABLE 17







Additional exemplary cell or tissue-specific promoters









Promoter
Gene Description
Gene Specificity





APOA2
Apolipoprotein A-II
Hepatocytes (from hepatocyte




progenitors)


SERPINA1
Serpin peptidase inhibitor, clade A
Hepatocytes


(hAAT)
(alpha-1 antiproteinase,
(from definitive endoderm stage)



antitrypsin), member 1




(also named alpha 1 anti-tryps in)



CYP3A
Cytochrome P450, family 3,
Mature Hepatocytes



subfamily A, polypeptide



MIR122
MicroRNA 122
Hepatocytes




(from early stage embryonic




liver cells) and endoderm







Pancreatic specific promoters









INS
Insulin
Pancreatic beta cells




(from definitive endoderm stage)


IRS2
Insulin receptor substrate 2
Pancreatic beta cells


Pdx1
Pancreatic and duodenal
Pancreas



homeobox 1
(from definitive endoderm stage)


Alx3
Aristaless-like homeobox 3
Pancreatic beta cells




(from definitive endoderm stage)


Ppy
Pancreatic polypeptide
PP pancreatic cells




(gamma cells)







Cardiac specific promoters









Myh6
Myosin, heavy chain 6, cardiac
Late differentiation marker of cardiac


(aMHC)
muscle, alpha
muscle cells (atrial specificity)


MYL2
Myosin, light chain 2, regulatory,
Late differentiation marker of cardiac


(MLC-2v)
cardiac, slow
muscle cells (ventricular specificity)


ITNN13
Troponin I type 3 (cardiac)
Cardiomyocytes


(cTn1)

(from immature state)


ITNN13
Troponin I type 3 (cardiac)
Cardiomyocytes


(cTn1)

(from immature state)


NPPA
Natriuretic peptide precursor A (also
Atrial specificity in adult cells


(ANF)
named Atrial Natriuretic Factor)



Slc8a1
Solute carrier family 8
Cardiomyocytes from early


(Ncx1)
(sodium/calcium exchanger),
developmental stages



member 1








CNS specific promoters









SYN1
Synapsin I
Neurons


(hSyn)




GFAP
Glial fibrillary acidic protein
Astrocytes


INA
Internexin neuronal intermediate
Neuroprogenitors



filament protein, alpha (a-internexin)



NES
Nestin
Neuroprogenitors and ectoderm


MOBP
Myelin-associated oligodendrocyte
Oligodendrocytes



basic protein



MBP
Myelin basic protein
Oligodendrocytes


TH
Tyrosine hydroxylase
Dopaminergic neurons


FOXA2
Forkhead box A2
Dopaminergic neurons (also used as a


(HNF3

marker of endoderm)


beta)









Skin specific promoters









FLG
Filaggrin
Keratinocytes from granular layer


K14
Keratin 14
Keratinocytes from granular




and basal layers


TGM3
Transglutaminase 3
Keratinocytes from granular layer







Immune cell specific promoters









ITGAM
Integrin, alpha M (complement
Monocytes, macrophages, granulocytes,


(CD11B)
component 3 receptor 3 subunit)
natural killer cells







Urogential cell specific promoters









Pbsn
Probasin
Prostatic epithelium


Upk2
Uroplakin 2
Bladder


Sbp
Spermine binding protein
Prostate


Fer114
Fer-1-like 4
Bladder







Endothelial cell specific promoters









ENG
Endoglin
Endothelial cells







Pluripotent and embryonic cell specific promoters









Oct4
POU class 5 homeobox 1
Pluripotent cells


(POU5F1)

(germ cells, ES cells, iPS cells)


NANOG
Nanog homeobox
Pluripotent cells




(ES cells, iPS cells)


Synthetic
Synthetic promoter based on a Oct-4
Pluripotent cells (ES cells, iPS cells)


Oct4
core enhancer element



T
Brachyury
Mesoderm


brachyury




NES
Nestin
Neuroprogenitors and Ectoderm


SOX17
SRY (sex determining region Y)-box 17
Endoderm


FOXA2
Forkhead box A2
Endoderm (also used as a marker of


(HNFJ

dopaminergic neurons)


beta)




MIR122
MicroRNA 122
Endoderm and hepatocytes




(from early stage embryonic liver cells~









Depending on the host/vector system utilized, any of a number of suitable transcription and translation control elements, including constitutive and inducible promoters, transcription enhancer elements, transcription terminators, etc. may be used in the expression vector (see e.g., Bitter et al. (1987) Methods in Eta-yinology, 153:516-544; incorporated herein by reference in its entirety).


In some embodiments, a nucleic acid encoding a gene modifying protein or template nucleic acid is operably linked to a control element, e.g., a transcriptional control element, such as a promoter. The transcriptional control element may, in some embodiment, be functional in either a eukaryotic cell, e.g., a mammalian cell; or a prokaryotic cell (e.g., bacterial or archaeal cell). In some embodiments, a nucleotide sequence encoding a polypeptide is operably linked to multiple control elements, e.g., that allow expression of the nucleotide sequence encoding the polypeptide in both prokaryotic and eukaryotic cells.


For illustration purposes, examples of spatially restricted promoters include, but are not limited to, neuron-specific promoters, adipocyte-specific promoters, cardiomyocyte-specific promoters, smooth muscle-specific promoters, photoreceptor-specific promoters, etc. Neuron-specific spatially restricted promoters include, but are not limited to, a neuron-specific enolase (NSE) promoter (see, ENTBL HSENO2, X51956); an aromatic amino acid decarboxiase (AADC) promoter, a neurofilament promoter (see, e.g., GenBank HUMNFL, L04147); a syna.psin promoter (see, e.g., GenBank liUMSYNIB,1V155301); a thy-1 promoter (see, e.g., Chen et al. (1987) Cell 51:7-19; and Llewellyn, et al. (2010) Nat. Med. 16(10): 1161-1166); a serotonin receptor promoter (see, e.g., GenBank S62283); a tyrosine hydroxylase promoter (TH) (see, e.g., Oh et al. (2009) Gene Ther 16:437; Sasaoka et al. (1992) Mol. Brain Res. 16:274; Boundy et al. (1998) J. Neurosci. 18:9989; and Kaneda et al. (1991) Neuron 6:583-594), a GnRH promoter (see, e.g., Radovick et al. (1991) Proc. Natl. Aca.d. Sci. USA 88:3402-3406); an L7 promoter (see, e.g., Oherdick et al. (1990) Science 248:223-226); a DNMT promoter (see, e.g., Bartge et al. (1988) Proc. Natl. Acad Sci. USA 85:3648-3652); an enkephalin promoter (see, e.g., Comb et al. (1988) EMBO J. 17:3793-3805); a myelin basic protein (MBP) promoter; a Ca2+-calmodulin-dependent protein kinase (CainK110.) promoter (see, e.g., Mayford et al. (1996) Proc. Natl. Acad. Sci. USA 93:13250; and Casanova et al. (2001) Genesis 31:37); a CMV enhancer/platelet-derived growth factor-13 promoter (see, e.g., Liu et al. (2004) Gene Therapy 11:52-60); and the like.


Adipocyte-specific spatially restricted promoters include, but are not limited to, the aP2 gene promoter/enhancer, region from kb to +21 hp of a human aP2 gene (see, e.g., Tozzo et al. (1997) Endocrinol. 138:1604; Ross et al. (1990) Proc. Natl. Acad. Sci. USA 87:9590; and Payjani. et al. (2005) Nat. Med. 11:797); a glucose transporter-4 (GLI-l174) promoter (see, e.g., Knight et al. (2003) Proc. Natl. Acad. Sci. USA 100:14725); a fatty acid translocase (FAT/CD36) promoter (see, e.g., Kuriki et al. (2002) Biol. Pharm. Ball. 2511476, and Sato et al. (2002) J. Biol. Chem. 277:15703); a stearoyl-CoA desaturase-1 (SCD1) promoter (Tabor et al. (1999) J. Biol. Chem. 274:20603); a leptin promoter (see, e.g., Mason et al. (1998) Endocrinol. 139:1013; and (Then et al. (1999) Biochem. Biophys. Res. Comm. 262:187); an adiponectin promoter (see, e.g., Kita et al. (2005) Biochem. Biophys. Res. Comm. 331:484; and Chakraharti (2010) Endocrinol. 151:2408); an adipsin promoter (see, e.g., Platt et al. (1989) Proc. Natl. Acad. Sci. USA 86:7490); a resistin promoter (see, e.g., Seo et al. (2003) Molec. Endocrinol. 17:1522); and the like.


Cardiomyocyte-specific spatially restricted promoters include, but are not limited to; control sequences derived from the following genes: myosin light chain-2, α-myosin heavy chain, AE3, cardiac troponin C, cardiac actin, and the like. Franz et al (1997) Cardiova sc. Res. 35:560-566; Robbins et al. (1995) Ann. N.Y. Acad. Sci. 752:492-505; Linn et al. (1995) Circ. Res. 76:584-591; Parmacek et al. (1994) Mol. Cell. Biol. 14:1870-1885; Hunter et al. (1993) Hypertension 22:608-617; and Sartorelli et al. (1992) Proc. Natl. Acad. Sci. USA 89:4047-4051.


Smooth muscle-specific spatially restricted promoters include, but are not limited to an SM22u, promoter (see, e.g., Akvarek et al. (2000) Mol. Med. 6:983; and U.S. Pat. No. 7,169,874); a smoothelin promoter (see, WO 2001/018048); an α--smooth muscle actin promoter; and the like. For example, a 0.4 kb region of the SM22u promoter, within which lie two CArG elements, has been shown to mediate vascular smooth muscle cell-specific expression (see, e.g., Kim, et al. (1997) Mol. Cell. Biol. 17, 2266-2278; Li, et as, (1996) J. Cell Biol. 132, 849-859; and Moessier, et al. (1996) Development 122, 2415-2425).


Photoreceptor-specific spatially restricted promoters include, but are not limited to, a rhodopsin promoter; a rhodopsin kinase promoter (Young et al. (2003) Ophthalmol. Vis. Sci. 44:4076); a beta phosphodiesterase gene promoter (Nicoud et al. (2007) J. Gene Med. 9:1015); a retinitis pigmentosa gene promoter (Nicoud et al. (2007) supra); an interphotoreceptor retinoid-binding protein (IRBP) gene enhancer (Nicoud et al. (2007) supra); an IRBP gene promoter (Yokoyama et al. (1992) Exp Eye Res. 55:225); and the like.


In some embodiments, a gene modifying system, e.g., DNA encoding a gene modifying polypeptide, DNA encoding a template RNA, or DNA or RNA encoding a heterologous object sequence, is designed such that one or more elements is operably linked to a tissue-specific promoter, e.g., a promoter that is active in T-cells. In further embodiments, the T-cell active promoter is inactive in other cell types, e.g., B-cells, NK cells. In some embodiments, the T-cell active promoter is derived from a promoter for a gene encoding a component of the T-cell receptor, e.g., TRAC, TRBC, TRGC, TRDC. In some embodiments, the T-cell active promoter is derived from a promoter for a gene encoding a component of a T-cell-specific cluster of differentiation protein, e.g., CD3, e.g., CD3D, CD3E, CD3G, CD3Z. In some embodiments, T-cell-specific promoters in gene modifying systems are discovered by comparing publicly available gene expression data across cell types and selecting promoters from the genes with enhanced expression in T-cells. In some embodiments, promoters may be selecting depending on the desired expression breadth, e.g., promoters that are active in T-cells only, promoters that are active in NK cells only, promoters that are active in both T-cells and NK cells.


Cell-specific promoters known in the art may be used to direct expression of a gene modifying protein, e.g., as described herein. Nonlimiting exemplary mammalian cell-specific promoters have been characterized and used in mice expressing Cre recombinase in a cell-specific manner. Certain nonlimiting exemplary mammalian cell-specific promoters are listed in Table 1 of U.S. Pat. No. 9,845,481, incorporated herein by reference.


In some embodiments, a vector as described herein comprises an expression cassette. Typically, an expression cassette comprises the nucleic acid molecule of the instant invention operatively linked to a promoter sequence. For example, a promoter is operatively linked with a. coding sequence when it is capable of affecting the expression of that coding sequence (e.g., the coding sequence is under the transcriptional control of the promoter). Encoding sequences can be operatively linked to regulatory sequences in sense or antisense orientation. In certain embodiments, the promoter is a heterologous promoter. In certain embodiments, an expression cassette may comprise additional elements, for example, an intron, an enhancer, a polyadenylation site, a woodchuck response element (WRE), and/or other elements known to affect expression levels of the encoding sequence. A promoter typically controls the expression of a coding sequence or functional RNA. In certain embodiments, a promoter sequence comprises proximal and more distal upstream elements and can further comprise an enhancer element. An enhancer can typically stimulate promoter activity and may be an innate element of the promoter or a heterologous element inserted to enhance the level. or tissue-specificity of a promoter. In certain embodiments, the promoter is derived in its entirety from a native gene. In certain embodiments, the promoter is composed of different elements derived from different naturally occurring promoters. In certain embodiments, the promoter comprises a synthetic nucleotide sequence. It will be understood by those skilled. in the art that different promoters will direct the expression of a gene in different tissues or cell types, or at different stages of development, or in response to different environmental conditions or to the presence or the absence of a drug or transcriptional co-factor. Ubiquitous, cell-type-specific, tissue-specific, developmental stage-specific, and conditional promoters, for example, drug-responsive promoters (e.g., tetracycline-responsive promoters) are well known to those of skill in the art. Exemplary promoters include, but are not limited to, the phosphoglycerate kinase (PKG) promoter, CAG (composite of the CMV enhancer the chicken beta actin promoter (CBA and the rabbit beta globin intron), NSE (neuronal specific enolase), synapsin or NeuN promoters, the SV40 early promoter, mouse mammary tumor virus LTR promoter; adenovirus major late promoter (Ad MLP), a herpes simplex virus (HSV) promoter, a cytomegalovirus (CMV) promoter such as the CMV immediate early promoter region (CMV IE), SFFV promoter, rous sarcoma virus (RSV) promoter, synthetic promoters, hybrid promoters, and the like. Other promoters can be of human origin or from other species, including from mice. Common promoters include, e.g., the human cytomegalovirus (CMV) immediate early gene promoter, the SV40 early promoter, the Rous sarcoma virus long terminal repeat, [beta]-actin, rat insulin promoter, the phosphoglycerate kinase promoter, the human alpha-1 antitrypsin (hAAT) promoter, the transthyretin promoter, the TBG promoter and other liver-specific, promoters, the desmin promoter and similar muscle-specific promoters, the EF1-alpha promoter, hybrid promoters with multi-tissue specificity, promoters specific for neurons like synapsin and glyceraldehyde-3-phosphate dehydrogenase promoter, all of which are promoters well known and readily available to those of skill in the art, can be used to obtain high-level expression of the coding sequence of interest. In addition, sequences derived from non-viral genes, such as the murine metallothionein gene, will also find use herein. Such promoter sequences are commercially available from, e.g., Stratagene (San Diego, CA). Additional exemplary promoter sequences are described, for example, in WO2018213786A1 (incorporated by reference herein in its entirety).


In some embodiments, the apolipoprotein E enhancer (ApoE) or a functional fragment thereof is used, e.g., to drive expression in the liver. In some embodiments, two copies of the ApoE enhancer or a functional fragment thereof are used. In some embodiments, the ApoE enhancer or functional fragment thereof is used in combination with a promoter, e.g., the human alpha-1 antitrypsin (hAAT) promoter.


In some embodiments, the regulatory sequences impart tissue-specific gene expression capabilities. In some cases, the tissue-specific regulatory sequences bind tissue-specific transcription factors that induce transcription in a tissue specific manner, Various tissue-specific regulatory sequences (e.g., promoters, enhancers, etc.) are known in the art, Exemplary tissue-specific regulatory sequences include, but are not limited to, the following tissue-specific promoters: a liver-specific thyroxin binding globulin (TBG) promoter, a insulin promoter, a glucagon promoter, a somatostatin promoter, a pancreatic polypeptide (PPY) promoter, a synapsin-1 (Syn) promoter, a creatine kinase (MCI( )promoter, a mammalian destnin (DES) promoter, a α-myosin heavy Chain (α-MHC) promoter, or a cardiac Troponin T (cTnT) promoter. Other exemplary promoters include Beta-actin promoter, hepatitis B virus core promoter, Sandig et al., Gene Ther., 3:1002-9 (1996); alpha-fetoprotein (ALT) promoter, Arbuthnot et al., Hum. Gene Ther., 7:1503-14 (1996)), hone osteocalcin promoter (Stein et al., Mol. Biol. Rep., 24:185-96 (1997)); bone sialoprotein promoter (Chen et al., I. Bone Miner. Res. 11:654-64 (1996)), CD2 promoter (Hansal et al., I. Immunol., 161:1063-8 (1998); immunoglobulin heavy chain promoter, T cell receptor α-chain promoter, neuronal. such as neuron-specific enolase (NSE) promoter (Andersen et al., Cell. Md. INeurobiol., 13:503-15 (1993)), neurofilament light-chain gene promoter (Piccioli et al., Proc. Natl. Acad. Sci. USA, 88:5611-5 (1991)), and the neuron-specific vgf gene promoter (Piccioli et al., Neuron, 15:373-84 (1995)), and others. Additional exemplary promoter sequences are described, for example, in U.S. patent Ser. No. 10/300,146 (incorporated herein by reference in its entirety). In some embodiments, a tissue-specific regulatory element, e.g., a tissue-specific promoter, is selected from one known to be operably linked to a gene that is highly expressed in a given tissue, e.g., as measured by RNA-seq or protein expression data, or a combination thereof. Methods for analyzing tissue specificity by expression are taught in Fagerberg et al. Mol Cell Proteomics 13(2):397-406 (2014), which is incorporated herein by reference in its entirety.


In some embodiments, a vector described herein is a multicistronic expression construct. Multicistronic expression constructs include, for example, constructs harboring a first expression cassette, e.g. comprising a first promoter and a first encoding nucleic acid sequence, and a second expression cassette, e.g. comprising a second promoter and a second encoding nucleic acid sequence. Such multicistronic expression constructs may, in some instances, be particularly useful in the delivery of non-translated gene products, such as hairpin RNAs, together with a polypeptide, for example, a gene modifying polypeptide and gene modifying template. In some embodiments, multicistronic expression constructs may exhibit reduced expression levels of one or more of the included transgenes, for example, because of promoter interference or the presence of incompatible nucleic acid elements in close proximity. If a multicistronic expression construct is part of a viral vector, the presence of a self-complementary nucleic acid sequence may; in some instances, interfere with the formation of structures necessary for viral reproduction or packaging.


In some embodiments, the sequence encodes an RNA with a hairpin. In some embodiments, the hairpin RNA is a guide RNA, a template RNA, a shRNA, or a microRNA. In some embodiments, the first promoter is an RNA polymerase 1 promoter. In some embodiments, the first promoter is an RNA polymerase H promoter. In some embodiments, the second promoter is an RNA polymerase Iii promoter. In some embodiments, the second promoter is a. U6 or H1 promoter.


Without wishing to be bound by theory, multicistronic expression constructs may not achieve optimal expression levels as compared to expression systems containing only one cistron. One of the suggested causes of lower expression levels achieved with multicistronic expression constructs comprising two or more promoter elements is the phenomenon of promoter interference (see, e.g., Curtin J A, Dane A P, Swanson A, Alexander I E, Ohm S L. Bidirectional promoter interference between two widely used internal heterologous promoters in a late-generation lentiviral construct. Gene Ther. 2008 March; 15(5):384-90; and Martin-Duque P, Jezzard S, Kaftansis L, Vassaux G. Direct comparison ofthe insulating properties of two genetic elements in an adenoviral vector containing two different expression cassettes. Hum Gene Tiler. 2004 October; 15(10):995-1002; both references incorporated herein by reference for disclosure of promoter interference phenomenon). In some embodiments, the problem of promoter interference may be overcome, e.g., by producing multicistronic expression constructs comprising only one promoter driving transcription of multiple encoding nucleic acid sequences separated by internal ribosomal entry sites, or by separating cistrons comprising their own promoter with transcriptional insulator elements. In some embodiments, single-promoter driven expression of multiple cistrons may result in uneven expression levels of the cistrons. In some embodiments, a promoter cannot efficiently: be isolated and isolation elements may not be compatible with some gene transfer vectors, for example, some retroviral vectors.


MicroRNAs


MicroRNAs (miRNAs) and other small interfering nucleic acids generally regulate gene expression via target RNA transcript cleavageldegradation or translational repression of the target messenger RNA (mRNA). miRNAs may, in some instances, be natively expressed, typically as final 19-25 non-translated RNA products. miRNAs generally exhibit their activity through sequence-.specific interactions with the 3′ untranslated regions (UTR) of target mRNAs. These endogenously expressed miRNAs may form hairpin precursors that are subsequently processed into an miRNA duplex, and further into a mature single stranded miRNA molecule This mature miRNA generally guides a multi protein complex, miRISC, which identifies target 3′ regions of target mRNAs based upon their complementarity to the mature miRNA. Useful transgene products may include, for example, miRNAs or miRNA binding sites that regulate the expression of a linked polypeptide. A non-limiting list of miRNA genes; the products of these genes and their homologues are useful as transgenes or as targets for small interfering nucleic acids (e.g., miRINA sponges, antisense oligonucleotides), e.g., in methods such as those listed in U.S. Ser. No. 10/300,146, 22:2525:48, are herein incorporated by reference. In some embodiments, one or more binding sites for one or more of the foregoing miRINAs are incorporated in a transgene, e.g., a transgene delivered by a rAAV vector, e.g., to inhibit the expression of the transgene in one or more tissues of an animal harboring the transgene. In some embodiments, a binding site may be selected to control the expression of a transgene in a tissue specific manner. For example, binding sites fix the liver-specific miR-122 may be incorporated into a transgene to inhibit expression of that transgene in the liver. Additional exemplary miRNA sequences are described, for example, in U.S. Pat. No. 10,300,146 (incorporated herein by reference in its entirety).


An miR inhibitor or miRNA inhibitor is generally an agent that blocks miRNA expression and/or processing. Examples of such agents include, but are not limited to, microRNA antagonists, microRNA specific antisense, microRNA sponges; and microRNA oligonucleotides (double-stranded, hairpin, short oligonucleotides) that inhibit miRNA interaction with a Drosha complex. MicroRNA inhibitors, e.g., miRNA sponges; can be expressed in cells from transgenes (e.g., as described in Ebert, M. S. Nature Methods, Epub Aug. 12, 2007; incorporated by reference herein in its entirety). In some embodiments, microRNA sponges, or other miR inhibitors, are used with the AAVs. InicroRNA sponges generally specifically inhibit miRNAs through a complementary heptameric seed sequence. In some embodiments, an entire family of miRNAs can be silenced using a single sponge sequence. Other methods for silencing miRNA function. (derepression of miRNA targets) in cells will be apparent to one of ordinary skill in the art.


In some embodiments, a gene modifying system, template RNA, or polypeptide described herein is administered to or is active in (e.g., is more active in) a target tissue, e.g., a first tissue. In some embodiments, the gene modifying system, template RNA, or polypeptide is not administered to or is less active in (e.g., not active in) a non-target tissue. In some embodiments, a gene modifying system, template RNA, or polypeptide described herein is useful for modifying DNA in a target tissue, e.g., a first tissue, (e.g., and not modifying DNA in a non-target tissue).


In some embodiments, a gene modifying system comprises (a) a polypeptide described herein or a nucleic acid encoding the same, (b) a template nucleic acid (e.g., template RNA) described herein, and (c) one or more first tissue-specific expression-control sequences specific to the target tissue, wherein the one or more first tissue-specific expression-control sequences specific to the target tissue are in operative association with (a), (b), or (a) and (b), wherein, when associated with (a), (a) comprises a nucleic acid encoding the polypeptide.


In some embodiments, the nucleic acid in (b) comprises RNA.


In some embodiments, the nucleic acid in (b) comprises DNA.


In some embodiments, the nucleic acid in (b): (i) is single-stranded or comprises a single-stranded segment, e.g., is single-stranded DNA or comprises a single-stranded segment and one or more double stranded segments; (ii) has inverted terminal repeats; or (iii) both (i) and (ii).


In some embodiments, the nucleic acid in (b) is double-stranded or comprises a double-stranded segment.


In some embodiments, (a) comprises a nucleic acid encoding the polypeptide.


In some embodiments, the nucleic acid in (a) comprises RNA.


In some embodiments, the nucleic acid in (a) comprises DNA.


In some embodiments, the nucleic acid in (a): (i) is single-stranded or comprises a single-stranded segment, e.g., is single-stranded DNA or comprises a single-stranded segment and one or more double stranded segments; (ii) has inverted terminal repeats; or (iii) both (i) and (ii).


In some embodiments, the nucleic acid in (a) is double-stranded or comprises a double-stranded segment.


In some embodiments, the nucleic acid in (a), (b), or (a) and (b) is linear.


In some embodiments, the nucleic acid in (a), (b), or (a) and (b) is circular, e.g., a plasmid or minicircle.


In some embodiments, the heterologous object sequence is in operative association with a first promoter.


In some embodiments, the one or more first tissue-specific expression-control sequences comprises a tissue specific promoter.


In some embodiments, the tissue-specific promoter comprises a first promoter in operative association with: (i) the heterologous object sequence, (ii) a nucleic acid encoding the retroviral RT, or (iii) (i) and (ii).


In some embodiments, the one or more first tissue-specific expression-control sequences comprises a tissue-specific microRNA recognition sequence in operative association with: (i) the heterologous object sequence, (ii) a nucleic acid encoding the retroviral RT domain, or (iii) (i) and (ii).


In some embodiments, a system comprises a tissue-specific promoter, and the system further comprises one or more tissue-specific microRNA recognition sequences, wherein: (i) the tissue specific promoter is in operative association with: (I) the heterologous object sequence, (II) a nucleic acid encoding the retroviral RT domain, or (III) (I) and (II); and/or (ii) the one or more tissue-specific microRNA recognition sequences are in operative association with: (I) the heterologous object sequence, (II) a nucleic acid encoding the retroviral RT, or (III) (I) and (II).


In some embodiments, wherein (a) comprises a nucleic acid encoding the polypeptide, the nucleic acid comprises a promoter in operative association with the nucleic acid encoding the polypeptide.


In some embodiments, the nucleic acid encoding the polypeptide comprises one or more second tissue-specific expression-control sequences specific to the target tissue in operative association with the polypeptide coding sequence.


In some embodiments, the one or more second tissue-specific expression-control sequences comprises a tissue specific promoter.


In some embodiments, the tissue-specific promoter is the promoter in operative association with the nucleic acid encoding the polypeptide.


In some embodiments, the one or more second tissue-specific expression-control sequences comprises a tissue-specific microRNA recognition sequence.


In some embodiments, the promoter in operative association with the nucleic acid encoding the polypeptide is a tissue-specific promoter, the system further comprising one or more tissue-specific microRNA recognition sequences.


In some embodiments, a nucleic acid component of a system provided by the invention is a sequence (e.g., encoding the polypeptide or comprising a heterologous object sequence) flanked by untranslated regions (UTRs) that modify protein expression levels. Various 5′ and 3′ UTRs can affect protein expression. For example, in some embodiments, the coding sequence may be preceded by a 5′ UTR that modifies RNA stability or protein translation. In some embodiments, the sequence may be followed by a 3′ UTR that modifies RNA stability or translation. In some embodiments, the sequence may be preceded by a 5′ UTR and followed by a 3′ UTR that modify RNA stability or translation. In some embodiments, the 5′ and/or 3′ UTR may be selected from the 5′ and 3′ UTRs of complement factor 3 (C3) (CACTCCTCCCCATCCTCTCCCTCTGTCCCTCTGTCCCTCTGACCCTGCACTGTCCCAG CACC; SEQ ID NO: 11,004) or orosomucoid 1 (ORM1) (CAGGACACAGCCTTGGATCAGGACAGAGACTTGGGGGCCATCCTGCCCCTCCAACC CGACATGTGTACCTCAGCTTTTTCCCTCACTTGCATCAATAAAGCTTCTGTGTTTGGA ACAGCTAA; SEQ ID NO: 11,005) (Asrani et al. RNA Biology 2018). In certain embodiments, the 5′ UTR is the 5′ UTR from C3 and the 3′ UTR is the 3′ UTR from ORM1. In certain embodiments, a 5′ UTR and 3′ UTR for protein expression, e.g., mRNA (or DNA encoding the RNA) for a gene modifying polypeptide or heterologous object sequence, comprise optimized expression sequences. In some embodiments, the 5′ UTR comprises GGGAAAUAAGAGAGAAAAGAAGAGUAAGAAGAAAUAUAAGAGCCACC (SEQ ID NO: 11,006) and/or the 3′ UTR comprising UGAUAAUAGGCUGGAGCCUCGGUGGCCAUGCUUCUUGCCCCUUGGGCCUCCCCCC AGCCCCUCCUCCCCUUCCUGCACCCGUACCCCCGUGGUCUUUGAAUAAAGUCUGA (SEQ ID NO: 11,007), e.g., as described in Richner et al. Cell 168(6): P1114-1125 (2017), the sequences of which are incorporated herein by reference.


In some embodiments, a 5′ and/or 3′ UTR may be selected to enhance protein expression. In some embodiments, a 5′ and/or 3′ UTR may be selected to modify protein expression such that overproduction inhibition is minimized. In some embodiments, UTRs are around a coding sequence, e.g., outside the coding sequence and in other embodiments proximal to the coding sequence, In some embodiments, additional regulatory elements (e.g., miRNA binding sites, cis-regulatory sites) are included in the UTRs.


In some embodiments, an open reading frame of a gene modifying system, e.g., an ORF of an mRNA (or DNA encoding an mRNA) encoding a gene modifying polypeptide or one or more ORFs of an mRNA (or DNA encoding an mRNA) of a heterologous object sequence, is flanked by a 5′ and/or 3′ untranslated region (UTR) that enhances the expression thereof. In some embodiments, the 5′ UTR of an mRNA component (or transcript produced from a DNA component) of the system comprises the sequence 5′-GGGAAAUAAGAGAGAAAAGAAGAGUAAGAAGAAAUAUAAGAGCCACC-3′; SEQ ID NO: 11,008). In some embodiments, the 3′ UTR of an mRNA component (or transcript produced from a DNA component) of the system comprises the sequence 5′-UGAUAAUAGGCUGGAGCCUCGGUGGCCAUGCUUCUUGCCCCUUGGGCCUCCCCCC AGCCCCUCCUCCCCUUCCUGCACCCGUACCCCCGUGGUCUUUGAAUAAAGUCUGA-3′ (SEQ ID NO: 11,009). This combination of 5′ UTR and 3′ UTR has been shown to result in desirable expression of an operably linked ORF by Richner et al. Cell 168(6): P1114-1125 (2017), the teachings and sequences of which are incorporated herein by reference. In some embodiments, a system described herein comprises a DNA encoding a transcript, wherein the DNA comprises the corresponding 5′ UTR and 3′ UTR sequences, with T substituting for U in the above-listed sequence). In some embodiments, a DNA vector used to produce an RNA component of the system further comprises a promoter upstream of the 5′ UTR for initiating in vitro transcription, e.g., a T7, T3, or SP6 promoter. The 5′ UTR above begins with GGG, which is a suitable start for optimizing transcription using T7 RNA polymerase. For tuning transcription levels and altering the transcription start site nucleotides to fit alternative 5′ UTRs, the teachings of Davidson et al. Pac Symp Biocomput 433-443 (2010) describe T7 promoter variants, and the methods of discovery thereof, that fulfill both of these traits.


Viral Vectors and Components Thereof

Viruses are a useful source of delivery vehicles for the systems described herein, in addition to a source of relevant enzymes or domains as described herein, e.g., as sources of polymerases and polymerase functions used herein, e.g., DNA-dependent DNA polymerase, RNA-dependent RNA polymerase, RNA-dependent DNA polymerase, DNA-dependent RNA polymerase, reverse transcriptase. Some enzymes, e.g., reverse transcriptases, may have multiple activities, e.g., be capable of both RNA-dependent DNA polymerization and DNA-dependent DNA polymerization, e.g., first and second strand synthesis. In some embodiments, the virus used as a gene modifying delivery system or a source of components thereof may be selected from a group as described by Baltimore Bacteriol Rev 35(3):235-241 (1971).


In some embodiments, the virus is selected from a Group I virus, e.g., is a DNA virus and packages dsDNA into virions. In some embodiments, the Group I virus is selected from, e.g., Adenoviruses, Herpesviruses, Poxviruses.


In some embodiments, the virus is selected from a Group II virus, e.g., is a DNA virus and packages ssDNA into virions. In some embodiments, the Group II virus is selected from, e.g., Parvoviruses. In some embodiments, the parvovirus is a dependoparvovirus, e.g., an adeno-associated virus (AAV).


In some embodiments, the virus is selected from a Group III virus, e.g., is an RNA virus and packages dsRNA into virions. In some embodiments, the Group III virus is selected from, e.g., Reoviruses. In some embodiments, one or both strands of the dsRNA contained in such virions is a coding molecule able to serve directly as mRNA upon transduction into a host cell, e.g., can be directly translated into protein upon transduction into a host cell without requiring any intervening nucleic acid replication or polymerization steps.


In some embodiments, the virus is selected from a Group IV virus, e.g., is an RNA virus and packages ssRNA(+) into virions. In some embodiments, the Group IV virus is selected from, e.g., Coronaviruses, Picornaviruses, Togaviruses. In some embodiments, the ssRNA(+) contained in such virions is a coding molecule able to serve directly as mRNA upon transduction into a host cell, e.g., can be directly translated into protein upon transduction into a host cell without requiring any intervening nucleic acid replication or polymerization steps.


In some embodiments, the virus is selected from a Group V virus, e.g., is an RNA virus and packages ssRNA(−) into virions. In some embodiments, the Group V virus is selected from, e.g., Orthomyxoviruses, Rhabdoviruses. In some embodiments, an RNA virus with an ssRNA(−) genome also carries an enzyme inside the virion that is transduced to host cells with the viral genome, e.g., an RNA-dependent RNA polymerase, capable of copying the ssRNA(−) into ssRNA(+) that can be translated directly by the host.


In some embodiments, the virus is selected from a Group VI virus, e.g., is a retrovirus and packages ssRNA(+) into virions. In some embodiments, the Group VI virus is selected from, e.g., retroviruses. In some embodiments, the retrovirus is a lentivirus, e.g., HIV-1, HIV-2, SIV, BIV. In some embodiments, the retrovirus is a spumavirus, e.g., a foamy virus, e.g., HFV, SFV, BFV. In some embodiments, the ssRNA(+) contained in such virions is a coding molecule able to serve directly as mRNA upon transduction into a host cell, e.g., can be directly translated into protein upon transduction into a host cell without requiring any intervening nucleic acid replication or polymerization steps. In some embodiments, the ssRNA(+) is first reverse transcribed and copied to generate a dsDNA genome intermediate from which mRNA can be transcribed in the host cell. In some embodiments, an RNA virus with an ssRNA(+) genome also carries an enzyme inside the virion that is transduced to host cells with the viral genome, e.g., an RNA-dependent DNA polymerase, capable of copying the ssRNA(+) into dsDNA that can be transcribed into mRNA and translated by the host. In some embodiments, the reverse transcriptase from a Group VI retrovirus is incorporated as the reverse transcriptase domain of a gene modifying polypeptide.


In some embodiments, the virus is selected from a Group VII virus, e.g., is a retrovirus and packages dsRNA into virions. In some embodiments, the Group VII virus is selected from, e.g., Hepadnaviruses. In some embodiments, one or both strands of the dsRNA contained in such virions is a coding molecule able to serve directly as mRNA upon transduction into a host cell, e.g., can be directly translated into protein upon transduction into a host cell without requiring any intervening nucleic acid replication or polymerization steps. In some embodiments, one or both strands of the dsRNA contained in such virions is first reverse transcribed and copied to generate a dsDNA genome intermediate from which mRNA can be transcribed in the host cell. In some embodiments, an RNA virus with a dsRNA genome also carries an enzyme inside the virion that is transduced to host cells with the viral genome, e.g., an RNA-dependent DNA polymerase, capable of copying the dsRNA into dsDNA that can be transcribed into mRNA and translated by the host. In some embodiments, the reverse transcriptase from a Group VII retrovirus is incorporated as the reverse transcriptase domain of a gene modifying polypeptide.


In some embodiments, virions used to deliver nucleic acid in this invention may also carry enzymes involved in the process of gene modification. For example, a retroviral virion may contain a reverse transcriptase domain that is delivered into a host cell along with the nucleic acid. In some embodiments, an RNA template may be associated with a gene modifying polypeptide within a virion, such that both are co-delivered to a target cell upon transduction of the nucleic acid from the viral particle. In some embodiments, the nucleic acid in a virion may comprise DNA, e.g., linear ssDNA, linear dsDNA, circular ssDNA, circular dsDNA, minicircle DNA, dbDNA, ceDNA. In some embodiments, the nucleic acid in a virion may comprise RNA, e.g., linear ssRNA, linear dsRNA, circular ssRNA, circular dsRNA. In some embodiments, a viral genome may circularize upon transduction into a host cell, e.g., a linear ssRNA molecule may undergo a covalent linkage to form a circular ssRNA, a linear dsRNA molecule may undergo a covalent linkage to form a circular dsRNA or one or more circular ssRNA. In some embodiments, a viral genome may replicate by rolling circle replication in a host cell. In some embodiments, a viral genome may comprise a single nucleic acid molecule, e.g., comprise a non-segmented genome. In some embodiments, a viral genome may comprise two or more nucleic acid molecules, e.g., comprise a segmented genome. In some embodiments, a nucleic acid in a virion may be associated with one or proteins. In some embodiments, one or more proteins in a virion may be delivered to a host cell upon transduction. In some embodiments, a natural virus may be adapted for nucleic acid delivery by the addition of virion packaging signals to the target nucleic acid, wherein a host cell is used to package the target nucleic acid containing the packaging signals.


In some embodiments, a virion used as a delivery vehicle may comprise a commensal human virus. In some embodiments, a virion used as a delivery vehicle may comprise an anellovirus, the use of which is described in WO2018232017A1, which is incorporated herein by reference in its entirety.


AAV Administration

In some embodiments, an adeno-associated virus (AAV) is used in conjunction with the system, template nucleic acid, and/or polypeptide described herein. In some embodiments, an AAV is used to deliver, administer, or package the system, template nucleic acid, and/or polypeptide described herein. In some embodiments, the AAV is a recombinant AAV (rAAV).


In some embodiments, a system comprises (a) a polypeptide described herein or a nucleic acid encoding the same, (b) a template nucleic acid (e.g., template RNA) described herein, and (c) one or more first tissue-specific expression-control sequences specific to the target tissue, wherein the one or more first tissue-specific expression-control sequences specific to the target tissue are in operative association with (a), (b), or (a) and (b), wherein, when associated with (a), (a) comprises a nucleic acid encoding the polypeptide.


In some embodiments, a system described herein further comprises a first recombinant adeno-associated virus (rAAV) capsid protein; wherein the at least one of (a) or (b) is associated with the first rAAV capsid protein, wherein at least one of (a) or (b) is flanked by AAV inverted terminal repeats (ITRs).


In some embodiments, (a) and (b) are associated with the first rAAV capsid protein.


In some embodiments, (a) and (b) are on a single nucleic acid.


In some embodiments, the system further comprises a second rAAV capsid protein, wherein at least one of (a) or (b) is associated with the second rAAV capsid protein, and wherein the at least one of (a) or (b) associated with the second rAAV capsid protein is different from the at least one of (a) or (b) is associated with the first rAAV capsid protein.


In some embodiments, the at least one of (a) or (b) is associated with the first or second rAAV capsid protein is dispersed in the interior of the first or second rAAV capsid protein, which first or second rAAV capsid protein is in the form of an AAV capsid particle.


In some embodiments, the system further comprises a nanoparticle, wherein the nanoparticle is associated with at least one of (a) or (b).


In some embodiments, (a) and (b), respectively are associated with: a) a first rAAV capsid protein and a second rAAV capsid protein; b) a nanoparticle and a first rAAV capsid protein; c) a first rAAV capsid protein; d) a first adenovirus capsid protein; e) a first nanoparticle and a second nanoparticle; or f) a first nanoparticle.


Viral vectors are useful for delivering all or part of a system provided by the invention, e.g., for use in methods provided by the invention. Systems derived from different viruses have been employed for the delivery of polypeptides or nucleic acids; for example: integrase-deficient lentivirus, adenovirus, adeno-associated virus (AAV), herpes simplex virus, and baculovirus (reviewed in Hodge et al. Hum Gene Ther 2017; Narayanavari et al. Crit Rev Biochem Mol Biol 2017; Boehme et al. Curr Gene Ther 2015).


Adenoviruses are common viruses that have been used as gene delivery vehicles given well-defined biology, genetic stability, high transduction efficiency, and ease of large-scale production (see, for example, review by Lee et al. Genes & Diseases 2017). They possess linear dsDNA genomes and come in a variety of serotypes that differ in tissue and cell tropisms. In order to prevent replication of infectious virus in recipient cells, adenovirus genomes used for packaging are deleted of some or all endogenous viral proteins, which are provided in trans in viral production cells. This renders the genomes helper-dependent, meaning they can only be replicated and packaged into viral particles in the presence of the missing components provided by so-called helper functions. A helper-dependent adenovirus system with all viral ORFs removed may be compatible with packaging foreign DNA of up to −37 kb (Parks et al. J Virol 1997). In some embodiments, an adenoviral vector is used to deliver DNA corresponding to the polypeptide or template component of the gene modifying system, or both are contained on separate or the same adenoviral vector. In some embodiments, the adenovirus is a helper-dependent adenovirus (HD-AdV) that is incapable of self-packaging. In some embodiments, the adenovirus is a high-capacity adenovirus (HC-AdV) that has had all or a substantial portion of endogenous viral ORFs deleted, while retaining the necessary sequence components for packaging into adenoviral particles. For this type of vector, the only adenoviral sequences required for genome packaging are noncoding sequences: the inverted terminal repeats (ITRs) at both ends and the packaging signal at the 5′-end (Jager et al. Nat Protoc 2009). In some embodiments, the adenoviral genome also comprises stuffer DNA to meet a minimal genome size for optimal production and stability (see, for example, Hausl et al. Mol Ther 2010). In some embodiments, an adenovirus is used to deliver a gene modifying system to the liver.


In some embodiments, an adenovirus is used to deliver a gene modifying system to HSCs, e.g., HDAd5/35++. HDAd5/35++is an adenovirus with modified serotype 35 fibers that de-target the vector from the liver (Wang et al. Blood Adv 2019). In some embodiments, the adenovirus that delivers a gene modifying system to HSCs utilizes a receptor that is expressed specifically on primitive HSCs, e.g., CD46.


Adeno-associated viruses (AAV) belong to the parvoviridae family and more specifically constitute the dependoparvovirus genus. The AAV genome is composed of a linear single-stranded DNA molecule which contains approximately 4.7 kilobases (kb) and consists of two major open reading frames (ORFs) encoding the non-structural Rep (replication) and structural Cap (capsid) proteins. A second ORF within the cap gene was identified that encodes the assembly-activating protein (AAP). The DNAs flanking the AAV coding regions are two cis-acting inverted terminal repeat (ITR) sequences, approximately 145 nucleotides in length, with interrupted palindromic sequences that can be folded into energetically stable hairpin structures that function as primers of DNA replication. In addition to their role in DNA replication, the ITR sequences have been shown to be involved in viral DNA integration into the cellular genome, rescue from the host genome or plasmid, and encapsidation of viral nucleic acid into mature virions (Muzyczka, (1992) Curr. Top. Micro. Immunol. 158:97-129). In some embodiments, one or more gene modifying nucleic acid components is flanked by ITRs derived from AAV for viral packaging. See, e.g., WO2019113310.


In some embodiments, one or more components of the gene modifying system are carried via at least one AAV vector. In some embodiments, the at least one AAV vector is selected for tropism to a particular cell, tissue, organism. In some embodiments, the AAV vector is pseudotyped, e.g., AAV2/8, wherein AAV2 describes the design of the construct but the capsid protein is replaced by that from AAV8. It is understood that any of the described vectors could be pseudotype derivatives, wherein the capsid protein used to package the AAV genome is derived from that of a different AAV serotype. Without wishing to be limited in vector choice, a list of exemplary AAV serotypes can be found in Table 18. In some embodiments, an AAV to be employed for gene modifying may be evolved for novel cell or tissue tropism as has been demonstrated in the literature (e.g., Davidsson et al. Proc Natl Acad Sci USA 2019).


In some embodiments, the AAV delivery vector is a vector which has two AAV inverted terminal repeats (ITRs) and a nucleotide sequence of interest (for example, a sequence coding for a gene modifying polypeptideor a DNA template, or both), each of said ITRs having an interrupted (or noncontiguous) palindromic sequence, i.e., a sequence composed of three segments: a first segment and a last segment that are identical when read 5′—>3′ but hybridize when placed against each other, and a segment that is different that separates the identical segments. See, for example, WO2012123430.


Conventionally, AAV virions with capsids are produced by introducing a plasmid or plasmids encoding the rAAV or scAAV genome, Rep proteins, and Cap proteins (Grimm et al, 1998). Upon introduction of these helper plasmids in trans, the AAV genome is “rescued” (i.e., released and subsequently recovered) from the host genome, and is further encapsidated to produce infectious AAV. In some embodiments, one or more gene modifying nucleic acids are packaged into AAV particles by introducing the ITR-flanked nucleic acids into a packaging cell in conjunction with the helper functions.


In some embodiments, the AAV genome is a so called self-complementary genome (referred to as scAAV), such that the sequence located between the ITRs contains both the desired nucleic acid sequence (e.g., DNA encoding the gene modifying polypeptide or template, or both) in addition to the reverse complement of the desired nucleic acid sequence, such that these two components can fold over and self-hybridize. In some embodiments, the self-complementary modules are separated by an intervening sequence that permits the DNA to fold back on itself, e.g., forms a stem-loop. An scAAV has the advantage of being poised for transcription upon entering the nucleus, rather than being first dependent on ITR priming and second-strand synthesis to form dsDNA. In some embodiments, one or more gene modifying components is designed as an scAAV, wherein the sequence between the AAV ITRs contains two reverse complementing modules that can self-hybridize to create dsDNA.


In some embodiments, nucleic acid (e.g., encoding a polypeptide, or a template, or both) delivered to cells is closed-ended, linear duplex DNA (CELiD DNA or ceDNA). In some embodiments, ceDNA is derived from the replicative form of the AAV genome (Li et al. PLoS One 2013). In some embodiments, the nucleic acid (e.g., encoding a polypeptide, or a template DNA, or both) is flanked by ITRs, e.g., AAV ITRs, wherein at least one of the ITRs comprises a terminal resolution site and a replication protein binding site (sometimes referred to as a replicative protein binding site). In some embodiments, the ITRs are derived from an adeno-associated virus, e.g., AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, or a combination thereof. In some embodiments, the ITRs are symmetric. In some embodiments, the ITRs are asymmetric. In some embodiments, at least one Rep protein is provided to enable replication of the construct. In some embodiments, the at least one Rep protein is derived from an adeno-associated virus, e.g., AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, or a combination thereof. In some embodiments, ceDNA is generated by providing a production cell with (i) DNA flanked by ITRs, e.g., AAV ITRs, and (ii) components required for ITR-dependent replication, e.g., AAV proteins Rep78 and Rep52 (or nucleic acid encoding the proteins). In some embodiments, ceDNA is free of any capsid protein, e.g., is not packaged into an infectious AAV particle. In some embodiments, ceDNA is formulated into LNPs (see, for example, WO2019051289A1).


In some embodiments, the ceDNA vector consists of two self-complementary sequences, e.g., asymmetrical or symmetrical or substantially symmetrical ITRs as defined herein, flanking said expression cassette, wherein the ceDNA vector is not associated with a capsid protein. In some embodiments, the ceDNA vector comprises two self-complementary sequences found in an AAV genome, where at least one ITR comprises an operative Rep-binding element (RBE) (also sometimes referred to herein as “RBS”) and a terminal resolution site (trs) of AAV or a functional variant of the RBE. See, for example, WO2019113310.


In some embodiments, the AAV genome comprises two genes that encode four replication proteins and three capsid proteins, respectively. In some embodiments, the genes are flanked on either side by 145-bp inverted terminal repeats (ITRs). In some embodiments, the virion comprises up to three capsid proteins (Vp1, Vp2, and/or Vp3), e.g., produced in a 1:1:10 ratio. In some embodiments, the capsid proteins are produced from the same open reading frame and/or from differential splicing (Vp1) and alternative translational start sites (Vp2 and Vp3, respectively). Generally, Vp3 is the most abundant subunit in the virion and participates in receptor recognition at the cell surface defining the tropism of the virus. In some embodiments, Vp1 comprises a phospholipase domain, e.g., which functions in viral infectivity, in the N-terminus of Vp1.


In some embodiments, packaging capacity of the viral vectors limits the size of the gene modifying system that can be packaged into the vector. For example, the packaging capacity of the AAVs can be about 4.5 kb (e.g., about 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, or 6.0 kb), e.g., including one or two inverted terminal repeats (ITRs), e.g., 145 base ITRs.


In some embodiments, recombinant AAV (rAAV) comprises cis-acting 145-bp ITRs flanking vector transgene cassettes, e.g., providing up to 4.5 kb for packaging of foreign DNA. Subsequent to infection, rAAV can, in some instances, express a fusion protein of the invention and persist without integration into the host genome by existing episomally in circular head-to-tail concatemers. rAAV can be used, for example, in vitro and in vivo. In some embodiments, AAV-mediated gene delivery requires that the length of the coding sequence of the gene is equal or greater in size than the wild-type AAV genome.


AAV delivery of genes that exceed this size and/or the use of large physiological regulatory elements can be accomplished, for example, by dividing the protein(s) to be delivered into two or more fragments. In some embodiments, the N-terminal fragment is fused to an intein-N sequence. In some embodiments, the C-terminal fragment is fused to an intein-C sequence. In embodiments, the fragments are packaged into two or more AAV vectors.


In some embodiments, dual AAV vectors are generated by splitting a large transgene expression cassette in two separate halves (5′ and 3′ ends, or head and tail), e.g., wherein each half of the cassette is packaged in a single AAV vector (of <5 kb). The re-assembly of the full-length transgene expression cassette can, in some embodiments, then be achieved upon co-infection of the same cell by both dual AAV vectors. In some embodiments, co-infection is followed by one or more of: (1) homologous recombination (HR) between 5′ and 3′ genomes (dual AAV overlapping vectors); (2) ITR-mediated tail-to-head concatemerization of 5′ and 3′ genomes (dual AAV trans-splicing vectors); and/or (3) a combination of these two mechanisms (dual AAV hybrid vectors). In some embodiments, the use of dual AAV vectors in vivo results in the expression of full-length proteins. In some embodiments, the use of the dual AAV vector platform represents an efficient and viable gene transfer strategy for transgenes of greater than about 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, or 5.0 kb in size. In some embodiments, AAV vectors can also be used to transduce cells with target nucleic acids, e.g., in the in vitro production of nucleic acids and peptides. In some embodiments, AAV vectors can be used for in vivo and ex vivo gene therapy procedures (see, e.g., West et al., Virology 160:38-47 (1987); U.S. Pat. No. 4,797,368; WO 93/24641; Kotin, Human Gene Therapy 5:793-801 (1994); Muzyczka, J. Clin. Invest.94:1351 (1994); each of which is incorporated herein by reference in their entirety). The construction of recombinant AAV vectors is described in a number of publications, including U.S. Pat. No. 5,173,414; Tratschin et al., Mol. Cell. Biol. 5:3251-3260 (1985); Tratschin, et al., Mol. Cell. Biol. 4:2072-2081 (1984); Hermonat & Muzyczka, PNAS 81:6466-6470 (1984); and Samulski et al., J. Viro1.63:03822-3828 (1989) (incorporated by reference herein in their entirety).


In some embodiments, a gene modifying polypeptide described herein (e.g., with or without one or more guide nucleic acids) can be delivered using AAV, lentivirus, adenovirus or other plasmid or viral vector types, in particular, using formulations and doses from, for example, U.S. Pat. No. 8,454,972 (formulations, doses for adenovirus), U.S. Pat. No. 8,404,658 (formulations, doses for AAV) and U.S. Pat. No. 5,846,946 (formulations, doses for DNA plasmids) and from clinical trials and publications regarding the clinical trials involving lentivirus, AAV and adenovirus. For example, for AAV, the route of administration, formulation and dose can be as described in U.S. Pat. No. 8,454,972 and as in clinical trials involving AAV. For adenovirus, the route of administration, formulation and dose can be as described in U.S. Pat. No. 8,404,658 and as in clinical trials involving adenovirus. For plasmid delivery, the route of administration, formulation and dose can be as described in U.S. Pat. No. 5,846,946 and as in clinical studies involving plasmids. Doses can be based on or extrapolated to an average 70 kg individual (e.g. a male adult human), and can be adjusted for patients, subjects, mammals of different weight and species. Frequency of administration is within the ambit of the medical or veterinary practitioner (e.g., physician, veterinarian), depending on usual factors including the age, sex, general health, other conditions of the patient or subject and the particular condition or symptoms being addressed. In some embodiments, the viral vectors can be injected into the tissue of interest. For cell-type specific gene modifying, the expression of the gene modifying polypeptide and optional guide nucleic acid can, in some embodiments, be driven by a cell-type specific promoter.


In some embodiments, AAV allows for low toxicity, for example, due to the purification method not requiring ultracentrifugation of cell particles that can activate the immune response. In some embodiments, AAV allows low probability of causing insertional mutagenesis, for example, because it does not substantially integrate into the host genome.


In some embodiments, AAV has a packaging limit of about 4.4, 4.5, 4.6, 4.7, or 4.75 kb. In some embodiments, a gene modifying polypeptide-encoding sequence, promoter, and transcription terminator can fit into a single viral vector. SpCas9 (4.1 kb) may, in some instances, be difficult to package into AAV. Therefore, in some embodiments, a gene modifying polypeptide coding sequence is used that is shorter in length than other gene modifying polypeptide coding sequences or base editors. In some embodiments, the gene modifying polypeptide encoding sequences are less than about 4.5 kb, 4.4 kb, 4.3 kb, 4.2 kb, 4.1 kb, 4 kb, 3.9 kb, 3.8 kb, 3.7 kb, 3.6 kb, 3.5 kb, 3.4 kb, 3.3 kb, 3.2 kb, 3.1 kb, 3 kb, 2.9 kb, 2.8 kb, 2.7 kb, 2.6 kb, 2.5 kb, 2 kb, or 1.5 kb.


An AAV can be AAV1, AAV2, AAVS or any combination thereof. In some embodiments, the type of AAV is selected with respect to the cells to be targeted; e.g., AAV serotypes 1, 2, 5 or a hybrid capsid AAV1, AAV2, AAV5 or any combination thereof can be selected for targeting brain or neuronal cells; or AAV4 can be selected for targeting cardiac tissue. In some embodiments, AAV8 is selected for delivery to the liver. Exemplary AAV serotypes as to these cells are described, for example, in Grimm, D. et al, J. Viro1.82:5887-5911 (2008) (incorporated herein by reference in its entirety). In some embodiments, AAV refers all serotypes, subtypes, and naturally-occurring AAV as well as recombinant AAV. AAV may be used to refer to the virus itself or a derivative thereof. In some embodiments, AAV includes AAV1, AAV2, AAV3, AAV3B, AAV4, AAV5, AAV6, AAV6.2, AAV7, AAVrh.64R1, AAVhu.37, AAVrh.8, AAVrh.32.33, AAV8, AAV9, AAV-DJ, AAV2/8, AAVrh10, AAVLK03, AV10, AAV11, AAV 12, rhlO, and hybrids thereof, avian AAV, bovine AAV, canine AAV, equine AAV, primate AAV, non-primate AAV, and ovine AAV. The genomic sequences of various serotypes of AAV, as well as the sequences of the native terminal repeats (TRs), Rep proteins, and capsid subunits are known in the art. Such sequences may be found in the literature or in public databases such as GenBank. Additional exemplary AAV serotypes are listed in Table 18.









TABLE 18







Exemplary AAV serotypes.









Target Tissue
Vehicle
Reference





Liver
AAV (AAV81, AAVrh.81,
1. Wang et al., Mol. Ther. 18,



AAVhu.371, AAV2/8,
118-25 (2010)



AAV2/rh102, AAV9, AAV2,
2. Ginn et al., JHEPReports,



NP403, NP592,3, AAV3B5,
100065 (2019)



AAV-DJ4, AAV-LK014,
3. Paulk et al., Mol. Ther. 26,



AAV-LK024, AAV-LK034,
289-303 (2018).



AAV-LK194, AAV57
4. L. Lisowski et al., Nature.



Adenovirus (Ad5, HC-AdV6)
506, 382-6 (2014).




5. L. Wang et al., Mol. Ther.




23, 1877-87 (2015).




6. Hausl MolTher (2010)




7. Davidoff et al., Mol. Ther.




11, 875-88 (2005)


Lung
AAV (AAV4, AAV5,
1. Duncan et al., MolTher



AAV61, AAV9, H222)

Methods Clin Dev (2018)



Adenovirus (Ad5, Ad3,
2. Cooney et al., Am J Respir



Ad21, Ad14)3
Cell Mol Biol (2019)




3. Li et al., MolTherMethods




Clin Dev (2019)


Skin
AAV (AAV61, AAV-LK192)
1. Petek et al., Mol. Ther.




(2010)




2. L. Lisowski et al., Nature.




506, 382-6 (2014).


HSCs
Adenovirus (HDAd5/35++)
Wang et al. BloodAdv (2019)









In some embodiments, a pharmaceutical composition (e.g., comprising an AAV as described herein) has less than 10% empty capsids, less than 8% empty capsids, less than 7% empty capsids, less than 5% empty capsids, less than 3% empty capsids, or less than 1% empty capsids. In some embodiments, the pharmaceutical composition has less than about 5% empty capsids. In some embodiments, the number of empty capsids is below the limit of detection. In some embodiments, it is advantageous for the pharmaceutical composition to have low amounts of empty capsids, e.g., because empty capsids may generate an adverse response (e.g., immune response, inflammatory response, liver response, and/or cardiac response), e.g., with little or no substantial therapeutic benefit.


In some embodiments, the residual host cell protein (rHCP) in the pharmaceutical composition is less than or equal to 100 ng/ml rHCP per 1×1013 vg/ml, e.g., less than or equal to 40 ng/ml rHCP per 1×1013 vg/ml or 1-50 ng/ml rHCP per 1×1013 vg/ml. In some embodiments, the pharmaceutical composition comprises less than 10 ng rHCP per 1.0×1013 vg, or less than 5 ng rHCP per 1.0×1013 vg, less than 4 ng rHCP per 1.0×1013 vg, or less than 3 ng rHCP per 1.0×1013 vg, or any concentration in between. In some embodiments, the residual host cell DNA (hcDNA) in the pharmaceutical composition is less than or equal to 5×106 pg/ml hcDNA per 1×1013 vg/ml, less than or equal to 1.2×106 pg/ml hcDNA per 1×1013 vg/ml, or 1 ×105 pg/ml hcDNA per 1×1013 vg/ml. In some embodiments, the residual host cell DNA in said pharmaceutical composition is less than 5.0×105 pg per 1×1013 vg, less than 2.0×105 pg per 1.0×1013 vg, less than 1.1×105 pg per 1.0×1013 vg, less than 1.0×105 pg hcDNA per 1.0×1013 vg, less than 0.9×105 pg hcDNA per 1.0×1013 vg, less than 0.8×105 pg hcDNA per 1.0×1013 vg, or any concentration in between.


In some embodiments, the residual plasmid DNA in the pharmaceutical composition is less than or equal to 1.7×105 pg/ml per 1.0×1013 vg/ml, or 1×105 pg/ml per 1×1.0×1013 vg/ml, or 1.7×106 pg/ml per 1.0×1013 vg/ml. In some embodiments, the residual DNA plasmid in the pharmaceutical composition is less than 10.0×105 pg by 1.0×1013 vg, less than 8.0×105 pg by 1.0×1013 vg or less than 6.8×105 pg by 1.0×1013 vg. In embodiments, the pharmaceutical composition comprises less than 0.5 ng per 1.0×1013 vg, less than 0.3 ng per 1.0 ×1013 vg, less than 0.22 ng per 1.0×1013 vg or less than 0.2 ng per 1.0×1013 vg or any intermediate concentration of bovine serum albumin (BSA). In embodiments, the benzonase in the pharmaceutical composition is less than 0.2 ng by 1.0×1013 vg, less than 0.1 ng by 1.0×1013 vg, less than 0.09 ng by 1.0×1013 vg, less than 0.08 ng by 1.0×1013 vg or any intermediate concentration. In embodiments, Poloxamer 188 in the pharmaceutical composition is about 10 to 150 ppm, about 15 to 100 ppm or about 20 to 80 ppm. In embodiments, the cesium in the pharmaceutical composition is less than 50 pg/g (ppm), less than 30 pg/g (ppm) or less than 20 pg/g (ppm) or any intermediate concentration.


In embodiments, the pharmaceutical composition comprises total impurities, e.g., as determined by SDS-PAGE, of less than 10%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or any percentage in between. In embodiments, the total purity, e.g., as determined by SDS-PAGE, is greater than 90%, greater than 92%, greater than 93%, greater than 94%, greater than 95%, greater than 96%, greater than 97%, greater than 98%, or any percentage in between. In embodiments, no single unnamed related impurity, e.g., as measured by SDS-PAGE, is greater than 5%, greater than 4%, greater than 3% or greater than 2%, or any percentage in between. In embodiments, the pharmaceutical composition comprises a percentage of filled capsids relative to total capsids (e.g., peak 1+peak 2 as measured by analytical ultracentrifugation) of greater than 85%, greater than 86%, greater than 87%, greater than 88%, greater than 89%, greater than 90%, greater than 91%, greater than 91.9%, greater than 92%, greater than 93%, or any percentage in between. In embodiments of the pharmaceutical composition, the percentage of filled capsids measured in peak 1 by analytical ultracentrifugation is 20-80%, 25-75%, 30-75%, 35-75%, or 37.4-70.3%. In embodiments of the pharmaceutical composition, the percentage of filled capsids measured in peak 2 by analytical ultracentrifugation is 20-80%, 20-70%, 22-65%, 24-62%, or 24.9-60.1%.


In one embodiment, the pharmaceutical composition comprises a genomic titer of 1.0 to 5.0×1013 vg/mL, 1.2 to 3.0×1013 vg/mL or 1.7 to 2.3×1013 vg/ml. In one embodiment, the pharmaceutical composition exhibits a biological load of less than 5 CFU/mL, less than 4 CFU/mL, less than 3 CFU/mL, less than 2 CFU/mL or less than 1 CFU/mL or any intermediate contraction. In embodiments, the amount of endotoxin according to USP, for example, USP <85>(incorporated by reference in its entirety) is less than 1.0 EU/mL, less than 0.8 EU/mL or less than 0.75 EU/mL. In embodiments, the osmolarity of a pharmaceutical composition according to USP, for example, USP <785>(incorporated by reference in its entirety) is 350 to 450 mOsm/kg, 370 to 440 mOsm/kg or 390 to 430 mOsm/kg. In embodiments, the pharmaceutical composition contains less than 1200 particles that are greater than 25 μm per container, less than 1000 particles that are greater than 25 μm per container, less than 500 particles that are greater than 25 μm per container or any intermediate value. In embodiments, the pharmaceutical composition contains less than 10,000 particles that are greater than 10 μm per container, less than 8000 particles that are greater than 10 μm per container or less than 600 particles that are greater than 10 μm per container.


In one embodiment, the pharmaceutical composition has a genomic titer of 0.5 to 5.0×1013 vg/mL, 1.0 to 4.0×1013 vg/mL, 1.5 to 3.0×1013 vg/ml or 1.7 to 2.3×1013 vg/ml. In one embodiment, the pharmaceutical composition described herein comprises one or more of the following: less than about 0.09 ng benzonase per 1.0×1013 vg, less than about 30 pg/g (ppm) of cesium, about 20 to 80 ppm Poloxamer 188, less than about 0.22 ng BSA per 1.0×1013 vg, less than about 6.8×105 pg of residual DNA plasmid per 1.0×1013 vg, less than about 1.1×105 pg of residual hcDNA per 1.0×1013 vg, less than about 4 ng of rHCP per 1.0×1013 vg, pH 7.7 to 8.3, about 390 to 430 mOsm/kg, less than about 600 particles that are >25 μm in size per container, less than about 6000 particles that are >10 μm in size per container, about 1.7×1013-2.3×1013 vg/mL genomic titer, infectious titer of about 3.9×108 to 8.4×1010 IU per 1.0×1013 vg, total protein of about 100-300 μg per 1.0×1013 vg, mean survival of >24 days in A7SMA mice with about 7.5×1013 vg/kg dose of viral vector, about 70 to 130% relative potency based on an in vitro cell based assay and/or less than about 5% empty capsid. In various embodiments, the pharmaceutical compositions described herein comprise any of the viral particles discussed here, retain a potency of between ±20%, between ±15%, between ±10% or within ±5% of a reference standard. In some embodiments, potency is measured using a suitable in vitro cell assay or in vivo animal model.


Additional methods of preparation, characterization, and dosing AAV particles are taught in WO2019094253, which is incorporated herein by reference in its entirety.


Additional rAAV constructs that can be employed consonant with the invention include those described in Wang et al 2019, available at://doi.org/10.1038/s41573-019-0012-9, including Table 1 thereof, which is incorporated by reference in its entirety.


Lipid Nanoparticles

The methods and systems provided herein may employ any suitable carrier or delivery modality, including, in certain embodiments, lipid nanoparticles (LNPs). Lipid nanoparticles, in some embodiments, comprise one or more ionic lipids, such as non-cationic lipids (e.g., neutral or anionic, or zwitterionic lipids); one or more conjugated lipids (such as PEG-conjugated lipids or lipids conjugated to polymers described in Table 5 of WO2019217941; incorporated herein by reference in its entirety); one or more sterols (e.g., cholesterol); and, optionally, one or more targeting molecules (e.g., conjugated receptors, receptor ligands, antibodies); or combinations of the foregoing.


Lipids that can be used in nanoparticle formations (e.g., lipid nanoparticles) include, for example those described in Table 4 of WO2019217941, which is incorporated by reference—e.g., a lipid-containing nanoparticle can comprise one or more of the lipids in Table 4 of WO2019217941. Lipid nanoparticles can include additional elements, such as polymers, such as the polymers described in Table 5 of WO2019217941, incorporated by reference.


In some embodiments, conjugated lipids, when present, can include one or more of PEG-diacylglycerol (DAG) (such as 1-(monomethoxy-polyethyleneglycol)-2,3-dimyristoylglycerol (PEG-DMG)), PEG-dialkyloxypropyl (DAA), PEG-phospholipid, PEG-ceramide (Cer), a pegylated phosphatidylethanoloamine (PEG-PE), PEG succinate diacylglycerol (PEGS-DAG) (such as 4-O-(2′,3′-di(tetradecanoyloxy)propyl-1-O-(w-methoxy(polyethoxy)ethyl) butanedioate (PEG-S-DMG)), PEG dialkoxypropylcarbam, N-(carbonyl-methoxypoly ethylene glycol 2000)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine sodium salt, and those described in Table 2 of WO2019051289 (incorporated by reference), and combinations of the foregoing.


In some embodiments, sterols that can be incorporated into lipid nanoparticles include one or more of cholesterol or cholesterol derivatives, such as those in WO2009/127060 or US2010/0130588, which are incorporated by reference. Additional exemplary sterols include phytosterols, including those described in Eygeris et al (2020), dx.doi.org/10.1021/acs.nanolett.0c01386, incorporated herein by reference.


In some embodiments, the lipid particle comprises an ionizable lipid, a non-cationic lipid, a conjugated lipid that inhibits aggregation of particles, and a sterol. The amounts of these components can be varied independently and to achieve desired properties. For example, in some embodiments, the lipid nanoparticle comprises an ionizable lipid is in an amount from about 20 mol % to about 90 mol % of the total lipids (in other embodiments it may be 20-70% (mol), 30-60% (mol) or 40-50% (mol); about 50 mol % to about 90 mol % of the total lipid present in the lipid nanoparticle), a non-cationic lipid in an amount from about 5 mol % to about 30 mol % of the total lipids, a conjugated lipid in an amount from about 0.5 mol % to about 20 mol % of the total lipids, and a sterol in an amount from about 20 mol % to about 50 mol % of the total lipids. The ratio of total lipid to nucleic acid (e.g., encoding the gene modifying polypeptide or template nucleic acid) can be varied as desired. For example, the total lipid to nucleic acid (mass or weight) ratio can be from about 10:1 to about 30:1.


In some embodiments, an ionizable lipid may be a cationic lipid, an ionizable cationic lipid, e.g., a cationic lipid that can exist in a positively charged or neutral form depending on pH, or an amine-containing lipid that can be readily protonated. In some embodiments, the cationic lipid is a lipid capable of being positively charged, e.g., under physiological conditions. Exemplary cationic lipids include one or more amine group(s) which bear the positive charge. In some embodiments, the lipid particle comprises a cationic lipid in formulation with one or more of neutral lipids, ionizable amine-containing lipids, biodegradable alkyn lipids, steroids, phospholipids including polyunsaturated lipids, structural lipids (e.g., sterols), PEG, cholesterol and polymer conjugated lipids. In some embodiments, the cationic lipid may be an ionizable cationic lipid. An exemplary cationic lipid as disclosed herein may have an effective pKa over 6.0. In embodiments, a lipid nanoparticle may comprise a second cationic lipid having a different effective pKa (e.g., greater than the first effective pKa), than the first cationic lipid. A lipid nanoparticle may comprise between 40 and 60 mol percent of a cationic lipid, a neutral lipid, a steroid, a polymer conjugated lipid, and a therapeutic agent, e.g., a nucleic acid (e.g., RNA) described herein (e.g., a template nucleic acid or a nucleic acid encoding a gene modifying polypeptide), encapsulated within or associated with the lipid nanoparticle. In some embodiments, the nucleic acid is co-formulated with the cationic lipid. The nucleic acid may be adsorbed to the surface of an LNP, e.g., an LNP comprising a cationic lipid. In some embodiments, the nucleic acid may be encapsulated in an LNP, e.g., an LNP comprising a cationic lipid. In some embodiments, the lipid nanoparticle may comprise a targeting moiety, e.g., coated with a targeting agent. In embodiments, the LNP formulation is biodegradable. In some embodiments, a lipid nanoparticle comprising one or more lipid described herein, e.g., Formula (i), (ii), (ii), (vii) and/or (ix) encapsulates at least 1%, at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 92%, at least 95%, at least 97%, at least 98% or 100% of an RNA molecule, e.g., template RNA and/or a mRNA encoding the gene modifying polypeptide.


In some embodiments, the lipid to nucleic acid ratio (mass/mass ratio; w/w ratio) can be in the range of from about 1:1 to about 25:1, from about 10:1 to about 14:1, from about 3:1 to about 15:1, from about 4:1 to about 10:1, from about 5:1 to about 9:1, or about 6:1 to about 9:1. The amounts of lipids and nucleic acid can be adjusted to provide a desired N/P ratio, for example, N/P ratio of 3, 4, 5, 6, 7, 8, 9, 10 or higher. Generally, the lipid nanoparticle formulation's overall lipid content can range from about 5 mg/ml to about 30 mg/mL.


Exemplary ionizable lipids that can be used in lipid nanoparticle formulations include, without limitation, those listed in Table 1 of WO2019051289, incorporated herein by reference. Additional exemplary lipids include, without limitation, one or more of the following formulae: X of US2016/0311759; I of US20150376115 or in US2016/0376224; I, II or III of US20160151284; I, IA, II, or IIA of US20170210967; I-c of US20150140070; A of US2013/0178541; I of US2013/0303587 or US2013/0123338; I of US2015/0141678; II, III, IV, or V of US2015/0239926; I of US2017/0119904; I or II of WO2017/117528; A of US2012/0149894; A of US2015/0057373; A of WO2013/116126; A of US2013/0090372; A of US2013/0274523; A of US2013/0274504; A of US2013/0053572; A of WO2013/016058; A of WO2012/162210; I of US2008/042973; I, II, III, or IV of US2012/01287670; I or II of US2014/0200257; I, II, or III of US2015/0203446; I or III of US2015/0005363; I, IA, IB, IC, ID, II, IIA, IIB, IIC, IID, or III-XXIV of US2014/0308304; of US2013/0338210; I, II, III, or IV of WO2009/132131; A of US2012/01011478; I or XXXV of US2012/0027796; XIV or XVII of US2012/0058144; of US2013/0323269; I of US2011/0117125; I, II, or III of US2011/0256175; I, II, III, IV, V, VI, VII, VIII, IX, X, XI, XII of US2012/0202871; I, II, III, IV, V, VI, VII, VIII, X, XII, XIII, XIV, XV, or XVI of US2011/0076335; I or II of US2006/008378; I of US2013/0123338; I or X-A-Y-Z of US2015/0064242; XVI, XVII, or XVIII of US2013/0022649; I, II, or III of US2013/0116307; I, II, or III of US2013/0116307; I or II of US2010/0062967; I-X of US2013/0189351; I of US2014/0039032; V of US2018/0028664; I of US2016/0317458; I of US2013/0195920; 5, 6, or 10 of U.S. Pat. No. 10,221,127; 111-3 of WO2018/081480; 1-5 or 1-8 of WO2020/081938; 18 or 25 of U.S. Pat. No. 9,867,888; A of US2019/0136231; II of WO2020/219876; 1 of US2012/0027803; OF-02 of US2019/0240349; 23 of U.S. Pat. No. 10,086,013; cKK-E12/A6 of Miao et al (2020); C12-200 of WO2010/053572; 7C1 of Dahlman et al (2017); 304-013 or 503-013 of Whitehead et al; TS-P4C2 of U.S. Pat. No. 9,708,628; I of WO2020/106946; I of WO2020/106946.


In some embodiments, the ionizable lipid is MC3 (6Z,9Z,28Z,3 1Z)-heptatriaconta-6,9,28,3 1-tetraen-19-yl-4-(dimethylamino) butanoate (DLin-MC3-DMA or MC3), e.g., as described in Example 9 of WO2019051289A9 (incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is the lipid ATX-002, e.g., as described in Example 10 of WO2019051289A9 (incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is (13Z,16Z)-A,A-dimethyl-3-nonyldocosa-13,16-dien-1-amine (Compound 32), e.g., as described in Example 11 of WO2019051289A9 (incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is Compound 6 or Compound 22, e.g., as described in Example 12 of WO2019051289A9 (incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate (SM-102); e.g., as described in Example 1 of U.S. Pat. No. 9,867,888(incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is 9Z,12Z)-3-((4,4-bis(octyloxy)butanoyl)oxy)-2-((((3-(diethylamino)propoxy)carbonyl)oxy)methyl)propyl octadeca-9,12-dienoate (LP01) e.g., as synthesized in Example 13 of WO2015/095340(incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is Di((Z)-non-2-en-1-yl) 9-((4-dimethylamino)butanoyl)oxy)heptadecanedioate (L319), e.g. as synthesized in Example 7, 8, or 9 of US2012/0027803(incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is 1,1′-((2-(4-(2-((2-(Bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl) amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-01) (C12-200), e.g., as synthesized in Examples 14 and 16 of WO2010/053572(incorporated by reference herein in its entirety). In some embodiments, the ionizable lipid is; Imidazole cholesterol ester (ICE) lipid (3S, 10R, 13R, 17R)-10, 13-dimethyl-17-((R)-6-methylheptan-2-yl)-2, 3, 4, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl3-(1H-imidazol-4-yl)propanoate, e.g., Structure (I) from WO2020/106946 (incorporated by reference herein in its entirety).


Some non-limiting examples of lipid compounds that may be used (e.g., in combination with other lipid components) to form lipid nanoparticles for the delivery of compositions described herein, e.g., nucleic acid (e.g., RNA) described herein (e.g., a template nucleic acid or a nucleic acid encoding a gene modifying polypeptide) includes,




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In some embodiments an LNP comprising Formula (i) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (ii) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (iii) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (v) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (vi) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (viii) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (ix) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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wherein


X1 is O, NR1, or a direct bond, X2 is C2-5 alkylene, X3 is C(=0) or a direct bond, R1 is H or Me, R3 is Ci-3 alkyl, R2 is Ci-3 alkyl, or R2 taken together with the nitrogen atom to which it is attached and 1-3 carbon atoms of X2 form a 4-, 5-, or 6-membered ring, or X1 is NR1, R1 and R2 taken together with the nitrogen atoms to which they are attached form a 5-or 6-membered ring, or R2 taken together with R2 and the nitrogen atom to which they are attached form a 5-, 6-, or 7-membered ring, Y1 is C2-12 alkylene. Y2 is selected from




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n is 0 to 3, R4 is Ci-15 alkyl, Z1 is Ci-6 alkylene or a direct bond,


Z2 is



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(in either orientation.) or absent, provided that if Z1 is a direct bond, Z2 is absent.


R5 is C5-9 alkyl or C6-10 alkoxy, R6 is C5-9 alkyl or C6-10 alkoxy, W is methylene or a direct bond, and R7 is H or Me, or a salt thereof provided that if R3 and R2 are C2 alkyls, X1 is O, X2 is linear C3 alkylene, X2 is C(═O), Y1 is linear Ce alkylene, (Y2)n-R4 is




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R4 is linear C5 alkyl, Z1 is C2 alkylene, Z2 is absent, W is methylene, and R7 is H, then R5 and R2 are not Cx alkoxy.


In some embodiments an LNP comprising Formula (xii) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising Formula (xi) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprises a compound of Formula (xiii) and a compound of Formula (xiv).




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In some embodiments an LNP comprising Formula (xv) is used to deliver a gene modifying composition described herein to the liver and/or hepatocyte cells.




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In some embodiments an LNP comprising a formulation of Formula (xvi) is used to deliver a gene modifying composition described herein to the lung endothelial cells.




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In some embodiments, a lipid compound used to form lipid nanoparticles for the delivery of compositions described herein, e.g., nucleic acid (e.g., RNA) described herein (e.g., a template nucleic acid or a nucleic acid encoding a gene modifying polypeptide) is made by one of the following reactions:




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Exemplary non-cationic lipids include, but are not limited to, distearoyl-sn-glycero-phosphoethanolamine, di stearoylphosphatidylcholine (DSPC), dioleoylphosphatidylcholine (DOPC), dipalmitoylphosphatidylcholine (DPPC), dioleoylphosphatidylglycerol (DOPG), dipalmitoylphosphatidylglycerol (DPPG), dioleoyl-phosphatidylethanolamine (DOPE), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), palmitoyloleoylphosphatidylcholine (POPC), palmitoyloleoylphosphatidylethanolamine (POPE), dioleoyl-phosphatidylethanolamine 4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (DOPE-mal), dipalmitoyl phosphatidyl ethanolamine (DPPE), dimyristoylphosphoethanolamine (DMPE), distearoyl-phosphatidyl-ethanolamine (DSPE), monomethyl-phosphatidylethanolamine (such as 16-O-monomethyl PE), dimethyl-phosphatidylethanolamine (such as 16-O-dimethyl PE), 18-1-trans PE, 1-stearoyl-2-oleoyl-phosphatidyethanolamine (SOPE), hydrogenated soy phosphatidylcholine (HSPC), egg phosphatidylcholine (EPC), dioleoylphosphatidylserine (DOPS), sphingomyelin (SM), dimyristoyl phosphatidylcholine (DMPC), dimyristoyl phosphatidylglycerol (DMPG), di stearoylphosphatidylglycerol (DSPG), dierucoylphosphatidylcholine (DEPC), palmitoyloleyolphosphatidylglycerol (POPG), dielaidoyl-phosphatidylethanolamine (DEPE), lecithin, phosphatidylethanolamine, lysolecithin, lysophosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, egg sphingomyelin (ESM), cephalin, cardiolipin, phosphatidicacid,cerebrosides, dicetylphosphate, lysophosphatidylcholine, dilinoleoylphosphatidylcholine, or mixtures thereof. It is understood that other diacylphosphatidylcholine and diacylphosphatidylethanolamine phospholipids can also be used. The acyl groups in these lipids are preferably acyl groups derived from fatty acids having C10-C24 carbon chains, e.g., lauroyl, myristoyl, paimitoyl, stearoyl, or oleoyl. Additional exemplary lipids, in certain embodiments, include, without limitation, those described in Kim et al. (2020) dx.doi.org/10.1021/acs.nanolett.0c01386, incorporated herein by reference. Such lipids include, in some embodiments, plant lipids found to improve liver transfection with mRNA (e.g., DGTS). In some embodiments, the non-cationic lipid may have the following structure,




text missing or illegible when filed


Other examples of non-cationic lipids suitable for use in the lipid nanopartieles include, without limitation, nonphosphorous lipids such as, e.g., stearylamine, dodeeylamine, hexadecylamine, acetyl palmitate, glycerol ricinoleate, hexadecyl stereate, isopropyl myristate, amphoteric acrylic polymers, triethanolamine-lauryl sulfate, alkyl-aryl sulfate polyethyloxylated fatty acid amides, dioctadecyl dimethyl ammonium bromide, ceramide, sphingomyelin, and the like. Other non-cationic lipids are described in WO2017/099823 or US patent publication US2018/0028664, the contents of which is incorporated herein by reference in their entirety.


In some embodiments, the non-cationic lipid is oleic acid or a compound of Formula I, II, or IV of US2018/0028664, incorporated herein by reference in its entirety. The non-cationic lipid can comprise, for example, 0-30% (mol) of the total lipid present in the lipid nanoparticle. In some embodiments, the non-cationic lipid content is 5-20% (mol) or 10-15% (mol) of the total lipid present in the lipid nanoparticle. In embodiments, the molar ratio of ionizable lipid to the neutral lipid ranges from about 2:1 to about 8:1 (e.g., about 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, or 8:1).


In some embodiments, the lipid nanoparticles do not comprise any phospholipids.


In some aspects, the lipid nanoparticle can further comprise a component, such as a sterol, to provide membrane integrity. One exemplary sterol that can be used in the lipid nanoparticle is cholesterol and derivatives thereof. Non-limiting examples of cholesterol derivatives include polar analogues such as 5α-choiestanol, 53-coprostanol, choiesteryl-(2-hydroxy)-ethyl ether, choiesteryl-(4′-hydroxy)-butyl ether, and 6-ketocholestanol; non-polar analogues such as 5α-cholestane, cholestenone, 5α-cholestanone, 5p-cholestanone, and cholesteryl decanoate; and mixtures thereof. In some embodiments, the cholesterol derivative is a polar analogue, e.g., choiesteryl-(4′-hydroxy)-butyl ether. Exemplary cholesterol derivatives are described in PCT publication WO2009/127060 and US patent publication US2010/0130588, each of which is incorporated herein by reference in its entirety.


In some embodiments, the component providing membrane integrity, such as a sterol, can comprise 0-50% (mol) (e.g., 0-10%, 10-20%, 20-30%, 30-40%, or 40-50%) of the total lipid present in the lipid nanoparticle. In some embodiments, such a component is 20-50% (mol) 30-40% (mol) of the total lipid content of the lipid nanoparticle.


In some embodiments, the lipid nanoparticle can comprise a polyethylene glycol (PEG) or a conjugated lipid molecule. Generally, these are used to inhibit aggregation of lipid nanoparticles and/or provide steric stabilization. Exemplary conjugated lipids include, but are not limited to, PEG-lipid conjugates, polyoxazoline (POZ)-lipid conjugates, polyamide-lipid conjugates (such as ATTA-lipid conjugates), cationic-polymer lipid (CPL) conjugates, and mixtures thereof. In some embodiments, the conjugated lipid molecule is a PEG-lipid conjugate, for example, a (methoxy polyethylene glycol)-conjugated lipid.


Exemplary PEG-lipid conjugates include, but are not limited to, PEG-diacylglycerol (DAG) (such as 1-(monomethoxy-polyethyleneglycol)-2,3-dimyristoylglycerol (PEG-DMG)), PEG-dialkyloxypropyl (DAA), PEG-phospholipid, PEG-ceramide (Cer), a pegylated phosphatidylethanoloamine (PEG-PE), 1,2-dimyristoyl-sn-glycerol, methoxypoly ethylene glycol (DMG-PEG-2K), PEG succinate diacylglycerol (PEGS-DAG) (such as 4-O-(2′,3′-di(tetradecanoyloxy)propyl-1-O-(w-methoxy(polyethoxy)ethyl) butanedioate (PEG-S-DMG)), PEG dialkoxypropylcarbam, N-(carbonyl-methoxypolyethylene glycol 2000)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine sodium salt, or a mixture thereof. Additional exemplary PEG-lipid conjugates are described, for example, in U.S. Pat. Nos. 5,885,613, 6,287,591, US2003/0077829, US2003/0077829, US2005/0175682, US2008/0020058, US2011/0117125, US2010/0130588, US2016/0376224, US2017/0119904, and US/099823, the contents of all of which are incorporated herein by reference in their entirety. In some embodiments, a PEG-lipid is a compound of Formula III, III-a-2, III-b-1, III-b-2, or V of US2018/0028664, the content of which is incorporated herein by reference in its entirety. In some embodiments, a PEG-lipid is of Formula II of US20150376115 or US2016/0376224, the content of both of which is incorporated herein by reference in its entirety. In some embodiments, the PEG-DAA conjugate can be, for example, PEG-dilauryloxypropyl, PEG-dimyristyloxypropyl, PEG-dipalmityloxypropyl, or PEG-distearyloxypropyl. The PEG-lipid can be one or more of PEG-DMG, PEG-dilaurylglycerol, PEG-dipalmitoylglycerol, PEG-di sterylglycerol, PEG-dilaurylglycamide, PEG-dimyristylglycamide, PEG-dipalmitoylglycamide, PEG-di sterylglycamide, PEG-cholesterol (1[8′-(Cholest-5-en-3[beta]-oxy)carboxamido-3′,6′-dioxaoctanyl] carbamoyl-[omega]-methyl-poly(ethylene glycol), PEG-DMB (3,4-Ditetradecoxylbenzyl-[omega]-methyl-poly(ethylene glycol) ether), and 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000]. In some embodiments, the PEG-lipid comprises PEG-DMG, 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000]. In some embodiments, the PEG-lipid comprises a structure selected from:




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In some embodiments, lipids conjugated with a molecule other than a PEG can also be used in place of PEG-lipid. For example, polyoxazoline (POZ)-lipid conjugates, polyamide-lipid conjugates (such as ATTA-lipid conjugates), and cationic-polymer lipid (GPL) conjugates can be used in place of or in addition to the PEG-lipid.


Exemplary conjugated lipids, i.e., PEG-lipids, (POZ)-lipid conjugates, ATTA-lipid conjugates and cationic polymer-lipids are described in the PCT and LIS patent applications listed in Table 2 of WO2019051289A9 and in WO2020106946A1, the contents of all of which are incorporated herein by reference in their entirety.


In some embodiments an LNP comprises a compound of Formula (xix), a compound of Formula (xxi) and a compound of Formula (xxv). In some embodiments an LNP comprising a formulation of Formula (xix), Formula (xxi) and Formula (xxv)is used to deliver a gene modifying composition described herein to the lung or pulmonary cells.


In some embodiments, a lipid nanoparticle may comprise one or more cationic lipids selected from Formula (i), Formula (ii), Formula (iii), Formula (vii), and Formula (ix). In some embodiments, the LNP may further comprise one or more neutral lipid, e.g., DSPC, DPPC, D1ViPC, DOPC, POPC, DOPE, SM, a steroid, e.g., cholesterol, and/or one or more polymer conjugated lipid, e.g., a pegylated lipid, e.g., PEG-DAG, PEG-PE, PEG-S-DAG, PEG-cer or a PEG dialkyoxypropylcarbamate.


In some embodiments, the PEG or the conjugated lipid can comprise 0-20% (mol) of the total lipid present in the lipid nanoparticle. In some embodiments, PEG or the conjugated lipid content is 0.5-10% or 2-5% (mol) of the total lipid present in the lipid nanoparticle. Molar ratios of the ionizable lipid, non-cationic-lipid, sterol, and PEG/conjugated lipid can be varied as needed. For example, the lipid particle can comprise 30-70% ionizable lipid by mole or by total weight of the composition, 0-60% cholesterol by mole or by total weight of the composition, 0-30% non-cationic-lipid by mole or by total weight of the composition and 1-10% conjugated lipid by mole or by total weight of the composition. Preferably, the composition comprises 30-40% ionizable lipid by mole or by total weight of the composition, 40-50% cholesterol by mole or by total weight of the composition, and 10-20% non-cationic-lipid by mole or by total weight of the composition. In some other embodiments, the composition is 50-75% ionizable lipid by mole or by total weight of the composition, 20-40% cholesterol by mole or by total weight of the composition, and 5 to 10% non-cationic-lipid, by mole or by total weight of the composition and 1-10% conjugated lipid by mole or by total weight of the composition. The composition may contain 60-70% ionizable lipid by mole or by total weight of the composition, 25-35% cholesterol by mole or by total weight of the composition, and 5-10% non-cationic-lipid by mole or by total weight of the composition. The composition may also contain up to 90% ionizable lipid by mole or by total weight of the composition and 2 to 15% non-cationic lipid by mole or by total weight of the composition. The formulation may also be a lipid nanoparticle formulation, for example comprising 8-30% ionizable lipid by mole or by total weight of the composition, 5-30% non-cationic lipid by mole or by total weight of the composition, and 0-20% cholesterol by mole or by total weight of the composition; 4-25% ionizable lipid by mole or by total weight of the composition, 4-25% non-cationic lipid by mole or by total weight of the composition, 2 to 25% cholesterol by mole or by total weight of the composition, 10 to 35% conjugate lipid by mole or by total weight of the composition, and 5% cholesterol by mole or by total weight of the composition; or 2-30% ionizable lipid by mole or by total weight of the composition, 2-30% non-cationic lipid by mole or by total weight of the composition, 1 to 15% cholesterol by mole or by total weight of the composition, 2 to 35% conjugate lipid by mole or by total weight of the composition, and 1-20% cholesterol by mole or by total weight of the composition; or even up to 90% ionizable lipid by mole or by total weight of the composition and 2-10% non-cationic lipids by mole or by total weight of the composition, or even 100% cationic lipid by mole or by total weight of the composition. In some embodiments, the lipid particle formulation comprises ionizable lipid, phospholipid, cholesterol and a PEG-ylated lipid in a molar ratio of 50:10:38.5:1.5. In some other embodiments, the lipid particle formulation comprises ionizable lipid, cholesterol and a PEG-ylated lipid in a molar ratio of 60:38.5:1.5.


In some embodiments, the lipid particle comprises ionizable lipid, non-cationic lipid (e.g. phospholipid), a sterol (e.g., cholesterol) and a PEG-ylated lipid, where the molar ratio of lipids ranges from 20 to 70 mole percent for the ionizable lipid, with a target of 40-60, the mole percent of non-cationic lipid ranges from 0 to 30, with a target of 0 to 15, the mole percent of sterol ranges from 20 to 70, with a target of 30 to 50, and the mole percent of PEG-ylated lipid ranges from 1 to 6, with a target of 2 to 5.


In some embodiments, the lipid particle comprises ionizable lipid/non-cationic-lipid/sterol/conjugated lipid at a molar ratio of 50:10:38.5:1.5.


In an aspect, the disclosure provides a lipid nanoparticle formulation comprising phospholipids, lecithin, phosphatidylcholine and phosphatidylethanolamine.


In some embodiments, one or more additional compounds can also be included. Those compounds can be administered separately or the additional compounds can be included in the lipid nanoparticles of the invention. In other words, the lipid nanoparticles can contain other compounds in addition to the nucleic acid or at least a second nucleic acid, different than the first. Without limitations, other additional compounds can be selected from the group consisting of small or large organic or inorganic molecules, monosaccharides, disaccharides, trisaccharides, oligosaccharides, polysaccharides, peptides, proteins, peptide analogs and derivatives thereof, peptidomimetics, nucleic acids, nucleic acid analogs and derivatives, an extract made from biological materials, or any combinations thereof.


In some embodiments, a lipid nanoparticle (or a formulation comprising lipid nanoparticles) lacks reactive impurities (e.g., aldehydes or ketones), or comprises less than a preselected level of reactive impurities (e.g., aldehydes or ketones). While not wishing to be bound by theory, in some embodiments, a lipid reagent is used to make a lipid nanoparticle formulation, and the lipid reagent may comprise a contaminating reactive impurity (e.g., an aldehyde or ketone). A lipid regent may be selected for manufacturing based on having less than a preselected level of reactive impurities (e.g., aldehydes or ketones). Without wishing to be bound by theory, in some embodiments, aldehydes can cause modification and damage of RNA, e.g., cross-linking between bases and/or covalently conjugating lipid to RNA (e.g., forming lipid-RNA adducts). This may, in some instances, lead to failure of a reverse transcriptase reaction and/or incorporation of inappropriate bases, e.g., at the site(s) of lesion(s), e.g., a mutation in a newly synthesized target DNA.


In some embodiments, a lipid nanoparticle formulation is produced using a lipid reagent comprising less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% total reactive impurity (e.g., aldehyde) content. In some embodiments, a lipid nanoparticle formulation is produced using a lipid reagent comprising less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% of any single reactive impurity (e.g., aldehyde) species. In some embodiments, a lipid nanoparticle formulation is produced using a lipid reagent comprising: (i) less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% total reactive impurity (e.g., aldehyde) content; and (ii) less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% of any single reactive impurity (e.g., aldehyde) species. In some embodiments, the lipid nanoparticle formulation is produced using a plurality of lipid reagents, and each lipid reagent of the plurality independently meets one or more criterion described in this paragraph. In some embodiments, each lipid reagent of the plurality meets the same criterion, e.g., a criterion of this paragraph.


In some embodiments, the lipid nanoparticle formulation comprises less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% total reactive impurity (e.g., aldehyde) content. In some embodiments, the lipid nanoparticle formulation comprises less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% of any single reactive impurity (e.g., aldehyde) species. In some embodiments, the lipid nanoparticle formulation comprises: (i) less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% total reactive impurity (e.g., aldehyde) content; and (ii) less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% of any single reactive impurity (e.g., aldehyde) species.


In some embodiments, one or more, or optionally all, of the lipid reagents used for a lipid nanoparticle as described herein or a formulation thereof comprise less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% total reactive impurity (e.g., aldehyde) content. In some embodiments, one or more, or optionally all, of the lipid reagents used for a lipid nanoparticle as described herein or a formulation thereof comprise less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% of any single reactive impurity (e.g., aldehyde) species. In some embodiments, one or more, or optionally all, of the lipid reagents used for a lipid nanoparticle as described herein or a formulation thereof comprise: (i) less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% total reactive impurity (e.g., aldehyde) content; and (ii) less than 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1% of any single reactive impurity (e.g., aldehyde) species.


In some embodiments, total aldehyde content and/or quantity of any single reactive impurity (e.g., aldehyde) species is determined by liquid chromatography (LC), e.g., coupled with tandem mass spectrometry (MS/MS), e.g., according to the method described in Example 40 of PCT/US21/20948. In some embodiments, reactive impurity (e.g., aldehyde) content and/or quantity of reactive impurity (e.g., aldehyde) species is determined by detecting one or more chemical modifications of a nucleic acid molecule (e.g., an RNA molecule, e.g., as described herein) associated with the presence of reactive impurities (e.g., aldehydes), e.g., in the lipid reagents. In some embodiments, reactive impurity (e.g., aldehyde) content and/or quantity of reactive impurity (e.g., aldehyde) species is determined by detecting one or more chemical modifications of a nucleotide or nucleoside (e.g., a ribonucleotide or ribonucleoside, e.g., comprised in or isolated from a template nucleic acid, e.g., as described herein) associated with the presence of reactive impurities (e.g., aldehydes), e.g., in the lipid reagents, e.g., according to the method described in Example 41 of PCT/US21/20948. In embodiments, chemical modifications of a nucleic acid molecule, nucleotide, or nucleoside are detected by determining the presence of one or more modified nucleotides or nucleosides, e.g., using LC-MS/MS analysis, e.g., according to the method described in Example 41 of PCT/US21/20948.


In some embodiments, a nucleic acid (e.g., RNA) described herein (e.g., a template nucleic acid or a nucleic acid encoding a gene modifying polypeptide) does not comprise an aldehyde modification, or comprises less than a preselected amount of aldehyde modifications. In some embodiments, on average, a nucleic acid has less than 50, 20, 10, 5, 2, or 1 aldehyde modifications per 1000 nucleotides, e.g., wherein a single cross-linking of two nucleotides is a single aldehyde modification. In some embodiments, the aldehyde modification is an RNA adduct (e.g., a lipid-RNA adduct). In some embodiments, the aldehyde-modified nucleotide is cross-linking between bases. In some embodiments, a nucleic acid (e.g., RNA) described herein comprises less than 50, 20, 10, 5, 2, or 1 cross-links between nucleotide.


In some embodiments, LNPs are directed to specific tissues by the addition of targeting domains. For example, biological ligands may be displayed on the surface of LNPs to enhance interaction with cells displaying cognate receptors, thus driving association with and cargo delivery to tissues wherein cells express the receptor. In some embodiments, the biological ligand may be a ligand that drives delivery to the liver, e.g., LNPs that display GalNAc result in delivery of nucleic acid cargo to hepatocytes that display asialoglycoprotein receptor (ASGPR). The work of Akinc et al. Mol Ther 18(7):1357-1364 (2010) teaches the conjugation of a trivalent GalNAc ligand to a PEG-lipid (GalNAc-PEG-DSG) to yield LNPs dependent on ASGPR for observable LNP cargo effect (see, e.g., FIG. 6 therein). Other ligand-displaying LNP formulations, e.g., incorporating folate, transferrin, or antibodies, are discussed in WO2017223135, which is incorporated herein by reference in its entirety, in addition to the references used therein, namely Kolhatkar et al., Curr Drug Discov Technol. 2011 8:197-206; Musacchio and Torchilin, Front Biosci. 2011 16:1388-1412; Yu et al., Mol Membr Biol. 2010 27:286-298; Patil et al., Crit Rev Ther Drug Carrier Syst. 2008 25:1-61; Benoit et al., Biomacromolecules. 2011 12:2708-2714; Zhao et al., Expert Opin Drug Deliv. 2008 5:309-319; Akinc et al., Mol Ther. 2010 18:1357-1364; Srinivasan et al., Methods Mol Biol. 2012 820:105-116; Ben-Arie et al., Methods Mol Biol. 2012 757:497-507; Peer 2010 J Control Release. 20:63-68; Peer et al., Proc Natl Acad Sci USA. 2007 104:4095-4100; Kim et al., Methods Mol Biol. 2011 721:339-353; Subramanya et al., Mol Ther. 2010 18:2028-2037; Song et al., Nat Biotechnol. 2005 23:709-717; Peer et al., Science. 2008 319:627-630; and Peer and Lieberman, Gene Ther. 2011 18:1127-1133.


In some embodiments, LNPs are selected for tissue-specific activity by the addition of a Selective ORgan Targeting (SORT) molecule to a formulation comprising traditional components, such as ionizable cationic lipids, amphipathic phospholipids, cholesterol and poly(ethylene glycol) (PEG) lipids. The teachings of Cheng et al. Nat Nanotechnol 15(4):313-320 (2020) demonstrate that the addition of a supplemental “SORT” component precisely alters the in vivo RNA delivery profile and mediates tissue-specific (e.g., lungs, liver, spleen) gene delivery and editing as a function of the percentage and biophysical property of the SORT molecule.


In some embodiments, the LNPs comprise biodegradable, ionizable lipids. In some embodiments, the LNPs comprise (9Z,12Z)-3-((4,4-bis(octyloxy)butanoyl)oxy)-2-((((3-(diethylamino)propoxy)carbonyl)oxy)methyl)propyl octadeca-9,12-dienoate, also called 3-((4,4-bis(octyloxy)butanoyl)oxy)-2-((((3-(diethylamino)propoxy)carbonyl)oxy)methyl)propyl (9Z,12Z)-octadeca-9,12-dienoate) or another ionizable lipid. See, e.g, lipids of WO2019/067992, WO/2017/173054, WO2015/095340, and WO2014/136086, as well as references provided therein. In some embodiments, the term cationic and ionizable in the context of LNP lipids is interchangeable, e.g., wherein ionizable lipids are cationic depending on the pH.


In some embodiments, an LNP described herein comprises a lipid described in Table 19.









TABLE 19







Exemplary Lipids












Molecular



LIPID ID
Chemical Name
Weight
Structure





LIPIDV003
(9Z,12Z)-3-((4,4- bis(octyloxy)butanoyl) oxy)-2-((((3- (diethylamino)propoxy) carbonyl)oxy)methyl) propyl octadeca-9,12- dienoate
852.29


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LIPIDV004
Heptadecan-9-y1 8-((2- hydroxyethyl)(8- (nonyloxy)-8- oxooctyl)amino)octanoate
710.18


embedded image







LIPIDV005

919.56


embedded image











In some embodiments, multiple components of a gene modifying system may be prepared as a single LNP formulation, e.g., an LNP formulation comprises mRNA encoding for the gene modifying polypeptide and an RNA template. Ratios of nucleic acid components may be varied in order to maximize the properties of a therapeutic. In some embodiments, the ratio of RNA template to mRNA encoding a gene modifying polypeptide is about 1:1 to 100:1, e.g., about 1:1 to 20:1, about 20:1 to 40:1, about 40:1 to 60:1, about 60:1 to 80:1, or about 80:1 to 100:1, by molar ratio. In other embodiments, a system of multiple nucleic acids may be prepared by separate formulations, e.g., one LNP formulation comprising a template RNA and a second


LNP formulation comprising an mRNA encoding a gene modifying polypeptide. In some embodiments, the system may comprise more than two nucleic acid components formulated into LNPs. In some embodiments, the system may comprise a protein, e.g., a gene modifying polypeptide, and a template RNA formulated into at least one LNP formulation.


In some embodiments, the average LNP diameter of the LNP formulation may be between 10s of nm and 100s of nm, e.g., measured by dynamic light scattering (DLS). In some embodiments, the average LNP diameter of the LNP formulation may be from about 40 nm to about 150 nm, such as about 40 nm, 45 nm, 50 nm, 55 nm, 60 nm, 65 nm, 70 nm, 75 nm, 80 nm, 85 nm, 90 nm, 95 nm, 100 nm, 105 nm, 110 nm, 115 nm, 120 nm, 125 nm, 130 nm, 135 nm, 140 nm, 145 nm, or 150 nm. In some embodiments, the average LNP diameter of the LNP formulation may be from about 50 nm to about 100 nm, from about 50 nm to about 90 nm, from about 50 nm to about 80 nm, from about 50 nm to about 70 nm, from about 50 nm to about 60 nm, from about 60 nm to about 100 nm, from about 60 nm to about 90 nm, from about 60 nm to about 80 nm, from about 60 nm to about 70 nm, from about 70 nm to about 100 nm, from about 70 nm to about 90 nm, from about 70 nm to about 80 nm, from about 80 nm to about 100 nm, from about 80 nm to about 90 nm, or from about 90 nm to about 100 nm. In some embodiments, the average LNP diameter of the LNP formulation may be from about 70 nm to about 100 nm. In a particular embodiment, the average LNP diameter of the LNP formulation may be about 80 nm. In some embodiments, the average LNP diameter of the LNP formulation may be about 100 nm. In some embodiments, the average LNP diameter of the LNP formulation ranges from about 1 mm to about 500 mm, from about 5 mm to about 200 mm, from about 10 mm to about 100 mm, from about 20 mm to about 80 mm, from about 25 mm to about 60 mm, from about 30 mm to about 55 mm, from about 35 mm to about 50 mm, or from about 38 mm to about 42 mm.


An LNP may, in some instances, be relatively homogenous. A polydispersity index may be used to indicate the homogeneity of an LNP, e.g., the particle size distribution of the lipid nanoparticles. A small (e.g., less than 0.3) polydispersity index generally indicates a narrow particle size distribution. An LNP may have a polydispersity index from about 0 to about 0.25, such as 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20, 0.21, 0.22, 0.23, 0.24, or 0.25. In some embodiments, the polydispersity index of an LNP may be from about 0.10 to about 0.20.


The zeta potential of an LNP may be used to indicate the electrokinetic potential of the composition. In some embodiments, the zeta potential may describe the surface charge of an LNP. Lipid nanoparticles with relatively low charges, positive or negative, are generally desirable, as more highly charged species may interact undesirably with cells, tissues, and other elements in the body. In some embodiments, the zeta potential of an LNP may be from about −10 mV to about +20 mV, from about −10 mV to about +15 mV, from about −10 mV to about +10 mV, from about −10 mV to about +5 mV, from about −10 mV to about 0 mV, from about −10 mV to about −5 mV, from about −5 mV to about +20 mV, from about −5 mV to about +15 mV, from about −5 mV to about +10 mV, from about −5 mV to about +5 mV, from about −5 mV to about 0 mV, from about 0 mV to about +20 mV, from about 0 mV to about +15 mV, from about 0 mV to about +10 mV, from about 0 mV to about +5 mV, from about +5 mV to about +20 mV, from about +5 mV to about +15 mV, or from about +5 mV to about +10 mV.


The efficiency of encapsulation of a protein and/or nucleic acid, e.g., gene modifying polypeptide or mRNA encoding the polypeptide, describes the amount of protein and/or nucleic acid that is encapsulated or otherwise associated with an LNP after preparation, relative to the initial amount provided. The encapsulation efficiency is desirably high (e.g., close to 100%). The encapsulation efficiency may be measured, for example, by comparing the amount of protein or nucleic acid in a solution containing the lipid nanoparticle before and after breaking up the lipid nanoparticle with one or more organic solvents or detergents. An anion exchange resin may be used to measure the amount of free protein or nucleic acid (e.g., RNA) in a solution. Fluorescence may be used to measure the amount of free protein and/or nucleic acid (e.g., RNA) in a solution. For the lipid nanoparticles described herein, the encapsulation efficiency of a protein and/or nucleic acid may be at least 50%, for example 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%. In some embodiments, the encapsulation efficiency may be at least 80%. In some embodiments, the encapsulation efficiency may be at least 90%. In some embodiments, the encapsulation efficiency may be at least 95%.


An LNP may optionally comprise one or more coatings. In some embodiments, an LNP may be formulated in a capsule, film, or table having a coating. A capsule, film, or tablet including a composition described herein may have any useful size, tensile strength, hardness or density.


Additional exemplary lipids, formulations, methods, and characterization of LNPs are taught by WO2020061457, which is incorporated herein by reference in its entirety.


In some embodiments, in vitro or ex vivo cell lipofections are performed using Lipofectamine MessengerMax (Thermo Fisher) or TranslT-mRNA Transfection Reagent (Minis Bio). In certain embodiments, LNPs are formulated using the GenVoy ILM ionizable lipid mix (Precision NanoSystems). In certain embodiments, LNPs are formulated using 2,2-dilinoleyl-4-dimethylaminoethyl-[1,3]-dioxolane (DLin-KC2-DMA) or dilinoleylmethyl-4-dimethylaminobutyrate (DLin-MC3-DMA or MC3), the formulation and in vivo use of which are taught in Jayaraman et al. Angew Chem Int Ed Engl 51(34):8529-8533 (2012), incorporated herein by reference in its entirety.


LNP formulations optimized for the delivery of CRISPR-Cas systems, e.g., Cas9-gRNA RNP, gRNA, Cas9 mRNA, are described in WO2019067992 and WO2019067910, both incorporated by reference.


Additional specific LNP formulations useful for delivery of nucleic acids are described in U.S. Pat. Nos. 8,158,601 and 8,168,775, both incorporated by reference, which include formulations used in patisiran, sold under the name ONPATTRO.


Exemplary dosing of gene modifying LNP may include about 0.1, 0.25, 0.3, 0.5, 1, 2, 3, 4, 5, 6, 8, 10, or 100 mg/kg (RNA). Exemplary dosing of AAV comprising a nucleic acid encoding one or more components of the system may include an MOI of about 1011, 1012, 1013, and 1014 vg/kg.


An mRNA encoding a gene modifying polypeptide may have a cap, 5′ UTR containing a Kozak, 3′ UTR, and polyA tail containing at least 60 As (SEQ ID NO: 25695). An mRNA encoding a gene modifying polypeptide may have a reduced Uridine content through codon selection/optimization. An mRNA encoding a gene modifying polypeptide may have uridines that are about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substituted with 5-methoxy uridine. An mRNA encoding a gene modifying polypeptide may have uridines that are about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substituted with N1-methyl-pseudouridine. An mRNA encoding a gene modifying polypeptide may have cytosines in the mRNA are about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substituted with 5-methylcytosine. An mRNA encoding a gene modifying polypeptide may have a combination of about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substitution of cytosine with 5-methylcytosine and about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substitution of uridine with 5-methoxy uridine. An mRNA encoding a gene modifying polypeptide may have a combination of about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substitution of cytosine with 5-methylcytosine and about 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% substitution of uridine with N1-methyl-pseudouridine.


A guide RNA may be synthesized by T7 RNA polymerase. A guide RNA may be chemically synthesized and contain modifications such as, e.g., 2′-O-methyl, 2′-Fluoro, and/or phosphorothioate. The 3 most terminal nucleotides of a guide RNA may contain 2′-O-methyl modifications with 3 phosphorothioate linkages between the nucleotides. A guide RNA may contain 2′-O-methyl modified nucleotides where there are cytosines and uridines, except at nucleotides found in the “seed” of the guide RNA where cytosines and uridines contain 2′-fluoro modifications.


A gene modifying mRNA and a guide RNA may be co-formulated in an LNP as described herein. They may be separately formulated. They may be combined prior to injection. They may be combined at a molar ratio in the range of about 1:10 to 1:250 mRNA:gRNA. They may be formulated in a molar ratio of about 1:20, 1:30, 1:40, 1:50, 1:60, 1:70, 1:80, 1:90, 1:100, 1:110, 1:120, 1:130, 1:140, 1:150, 1:160, 1:170, 1:180, 1:190, 1:200, 1:210, 1:220, 1:230, 1:240, or 1:250 mRNA:gRNA. The mRNA and guide RNA may be injected 30-180 minutes apart where the mRNA LNPs are delivered first followed by the guide RNA LNPs. The may be delivered about 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, or 180 minutes apart. The mRNA and/or gRNA may be dosed at 0.01-6 mg/kg either separately or together as a total amount of RNA-LNP. The RNA-LNPs may be injected IV bolus. The RNA-LNPs may be infused over a period of 30-360 minutes. The RNA-LNPs may be infused over a period of about 30, 60, 90, 120, 150, 180, 210, 240, 270, 300, 330 or 360 minutes.


Exemplary Gene Modifying Systems for Correcting an E342K Mutation

In some embodiments, the compositions of the gene modifying system used to correct the E342K mutation in the PiZ model, as described herein, are modified as follows to optimize efficiency and precision of editing.


Gene modifying polypeptide-encoding mRNA. In some embodiments, the gene modifying polypeptide comprises the bipartite SV40 NLS sequences (doi: 10.1074/jbc.M601718200) at its N-terminus and C-terminus. In some embodiments, The gene modifying system construct contains modified c-myc NLS and bipartite SV40 NLS at its N-terminus and at the C-terminus a modified bipartite SV40 NLS followed by a SV40 NLS is linked to the reverse transcriptase through a SGGS (SEQ ID NO: 25694) linker. In some embodiments, the linker between each NLS and the NLS and the fusion protein is a SGGS (SEQ ID NO: 25694) linker. In some embodiments, the 32 amino acid linker of the fusion protein encoded by the mRNA is:











(SEQ ID NO: 19525)



SGGSSGGSSGSETPGTSESATPESSGGSSGGSS.






In some embodiments, the catalytic mutation of the Cas9 domain to generate the Cas9 nickase activity is H840A or N863A. In some embodiments, the mRNA has a cap, 5′ UTR containing a Kozak sequence, 3′ UTR, and a polyA tail containing at least 60 As (SEQ ID NO: 25695). In some embodiments, the mRNA has a reduced uridine content through codon selection/optimization. In some embodiments, the uridines in the mRNA are 100% substituted with 5-methoxy uridine. In some embodiments, the uridines in the mRNA are 100% substituted with N1-methyl-pseudouridine. In some embodiments, the cytosines in the mRNA are 100% substituted with 5-methylcytosine. In some embodiments, the mRNA contains a combination of 100% substitution of cytosine with 5-methylcytosine and 100% substitution of uridine with 5-methoxy uridine. In some embodiments, the mRNA contains a combination of 100% substitution of cytosine with 5-methylcytosine and 100% substitution of uridine with N1-methyl-pseudouridine. In some embodiments, combinations of modifications described above include 0-100% substitution of unmodified nucleotides, e.g., 0-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or less than 90-100% substitution. In some embodiments, the gene modifying polypeptide encoded by the mRNA of the system comprises the sequence:










c-Myc NLS-BPSV40 NLS-SpCas9H840A-linker-M-



MLV_reverse_transcriptase-SGGS linker-BPSV40 NLS-SV40





(SEQ ID NO:19526)





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KFKVLGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEM





AKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKA





DLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAK





AILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDT





YDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQ





DLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELL





VKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYY





VGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPK





HSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDY





FKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDR





EMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFA





NRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELV





KVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQ





NEKLYLYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRG





KSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVE





TRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYH





HAHDAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYS





NIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEV





QTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLK





SVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGE





LQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKR





VILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTST







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VEFEPKKKRKV






Template RNA and Optional Second-Nick Guide RNA.


In some embodiments, the gene modifying system employs only a Template RNA in addition to the mRNA encoding the gene modifying polypeptide. In some embodiments, the gene modifying system additionally employs a second-nick guide RNA that targets the Cas9 nickase of the system to the non-edited strand of the target DNA. In some embodiments, the gene modifying Template RNA for targeting SERPINA1 is: UCCCCUCCAGGCCGUGCAUAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGG CUAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUCCUCUCGUCGAUGGU CAGCACAGCUUUAUGCACGGCCUGGAG (SEQ ID NO: 19527). In some embodiments, the optional guide RNA for second nicking is: GGUUUGUUGAACUUGACCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGC UAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGCUUUUU (SEQ ID NO: 19528). In some embodiments, the Template RNA and optional second-nick guide RNA are synthesized by T7 RNA polymerase. In some embodiments, the Template RNA and optional second-nick guide RNA are chemically synthesized and contain a combination of one or multiple modifications of the following: 2′-O-methyl, 2′-Fluoro, and/or Phosphorothioate. In some embodiments, the 3 most terminal nucleotides contain 2′-O-methyl modifications with 3 phosphorothioate linkages between the nucleotides. In some embodiments, the Template RNA and optional second-nick guide RNA contain 2′-O-methyl modified nucleotides, where there are cytosines and uridines, except at nucleotides found in the seed sequence of the gRNA spacers, e.g., the seed sequences in the 3′ end of the spacer regions, where cytosines and uridines contain 2′-fluoro modifications and/or combination of 2′-fluoro and 2′ hydroxyl. In some embodiments, combinations of modifications described above include 0-100% substitution of unmodified nucleotides, e.g., 0-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or less than 90-100% substitution.


Formulations. In some embodiments, the gene modifying polypeptide mRNA and Template RNA (and optional second-nick guide RNA) are separately formulated as described above, combined prior to injection at a 1:20 RNA molar ratio, mRNA:Template RNA (and optionally mRNA:second-nick guide RNA), respectively. In some embodiments, the gene modifying polypeptide mRNA and Template RNA (and optional second-nick guide RNA) are separately formulated as described above, combined prior to injection at a 1:50 RNA molar ratio, mRNA:guide RNAs (and optionally mRNA:second-nick guide RNA), respectively. In some embodiments, the gene modifying polypeptide mRNA and Template RNA (and optional second-nick guide RNA) are separately formulated, combined prior to injection at ratio ranges from 1:10-1:250, mRNA:Template RNA (and optionally mRNA:second-nick guide RNA), respectively. In some embodiments, the mRNA and Template RNA (and optional second-nick guide NRA) are mixed together at a 1:10-1:250, mRNA:Template RNA (and optionally mRNA:second-nick guide RNA), and then formulated as described above, where the RNA concentration going into formulation is 0.1 mg/mL. In some embodiments, the mRNA and Template RNA (and optional second-nick guide RNA) are formulated separately and are injected 30-180 minutes apart, where the mRNA LNPs are delivered first followed by the Template RNA (and optional second-nick guide RNA) LNPs. In some embodiments, the ionizable lipid is LIPIDV005 from Table 19.


Dosing. In some embodiments, the gene modifying polypeptide mRNA and/or Template RNA (and optional second-nick guide RNA) are dosed at 0.01-6 mg/kg, either separately or together as a total amount of RNA-LNP. In some embodiments, the RNA-LNPs is injected as an IV bolus. In some embodiments, the RNA-LNPs is infused over a period of 30-360 minutes.


Kits, Articles of Manufacture, and Pharmaceutical Compositions

In an aspect the disclosure provides a kit comprising a gene modifying polypeptide or a gene modifying system, e.g., as described herein. In some embodiments, the kit comprises a gene modifying polypeptide (or a nucleic acid encoding the polypeptide) and a template RNA (or DNA encoding the template RNA). In some embodiments, the kit further comprises a reagent for introducing the system into a cell, e.g., transfection reagent, LNP, and the like. In some embodiments, the kit is suitable for any of the methods described herein. In some embodiments, the kit comprises one or more elements, compositions (e.g., pharmaceutical compositions), gene modifying polypeptides, and/or gene modifying systems, or a functional fragment or component thereof, e.g., disposed in an article of manufacture. In some embodiments, the kit comprises instructions for use thereof.


In an aspect, the disclosure provides an article of manufacture, e.g., in which a kit as described herein, or a component thereof, is disposed.


In an aspect, the disclosure provides a pharmaceutical composition comprising a gene modifying polypeptide or a gene modifying system, e.g., as described herein. In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier or excipient. In some embodiments, the pharmaceutical composition comprises a template RNA and/or an RNA encoding the polypeptide. In embodiments, the pharmaceutical composition has one or more (e.g., 1, 2, 3, or 4) of the following characteristics:

    • (a) less than 1% (e.g., less than 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%) DNA template relative to the template RNA and/or the RNA encoding the polypeptide, e.g., on a molar basis;
    • (b) less than 1% (e.g., less than 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%) uncapped RNA relative to the template RNA and/or the RNA encoding the polypeptide, e.g., on a molar basis;
    • (c) less than 1% (e.g., less than 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%) partial length RNAs relative to the template RNA and/or the RNA encoding the polypeptide, e.g., on a molar basis;
    • (d) substantially lacks unreacted cap dinucleotides.


Chemistry, Manufacturing, and Controls (CMC)

Purification of protein therapeutics is described, for example, in Franks, Protein Biotechnology: Isolation, Characterization, and Stabilization, Humana Press (2013); and in Cutler, Protein Purification Protocols (Methods in Molecular Biology), Humana Press (2010).


In some embodiments, a gene modifying system, polypeptide, and/or template nucleic acid (e.g., template RNA) conforms to certain quality standards. In some embodiments, a gene modifying system, polypeptide, and/or template nucleic acid (e.g., template RNA) produced by a method described herein conforms to certain quality standards. Accordingly, the disclosure is directed, in some aspects, to methods of manufacturing a gene modifying system, polypeptide, and/or template nucleic acid (e.g., template RNA) that conforms to certain quality standards, e.g., in which said quality standards are assayed. The disclosure is also directed, in some aspects, to methods of assaying said quality standards in a gene modifying system, polypeptide, and/or template nucleic acid (e.g., template RNA). In some embodiments, quality standards include, but are not limited to, one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12) of the following:

    • (i) the length of the template RNA, e.g., whether the template RNA has a length that is above a reference length or within a reference length range, e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the template RNA present is greater than 100, 125, 150, 175, or 200 nucleotides long;
    • (ii) the presence, absence, and/or length of a polyA tail on the template RNA, e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the template RNA present contains a polyA tail (e.g., a polyA tail that is at least 5, 10, 20, 30, 50, 70, 100 nucleotides in length (SEQ ID NO: 25697));
    • (iii) the presence, absence, and/or type of a 5′ cap on the template RNA, e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the template RNA present contains a 5′ cap, e.g., whether that cap is a 7-methylguanosine cap, e.g., a O-Me-m7G cap;
    • (iv) the presence, absence, and/or type of one or more modified nucleotides (e.g., selected from pseudouridine, dihydrouridine, inosine, 7-methylguanosine, 1-N-methylpseudouridine (1-5-methoxyuridine (5-MO-U), 5-methylcytidine (5mC), or a locked nucleotide) in the template RNA, e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the template RNA present contains one or more modified nucleotides;
    • (v) the stability of the template RNA (e.g., over time and/or under a pre-selected condition), e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the template RNA remains intact (e.g., greater than 100, 125, 150, 175, or 200 nucleotides long) after a stability test;
    • (vi) the potency of the template RNA in a system for modifying DNA, e.g., whether at least 1% of target sites are modified after a system comprising the template RNA is assayed for potency;
    • (vii) the length of the polypeptide, first polypeptide, or second polypeptide, e.g., whether the polypeptide, first polypeptide, or second polypeptide has a length that is above a reference length or within a reference length range, e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the polypeptide, first polypeptide, or second polypeptide present is greater than 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450, 1500, 1600, 1700, 1800, 1900, or 2000 amino acids long (and optionally, no larger than 2500, 2000, 1500, 1400, 1300, 1200, 1100, 1000, 900, 800, 700, or 600 amino acids long);
    • (viii) the presence, absence, and/or type of post-translational modification on the polypeptide, first polypeptide, or second polypeptide, e.g., whether at least 80, 85, 90, 95, 96, 97, 98, or 99% of the polypeptide, first polypeptide, or second polypeptide contains phosphorylation, methylation, acetylation, myristoylation, palmitoylation, isoprenylation, glipyatyon, or lipoylation, or any combination thereof;
    • (ix) the presence, absence, and/or type of one or more artificial, synthetic, or non-canonical amino acids (e.g., selected from ornithine, (3-alanine, GABA, 6-Aminolevulinic acid, PABA, a D-amino acid (e.g., D-alanine or D-glutamate), aminoisobutyric acid, dehydroalanine, cystathionine, lanthionine, Djenkolic acid, Diaminopimelic acid, Homoalanine, Norvaline, Norleucine, Homonorleucine, homoserine, O-methyl-homoserine and O-ethyl-homoserine, ethionine, selenocysteine, selenohomocysteine, selenomethionine, selenoethionine, tellurocysteine, or telluromethionine) in the polypeptide, first polypeptide, or second polypeptide, e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the polypeptide, first polypeptide, or second polypeptide present contains one or more artificial, synthetic, or non-canonical amino acids;
    • (x) the stability of the polypeptide, first polypeptide, or second polypeptide (e.g., over time and/or under a pre-selected condition), e.g., whether at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the polypeptide, first polypeptide, or second polypeptide remains intact (e.g., greater than 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450, 1500, 1600, 1700, 1800, 1900, or 2000 amino acids long (and optionally, no larger than 2500, 2000, 1500, 1400, 1300, 1200, 1100, 1000, 900, 800, 700, or 600 amino acids long)) after a stability test;
    • (xi) the potency of the polypeptide, first polypeptide, or second polypeptide in a system for modifying DNA, e.g., whether at least 1% of target sites are modified after a system comprising the polypeptide, first polypeptide, or second polypeptide is assayed for potency; or
    • (xii) the presence, absence, and/or level of one or more of a pyrogen, virus, fungus, bacterial pathogen, or host cell protein, e.g., whether the system is free or substantially free of pyrogen, virus, fungus, bacterial pathogen, or host cell protein contamination.


In some embodiments, a system or pharmaceutical composition described herein is endotoxin free.


In some embodiments, the presence, absence, and/or level of one or more of a pyrogen, virus, fungus, bacterial pathogen, and/or host cell protein is determined. In embodiments, whether the system is free or substantially free of pyrogen, virus, fungus, bacterial pathogen, and/or host cell protein contamination is determined.


In some embodiments, a pharmaceutical composition or system as described herein has one or more (e.g., 1, 2, 3, or 4) of the following characteristics:

    • (a) less than 1% (e.g., less than 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%) DNA template relative to the template RNA and/or the RNA encoding the polypeptide, e.g., on a molar basis;
    • (b) less than 1% (e.g., less than 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%) uncapped RNA relative to the template RNA and/or the RNA encoding the polypeptide, e.g., on a molar basis;
    • (c) less than 1% (e.g., less than 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%) partial length RNAs relative to the template RNA and/or the RNA encoding the polypeptide, e.g., on a molar basis;
    • (d) substantially lacks unreacted cap dinucleotides.


EXAMPLES
Example 1: Screening Configurations of Template RNAs that Correct the AATD Mutation in a Genomic Landing Pad in Human Cells

This example describes the use of gene modifying system containing a gene modifying polypeptide and template RNAs comprising varied lengths of heterologous object sequences and PBS sequences to quantify the activity of template RNAs for correction of the AATD mutation. In this example, a template RNA contains:

    • (1) a gRNA spacer;
    • (2) a gRNA scaffold;
    • (3) a heterologous object sequence; and
    • (4) a primer binding site (PBS) sequence.


One or more template RNAs described in Tables 1-5 can be tested as described in this example. The heterologous object sequences and PBS sequences were designed to correct the AATD mutation in a landing pad by replacing an “A” nucleotide with a “G” nucleotide at the mutation site via gene editing, to reverse an E342K mutation in the corresponding protein.


A cell line is created to have a “landing pad” or a stable integration that mimics a region of the SERPINA1 gene that contains the SERPINA1 mutation site and flanking sequences. The DNA for the landing pad is chemically synthesized and cloned into the pLenti-N-tGFP vector. The cloned landing pad sequence in the lentiviral expression vector is confirmed and the sequence is verified by Sanger sequencing of the landing pad. The sequence verified plasmids (9 μg) along with the lentiviral packaging mix (9 μg, Biosettia) are transfected using Lipofectamine2000™ according to the manufacturer instructions into a packaging cell line, LentiX-293T (Takara Bio). The transfected cells are incubated at 37° C., 5% CO2 for 48 hours (including one medium change at 24 hrs) and the viral particle containing medium is collected from the cell culture dish. The collected medium is filtered through a 0.2 μm filter to remove cell debris and is prepared for transduction of HEK293T cells. The virus-containing medium is diluted in DMEM and mixed with polybrene to prepare a dilution series for transduction of HEK293T cells where the final concentration of polybrene is 8 pg/ml. The HEK293T cells are grown in virus containing medium for 48 hours and then split with fresh medium. The split cells are grown to confluence and transduction efficiency of the different dilutions of virus is measured by GFP expression via flow cytometry and ddPCR detection of the genomic integrated lentivirus that contained GFP and the SERPINA1 landing pads.


A gene modifying system comprising (i) a compatible gene modifying polypeptide described herein, e.g., having: an NLS of Table 11, a compatible Cas9 domain having a sequence of Table 8, a linker of Table 10, an RT sequence of Table 6 (e.g., MLVMS P03355 PLV919), and a second NLS of Table 11 and (ii) a template RNA of any of Tables 1-5 is transfected into the HEK293T landing pad cell line. The gene modifying polypeptide and the template RNAs are delivered by nucleofection in RNA format. Specifically, 1 μg of gene modifying polypeptide mRNA is combined with 10 μM template RNAs. The mRNA and template RNAs are added to 254, SF buffer containing 250,000 HEK293T landing pad cells and cells are nucleofected using program DS-150. After nucleofection, are were grown at 37° C., 5% CO2 for 3 days prior to cell lysis and genomic DNA extraction. To analyze gene editing activity, primers flanking the SERPINA1 site are used to amplify across the locus. Amplicons are analyzed via short read sequencing using an Illumina MiSeq. In some embodiments, the assay will indicate that at least 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of copies of the SERPINA1 gene in the sample are converted to the desired wild-type sequence.


Example 2: Gene Modifying Polypeptide Selection by Pooled Screening in HEK293T & U2OS Cells

This example describes the use of an RNA gene modifying system for the targeted editing of a coding sequence in the human genome. More specifically, this example describes the infection of HEK293T and U2OS cells with a library of gene modifying candidates, followed by transfection of a template guide RNA (tgRNA) for in vitro gene modifying in the cells, e.g., as a means of evaluating a new gene modifying polypeptide for editing activity in human cells by a pooled screening approach.


The gene modifying polypeptide library candidates assayed herein each comprise: 1) a S. pyogenes (Spy) Cas9 nickase containing an N863A mutation that inactivates one endonuclease active site; 2) one of the 122 peptide linkers depicted at Table 10; and 3) a reverse transcriptase (RT) domain from Table 6 of retroviral origin. The particular retroviral RT domains utilized were selected if they were expected to function as a monomer. For each selected RT domain, the wild-type sequences were tested, as well as versions with point mutations installed in the primary wild-type sequence. In particular, 143 RT domains were tested, either wild type or containing various mutations. In total, 17,446 Cas-linker-RT gene modifying polypeptides were tested.


The system described here is a two-component system comprising: 1) an expression plasmid encoding a human codon-optimized gene modifying polypeptide library candidate within a lentiviral cassette, and 2) a tgRNA expression plasmid expressing a non-coding tgRNA sequence that is recognized by Cas and localizes it to the genomic locus of interest, and that also templates reverse transcription of the desired edit into the genome by the RT domain, driven by a U6 promoter. The lentiviral cassette comprises: (i) a CMV promoter for expression in mammalian cells; (ii) a gene modifying polypeptide library candidate as shown; (iii) a self-cleaving T2A polypeptide; (iv) a puromycin resistance gene enabling selection in mammalian cells; and (v) a polyA tail termination signal.


To prepare a pool of cells expressing gene modifying polypeptide library candidates, HEK293T or U2OS cells were transduced with pooled lentiviral preparations of the gene modifying candidate plasmid library. HEK293 Lenti-X cells were seeded in 15 cm plates (12×106 cells) prior to lentiviral plasmid transfection. Lentiviral plasmid transfection using the Lentiviral Packaging Mix (Biosettia, 27 ug) and the plasmid DNA for the gene modifying candidate library (27 ug) was performed the following day using Lipofectamine 2000 and Opti-MEM media according to the manufacturer's protocol. Extracellular DNA was removed by a full media change the next day and virus-containing media was harvested 48 hours after. Lentiviral media was concentrated using Lenti-X Concentrator (TaKaRa Biosciences) and 5 mL lentiviral aliquots were made and stored at −80° C. Lentiviral titering was performed by enumerating colony forming units post Puromycin selection. HEK293T or U2OS cells carrying a BFP-expressing genomic landing pad were seeded at 6×107 cells in culture plates and transduced at a 0.3 multiplicity of infection (MOI) to minimize multiple infections per cell. Puromycin (2.5 ug/mL) was added 48 hours post infection to allow for selection of infected cells. Cells were kept under puromycin selection for at least 7 days and then scaled up for tgRNA electroporation.


To determine the genome-editing capacity of the gene modifying library candidates in the assay, infected BFP-expressing HEK293T or U2OS cells were then transfected by electroporation of 250,000 cells/well with 200 ng of a tgRNA (either g4 or g10) plasmid, designed to convert BFP to GFP, at sufficient cell count for >1000× coverage per library candidate.


The g4 tgRNA (5′ to 3′) is as follows: 20 nucleotide spacer region (GCCGAAGCACTGCACGCCGT; SEQ ID NO: 11,011), a scaffold region (GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAAA AAGTGGCACCGAGTCGGTGC; SEQ ID NO: 11,012), the template region encoding the single base pair substitution to change BFP to GFP (bold) and a PAM inactivation that introduces a synonymous point mutation in the SpyCas9 PAM (NGG to NCG) that prevents re-engagement of the gene modifying polypeptide upon completion of a functional gene modifying reaction (underline) (ACCCTGACGTACG; SEQ ID NO: 11,013), and the 13 nucleotide PBS (GCGTGCAGTGCTT; SEQ ID NO: 11,014).


Similarly, the g10 tgRNA (5′ to 3′) is as follows: 20 nucleotide spacer region (AGAAGTCGTGCTGCTTCATG; SEQ ID NO: 11,015), a scaffold region (GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAAA AAGTGGCACCGAGTCGGTGC; SEQ ID NO: 11,016), the template region encoding the single base pair substitution to change BFP to GFP (bold) and a PAM inactivation that introduces a synonymous point mutation in the SpyCas9 PAM (NGG to NGA) that prevents re-engagement of the gene modifying polypeptide upon completion of a functional gene modifying reaction (underline) (ACCCTGACCTACGGCGTGCAGTGCTTCGGCCGCTACCCCGATCACAT; SEQ ID NO: 11,017), and 13 nucleotide PBS (GAAGCAGCACGAC; SEQ ID NO: 11,018).


To assess the genome-editing capacity of the various constructs in the assay, cells were sorted by Fluorescence-Activated Cell Sorting (FACS) for GFP expression 6-7 days post-electroporation. Cells were sorted and harvested as distinct populations of unedited (BFP+) cells, edited (GFP+) cells and imperfect edit (BFP-, GFP-) cells. A sample of unsorted cells was also harvested as the input population to determine enrichment during analysis.


To determine which gene modifying library candidates have genome-editing capacity in this assay, genomic DNA (gDNA) was harvested from sorted and unsorted cell populations, and analyzed by sequencing the gene modifying library candidates in each population. Briefly, gene modifying sequences were amplified from the genome using primers specific to the lentiviral cassette, amplified in a second round of PCR to dilute genomic DNA, and then sequenced using Oxford Nanopore Sequencing Technology according to the manufacturer's protocol.


After quality control of sequencing reads, reads of at least 1500 and no more than 3200 nucleotides were mapped to the gene modifying polypeptide library sequences and those containing a minimum of an 80% match to a library sequence were considered to be successfully aligned to a given candidate. To identify gene modifying candidates capable of performing gene editing in the assay, the read count of each library candidate in the edited population was compared to its read count in the initial, unsorted population. For purposes of this pooled screen, gene modifying candidates with genome-editing capacity were selected as those candidates that were enriched in the converted (GFP+) population relative to unsorted (input) cells and wherein the enrichment was determined to be at or above the enrichment level of a reference (Element ID No: 17380).


A large number of gene modifying polypeptide candidates were determined to be enriched in the GFP+ cell populations. For example, of the 17,446 candidates tested, over 3,300 exhibited enrichment in GFP+sorted populations (relative to unsorted) that was at least equivalent to that of the reference under similar experimental conditions (HEK293T using g4 tgRNA; HEK293T cells using g10 tgRNA; or U2OS cells using g4 tgRNA), shown in Table D. Although the 17,446 candidates were also tested in U2OS cells using g10 tgRNA, the pooled screen did not yield candidates that were enriched in the converted (GFP+) population relative to unsorted (input) cells under that experimental condition; further investigation is required to explain these results.









TABLE D







Combinations of linker and RT sequences


screened. The amino acid sequence of


each RT in this table is provided in


Table 6.










SEQ



Linker amino
ID
RT domain


acid sequence
NO:
name





EAAAKGSS
12,001
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,002
MLVMS_P03355_PLV919


AK







PAPEAAAK
12,003
MLVFF_P26809_3mutA





EAAAKPAPGGG
12,004
MLVFF_P26809_3mutA





GSSGSSGSSGSSGSSGSS
12,005
PERV_Q4VFZ2_3mut





PAPGGGEAAAK
12,006
MLVAV_P03356_3mutA





AEAAAKEAAAKEAAAKEA
12,007
MLVMS_P03355_PLV919


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSEAAAK
12,008
MLVFF_P26809_3mutA





EAAAKPAPGGS
12,009
MLVFF_P26809_3mutA





GGSGGSGGSGGSGGSGGS
12,010
MLVFF_P26809_3mutA





AEAAAKEAAAKEAAAKEA
12,011
XMRV6_A1Z651_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







AEAAAKEAAAKEAAAKEA
12,012
PERV_Q4VFZ2_3mutA_WS


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKEAAAKEAAAK
12,013
MLVFF_P26809_3mutA





PAPEAAAKGSS
12,014
MLVFF_P26809_3mutA





AEAAAKEAAAKEAAAKEA
12,015
PERV_Q4VFZ2_3mutA_WS


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKEAAAKEAAAK
12,016
PERV_Q4VFZ2_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,017
AVIRE_P03360_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPAPAPAPAP
12,018
MLVCB_P08361_3mutA





PAPAPAPAPAP
12,019
MLVFF_P26809_3mutA





EAAAKGGSPAP
12,020
PERV_Q4VFZ2_3mutA_WS





PAP

MLVMS_P03355_PLV919





PAPGGGGSS
12,022
WMSV_P03359_3mutA





SGSETPGTSESATPES
12,023
MLVFF_P26809_3mutA





PAPEAAAKGSS
12,024
XMRV6_A1Z651_3mutA





EAAAKGGSGGG
12,025
MLVMS_P03355_PLV919





GGGGSGGGGS
12,026
MLVFF_P26809_3mutA





GGGPAPGSS
12,027
MLVAV_P03356_3mutA





GGSGGSGGSGGSGGSGGS
12,028
XMRV6_A1Z651_3mut





GGGGSGGGGSGGGGSGGG
12,029
MLVCB_P08361_3mutA


GSGGGGSGGGGS







GSSPAP
12,030
AVIRE_P03360_3mutA





EAAAKGSSPAP
12,031
MLVFF_P26809_3mutA





GSSGGGEAAAK
12,032
MLVFF_P26809_3mutA





GGSGGSGGSGGSGGSGGS
12,033
MLVMS_P03355_3mutA_WS





PAPAPAPAP
12,034
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,035
XMRV6_A1Z651_3mutA


AK







EAAAKGGSPAP
12,036
MLVMS_P03355_3mutA_WS





PAPGGSEAAAK
12,037
AVIRE_P03360_3mutA





GGGGSGGGGSGGGGSGGG
12,038
AVIRE_P03360_3mutA


GSGGGGSGGGGS







EAAAKGGGGSEAAAK
12,039
MLVCB_P08361_3mutA





AEAAAKEAAAKEAAAKEA
12,040
WMSV_P03359_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSS

MLVMS_P03355_PLV919





GSSGSSGSSGSS
12,042
MLVMS_P03355_PLV919





GSSPAPEAAAK
12,043
XMRV6_A1Z651_3mutA





GGSPAPEAAAK
12,044
MLVFF_P26809_3mutA





GGGEAAAKGGS
12,045
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,046
PERV_Q4VFZ2_3mutA_WS


AKEAAAK







GGGGGGGG
12,047
PERV_Q4VFZ2_3mut





GGGPAP
12,048
MLVCB_P08361_3mutA





PAPAPAPAPAPAP
12,049
MLVCB_P08361_3mutA





GGSGGSGGSGGSGGSGGS
12,050
MLVCB_P08361_3mutA





PAP

MLVMS_P03355_3mutA_WS





GGSGGSGGSGGSGGSGGS
12,052
PERV_Q4VFZ2_3mutA_WS





PAPAPAPAPAPAP
12,053
MLVMS_P03355_PLV919





EAAAKPAPGSS
12,054
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,055
MLVMS_P03355_3mutA_WS


AK







EAAAKGGS
12,056
MLVMS_P03355_3mutA_WS





GGGGSEAAAKGGGGS
12,057
MLVFF_P26809_3mutA





EAAAKPAPGSS
12,058
MLVFF_P26809_3mutA





GGGGSGGGGSGGGGSGGG
12,059
MLVMS_P03355_PLV919


GS







EAAAKGGGGGS
12,060
MLVMS_P03355_PLV919





GGSPAP
12,061
XMRV6_A1Z651_3mutA





EAAAKGGGPAP
12,062
MLVMS_P03355_PLV919





EAAAKEAAAKEAAAKEAA
12,063
MLVFF_P26809_3mutA


AKEAAAK







PAP

MLVCB_P08361_3mutA





EAAAK
12,065
XMRV6_A1Z651_3mutA





GGSGSSPAP
12,066
PERV_Q4VFZ2_3mutA_WS





GSSGSSGSSGSSGSSGSS
12,067
MLVMS_P03355_PLV919





GSSEAAAKGGG
12,068
MLVAV_P03356_3mutA





GGGEAAAKGGS
12,069
XMRV6_A1Z651_3mutA





EAAAKGGGGSEAAAK
12,070
MLVAV_P03356_3mutA





GGGGSGGGGSGGGGS
12,071
MLVFF_P26809_3mutA





GGGGSGGGGSGGGGSGGG
12,072
AVIRE_P03360_3mutA


GS







SGSETPGTSESATPES
12,073
AVIRE_P03360_3mutA





GGGEAAAKPAP
12,074
MLVFF_P26809_3mutA





EAAAKGSSGGG
12,075
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,076
WMSV_P03359_3mut


AKEAAAK







GGSGGSGGSGGS
12,077
XMRV6_A1Z651_3mutA





GGSEAAAKPAP
12,078
MLVFF_P26809_3mutA





EAAAKGSSGGG
12,079
XMRV6_A1Z651_3mutA





GGGGS
12,080
MLVFF_P26809_3mutA





GGGEAAAKGSS
12,081
MLVMS_P03355_PLV919





PAPAPAPAPAPAP
12,082
MLVAV_P03356_3mutA





GGGGSGGGGSGGGGSGGG
12,083
MLVCB_P08361_3mutA


GS







GGGEAAAKGSS
12,084
MLVCB_P08361_3mutA





PAPGGSGSS
12,085
MLVFF_P26809_3mutA





GSAGSAAGSGEF
12,086
MLVCB_P08361_3mutA





PAPGGSEAAAK
12,087
MLVMS_P03355_3mutA_WS





GGSGSS
12,088
XMRV6_A1Z651_3mutA





PAPGGGGSS
12,089
MLVMS_P03355_PLV919





GSSGSSGSS
12,090
XMRV6_A1Z651_3mut





AEAAAKEAAAKEAAAKEA
12,091
MLVMS_P03355_3mutA_WS


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAK
12,092
MLVMS_P03355_PLV919





GSSGSSGSSGSS
12,093
MLVFF_P26809_3mutA





PAPGGGGSS
12,094
MLVCB_P08361_3mutA





GGGEAAAKGGS
12,095
MLVCB_P08361_3mutA





PAPGGGEAAAK
12,096
MLVMS_P03355_PLV919





GGGGGSPAP
12,097
XMRV6_A1Z651_3mutA





EAAAKGGS
12,098
XMRV6_A1Z651_3mutA





EAAAKGSSPAP
12,099
XMRV6_A1Z651_3mut





PAPEAAAK
12,100
MLVAV_P03356_3mutA





GGSGGSGGSGGS
12,101
MLVMS_P03355_3mutA_WS





GGGPAPGGS
12,102
MLVMS_P03355_PLV919





GSSGSSGSSGSS
12,103
PERV_Q4VFZ2_3mutA_WS





EAAAKPAPGGS
12,104
MLVCB_P08361_3mutA





GSSGSS
12,105
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,106
MLVCB_P08361_3mutA


AK







EAAAKEAAAKEAAAKEAA
12,107
FLV_P10273_3mutA


AK







GSS

MLVFF_P26809_3mutA





EAAAKEAAAK
12,109
MLVMS_P03355_3mutA_WS





PAPEAAAKGGG
12,110
MLVAV_P03356_3mutA





GGSGSSEAAAK
12,111
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,112
PERV_Q4VFZ2


AKEAAAK







GSSEAAAKPAP
12,113
AVIRE_P03360_3mutA





EAAAKEAAAKEAAAKEAA
12,114
MLVCB_P08361_3mutA


AKEAAAK







EAAAKGGG
12,115
MLVFF_P26809_3mutA





GSSPAPGGG
12,116
MLVCB_P08361_3mutA





GGGPAPGSS
12,117
MLVMS_P03355_PLV919





GGGGGS
12,118
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,119
PERV_Q4VFZ2_3mut


AKEAAAKEAAAK







GGGGSGGGGSGGGGSGGG
12,120
WMSV_P03359_3mutA


GSGGGGS







EAAAKEAAAKEAAAK
12,121
PERV_Q4VFZ2_3mut





PAPAPAPAP
12,122
MLVCB_P08361_3mutA





GSSGSSGSSGSSGSS
12,123
PERV_Q4VFZ2_3mut





GGGGSSEAAAK
12,124
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGS
12,125
MLVCB_P08361_3mutA





PAPEAAAKGGS
12,126
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
12,127
MLVCB_P08361_3mutA


AKEAAAKEAAAK







EAAAKGGGGSEAAAK
12,128
MLVMS_P03355_PLV919





EAAAKGGGGSEAAAK
12,129
MLVMS_P03355_3mutA_WS





EAAAKGGGPAP
12,130
XMRV6_A1Z651_3mut





EAAAKEAAAKEAAAKEAA
12,131
MLVMS_P03355_3mutA_WS


AKEAAAK







AEAAAKEAAAKEAAAKEA
12,132
FLV_P10273_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSEAAAKGGG
12,133
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGSGGG
12,134
KORV_Q9TTC1-Pro_3mutA


GSGGGGSGGGGS







GGGPAPGGS
12,135
MLVCB_P08361_3mutA





PAPAPAPAPAPAP
12,136
XMRV6_A1Z651_3mutA





GGSGSSGGG
12,137
XMRV6_A1Z651_3mutA





GGSGSSGGG
12,138
MLVCB_P08361_3mutA





GGGEAAAKGGS
12,139
MLVMS_P03355_3mutA_WS





EAAAK
12,140
MLVCB_P08361_3mutA





GGSPAPGSS
12,141
MLVMS_P03355_3mutA_WS





GGGGSSEAAAK
12,142
PERV_Q4VFZ2_3mut





PAPAPAPAPAP
12,143
MLVBM_Q7SVK7_3mut





EAAAKEAAAKEAAAKEAA
12,144
MLVAV_P03356_3mutA


AK







GGGGGSGSS
12,145
MLVCB_P08361_3mutA





EAAAKGSSPAP
12,146
MLVMS_P03355_3mutA_WS





PAPAPAPAPAPAP
12,147
MLVMS_P03355_3mutA_WS





GSSGGGGGS
12,148
MLVMS_P03355_3mutA_WS





PAPGSSGGG
12,149
MLVMS_P03355_PLV919





GGSGGGPAP
12,150
MLVCB_P08361_3mutA





GGGGGGG
12,151
MLVCB_P08361_3mutA





GSSGSSGSSGSSGSSGSS
12,152
MLVCB_P08361_3mutA





GGGPAPGGS
12,153
MLVFF_P26809_3mutA





EAAAKGGSGGG
12,154
PERV_Q4VFZ2_3mut





EAAAKGGGGSS
12,155
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSSGSSGSS
12,156
MLVMS_P03355_3mut





GGGGSGGGGSGGGGSGGG
12,157
MLVBM_Q7SVK7_3mutA_WS


GS







PAPAPAPAPAP
12,158
MLVMS_P03355_PLV919





GGGEAAAKGGS
12,159
MLVMS_P03355_PLV919





AEAAAKEAAAKEAAAKEA
12,160
MLVMS_P03355_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSAGSAAGSGEF
12,161
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSSGSS
12,162
MLVFF_P26809_3mutA





EAAAKGGSGSS
12,163
MLVFF_P26809_3mutA





PAPGGG
12,164
MLVFF_P26809_3mutA





GGGPAPGSS
12,165
XMRV6_A1Z651_3mutA





PAPEAAAKGGS
12,166
AVIRE_P03360_3mutA





PAPGGGEAAAK
12,167
MLVFF_P26809_3mut





GGGGSSEAAAK
12,168
MLVCB_P08361_3mutA





EAAAK
12,169
MLVMS_P03355_PLV919





GGGGSGGGGSGGGGSGGG
12,170
BAEVM_P10272_3mutA


GGGGGSGGGGS







GGSGGGEAAAK
12,171
MLVMS_P03355_PLV919





AEAAAKEAAAKEAAAKEA
12,172
MLVFF_P26809_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSPAPGGS
12,173
XMRV6_A1Z651_3mutA





GGSGGGPAP
12,174
MLVMS_P03355_PLV919





EAAAK
12,175
AVIRE_P03360_3mutA





GSS

XMRV6_A1Z651_3mutA





GGSGGSGGS
12,177
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,178
AVIRE_P03360_3mut


AK







PAPEAAAKGGG
12,179
PERV_Q4VFZ2_3mutA_WS





GGGGGSEAAAK
12,180
BAEVM_P10272_3mutA





GGSGSSGGG
12,181
MLVMS_P03355_3mutA_WS





GGGGGGG
12,182
MLVMS_P03355_3mutA_WS





GSSEAAAKPAP
12,183
PERV_Q4VFZ2_3mut





GGGGGSEAAAK
12,184
WMSV_P03359_3mut





GGGGSGGGGSGGGGSGGG
12,185
MLVFF_P26809_3mut


GSGGGGS







GGGEAAAKGGS
12,186
AVIRE_P03360_3mutA





GGSPAPGGG
12,187
AVIRE_P03360_3mutA





GSAGSAAGSGEF
12,188
MLVAV_P03356_3mutA





EAAAK
12,189
MLVAV_P03356_3mutA





EAAAKPAPGSS
12,190
WMSV_P03359_3mutA





EAAAKEAAAKEAAAKEAA
12,191
PERV_Q4VFZ2_3mutA_WS


AKEAAAKEAAAK







GGSEAAAKPAP
12,192
MLVCB_P08361_3mutA





PAPAPAPAPAPAP
12,193
MLVBM_Q7SVK7_3mutA_WS





GGSPAPGGG
12,194
MLVMS_P03355_3mutA_WS





GGSEAAAKGGG
12,195
MLVMS_P03355_3mut





GGSGGSGGSGGS
12,196
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,197
MLVFF_P26809_3mutA


AKEAAAKEAAAK







GGG

AVIRE_P03360_3mutA





AEAAAKEAAAKEAAAKEA
12,199
PERV_Q4VFZ2_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSGGSGGSGGS
12,200
MLVMS_P03355_3mutA_WS





GGGEAAAK
12,201
MLVCB_P08361_3mutA





GSSGSSGSSGSSGSSGSS
12,202
MLVMS_P03355_3mutA_WS





GSSGGGPAP
12,203
MLVMS_P03355_3mutA_WS





GSSEAAAKPAP
12,204
MLVFF_P26809_3mutA





EAAAKEAAAK
12,205
MLVMS_P03355_PLV919





GGGGSGGGGGGGGSGGGG
12,206
MLVCB_P08361_3mut


SGGGGSGGGGS







GGGGGG
12,207
MLVMS_P03355_3mutA_WS





GGSGSSGGG
12,208
MLVFF_P26809_3mutA





GSSGGGEAAAK
12,209
PERV_Q4VFZ2_3mutA_WS





PAPAPAPAPAP
12,210
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
12,211
SFV3L_P27401_2mut


AKEAAAKEAAAK







EAAAKGGSGGG
12,212
BAEVM_P10272_3mutA





GGGGSSPAP
12,213
PERV_Q4VFZ2_3mutA_WS





GGGEAAAKPAP
12,214
MLVMS_P03355_PLV919





GGSGGGPAP
12,215
BAEVM_P10272_3mutA





PAPGSSGGS
12,216
MLVMS_P03355_PLV919





GGSGGGPAP
12,217
MLVMS_P03355_3mutA_WS





EAAAKGGSPAP
12,218
PERV_Q4VFZ2_3mutA_WS





EAAAKGGSGGG
12,219
MLVMS_P03355_3mutA_WS





PAPGSSGGG
12,220
MLVFF_P26809_3mutA





GSSEAAAKGGS
12,221
MLVFF_P26809_3mutA





PAPGSSEAAAK
12,222
MLVFF_P26809_3mutA





EAAAKGSSPAP
12,223
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAKEAAAKEAA
12,224
MLVBM_Q7SVK7_3mutA_WS


AKEAAAK







PAPGSSEAAAK
12,225
MLVMS_P03355_PLV919





EAAAKGSSGGG
12,226
MLVMS_P03355_3mutA_WS





EAAAKGGGGGS
12,227
AVIRE_P03360_3mutA





EAAAKEAAAKEAAAK
12,228
MLVMS_P03355_PLV919





PAPAPAPAPAPAP
12,229
MLVFF_P26809_3mutA





GGGGSGGGGSGGGGS
12,230
MLVCB_P08361_3mutA





PAPGGSEAAAK
12,231
MLVCB_P08361_3mutA





PAPGSSEAAAK
12,232
MLVBM_Q7SVK7_3mutA_WS





PAPEAAAKGSS
12,233
AVIRE_P03360_3mutA





GGSPAPGSS
12,234
WMSV_P03359_3mutA





PAPGGSGGG
12,235
MLVMS_P03355_PLV919





EAAAKGGSGSS
12,236
MLVMS_P03355_3mutA_WS





GGSGGG
12,237
MLVFF_P26809_3mutA





GGSEAAAKGSS
12,238
KORV_Q9TTC1_3mutA





AEAAAKEAAAKEAAAKEA
12,239
MLVCB_P08361_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPAPAPAPAPAP
12,240
PERV_Q4VFZ2_3mutA_WS





PAPEAAAK
12,241
MLVMS_P03355_3mutA_WS





GGSEAAAKGGG
12,242
MLVMS_P03355_PLV919





GSSPAP
12,243
MLVMS_P03355_3mutA_WS





GGGGSS
12,244
MLVMS_P03355_PLV919





GGGEAAAKPAP
12,245
AVIRE_P03360_3mutA





EAAAKPAPGGS
12,246
MLVAV_P03356_3mutA





EAAAKGGGPAP
12,247
MLVAV_P03356_3mutA





PAPGGSEAAAK
12,248
BAEVM_P10272_3mutA





PAPGGSGSS
12,249
MLVMS_P03355_3mutA_WS





PAPGGSGSS
12,250
AVIRE_P03360_3mutA





GGSGGGPAP
12,251
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,252
BAEVM_P10272_3mutA


AK







GGGGSGGGGSGGGGSGGG
12,253
MLVMS_P03355_PLV919


GSGGGGS







GGGGSSPAP
12,254
MLVCB_P08361_3mutA





GSSGGGPAP
12,255
MLVFF_P26809_3mutA





GGGGSSGGS
12,256
MLVMS_P03355_PLV919





GGSGGG
12,257
MLVCB_P08361_3mutA





GSSGGGGGS
12,258
MLVMS_P03355_PLV919





SGGSSGGSSGSETPGTSE
12,259
XMRV6_A1Z651_3mutA


SATPESSGGSSGGSS







GGGGGSGSS
12,260
KORV_Q9TTC1_3mut





GGGEAAAKGGS
12,261
BAEVM_P10272_3mutA





GGSGGG
12,262
BAEVM_P10272_3mutA





PAPAPAP
12,263
KORV_Q9TTC1-Pro_3mut





AEAAAKEAAAKEAAAKEA
12,264
SFV3L_P27401_2mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







AEAAAKEAAAKEAAAKEA
12,265
MLVBM_Q7SVK7_3mutA_WS


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSGSSGSSGSSGSS
12,266
MLVMS_P03355_3mutA_WS





GSSGGGEAAAK
12,267
MLVMS_P03355_3mutA_WS





GSSGGSEAAAK
12,268
MLVFF_P26809_3mutA





PAP

MLVMS_P03355_PLV919





EAAAKGGGGSEAAAK
12,270
MLVBM_Q7SVK7_3mutA_WS





PAPAP
12,271
AVIRE_P03360_3mutA





PAP

MLVFF_P26809_3mutA





GSSGGG
12,273
MLVMS_P03355_3mut





GSSPAPGGS
12,274
MLVFF_P26809_3mutA





PAPAPAPAP
12,275
XMRV6_A1Z651_3mutA





EAAAKGSSGGS
12,276
PERV_Q4VFZ2_3mut





PAPEAAAKGGG
12,277
KORV_Q9TTC1-Pro_3mutA





PAPGGS
12,278
MLVCB_P08361_3mutA





EAAAKGGG
12,279
MLVCB_P08361_3mutA





GSSEAAAKPAP
12,280
MLVMS_P03355_PLV919





PAPGGS
12,281
MLVFF_P26809_3mutA





EAAAKGGS
12,282
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
12,283
FLV_P10273_3mutA


AKEAAAKEAAAK







PAPGGSEAAAK
12,284
MLVAV_P03356_3mutA





GSS

MLVCB_P08361_3mutA





GSSGSSGSSGSS
12,286
AVIRE_P03360_3mutA





GSSGSSGSS
12,287
MLVFF_P26809_3mutA





GSSGGG
12,288
MLVMS_P03355_PLV919





EAAAK
12,289
MLVFF_P26809_3mutA





GGSPAPEAAAK
12,290
MLVCB_P08361_3mutA





GGSGSS
12,291
MLVCB_P08361_3mutA





GSSPAPGGG
12,292
MLVMS_P03355_PLV919





EAAAKEAAAKEAAAKEAA
12,293
MLVAV_P03356_3mutA


AKEAAAK







EAAAKGSSPAP
12,294
FLV_P10273_3mutA





GGGGSS
12,295
XMRV6_A1Z651_3mutA





GGSPAPGSS
12,296
MLVMS_P03355_PLV919





EAAAKEAAAKEAAAKEAA
12,297
MLVMS_P03355_3mutA_WS


AKEAAAK







PAPEAAAKGGG
12,298
FLV_P10273_3mutA





EAAAKPAPGGS
12,299
XMRV6_A1Z651_3mut





PAPAP
12,300
BAEVM_P10272_3mutA





EAAAKEAAAKEAAAKEAA
12,301
MLVMS_P03355_PLV919


AK







GSSPAPGGG
12,302
MLVMS_P03355_PLV919





EAAAKGGGPAP
12,303
KORV_Q9TTC1_3mutA





PAPEAAAK
12,304
MLVMS_P03355_PLV919





PAPGGGEAAAK
12,305
PERV_Q4VFZ2_3mutA_WS





EAAAKGSSGGS
12,306
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAK
12,307
MLVMS_P03355_PLV919





GSSEAAAK
12,308
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSS
12,309
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGSGGG
12,310
MLVMS_P03355_3mutA_WS


GS







EAAAKGGGGSEAAAK
12,311
MLVMS_P03355_3mut





GGS

MLVCB_P08361_3mutA





GGGGSGGGGSGGGGSGGG
12,313
XMRV6_A1Z651_3mutA


GSGGGGGGGGS







GGSGSSPAP
12,314
MLVCB_P08361_3mutA





GGGGSGGGGSGGGGS
12,315
XMRV6_A1Z651_3mutA





PAPAPAPAPAP
12,316
BAEVM_P10272_3mutA





PAPAPAPAPAP
12,317
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,318
MLVBM_Q7SVK7_3mut


AK







GGGGSGGGGSGGGGSGGG
12,319
BAEVM_P10272_3mutA


GSGGGGS







GGSGGSGGS
12,320
MLVMS_P03355_3mutA_WS





EAAAKPAPGSS
12,321
MLVMS_P03355_PLV919





GSS

MLVMS_P03355_3mutA_WS





PAPEAAAKGGS
12,323
MLVMS_P03355_3mutA_WS





GGGPAPGGS
12,324
MLVMS_P03355_3mutA_WS





EAAAKGGGGSS
12,325
MLVAV_P03356_3mutA





GSSGSSGSSGSSGSS
12,326
MLVFF_P26809_3mut





SGSETPGTSESATPES
12,327
PERV_Q4VFZ2_3mut





GGSEAAAKGGG
12,328
MLVMS_P03355_3mut





GSSGSSGSSGSSGSSGSS
12,329
AVIRE_P03360_3mutA





PAPAPAPAPAPAP
12,330
AVIRE_P03360_3mut





GGSGGS
12,331
XMRV6_A1Z651_3mutA





PAPGSSEAAAK
12,332
MLVCB_P08361_3mut





GGSPAPEAAAK
12,333
PERV_Q4VFZ2_3mut





EAAAKGGGGGS
12,334
MLVCB_P08361_3mutA





GGSGGSGGSGGS
12,335
MLVMS_P03355_PLV919





GGGGSSEAAAK
12,336
MLVMS_P03355_PLV919





GSSEAAAKGGG
12,337
MLVFF_P26809_3mutA





PAPGGS
12,338
MLVMS_P03355_3mutA_WS





EAAAKGGSGGG
12,339
MLVCB_P08361_3mutA





EAAAKGGG
12,340
PERV_Q4VFZ2_3mut





PAPGGS
12,341
XMRV6_A1Z651_3mutA





GSSPAPGGG
12,342
XMRV6_A1Z651_3mutA





PAPEAAAKGGG
12,343
MLVMS_P03355_3mutA_WS





GSSEAAAKGGG
12,344
PERV_Q4VFZ2_3mutA_WS





PAPGGSEAAAK
12,345
XMRV6_A1Z651_3mutA





GGGGGS
12,346
MLVMS_P03355_3mutA_WS





GGSPAPEAAAK
12,347
MLVMS_P03355_3mutA_WS





GGGPAP
12,348
MLVFF_P26809_3mutA





PAPGSSGGG
12,349
XMRV6_A1Z651_3mutA





PAPGSSGGG
12,350
MLVBM_Q7SVK7_3mutA_WS





GGGEAAAKGSS
12,351
MLVMS_P03355_3mutA_WS





GSSEAAAKGGS
12,352
MLVCB_P08361_3mutA





PAPGGSGSS
12,353
MLVCB_P08361_3mutA





EAAAKGGGGSEAAAK
12,354
BAEVM_P10272_3mutA





PAPAPAP
12,355
PERV_Q4VFZ2_3mutA_WS





GGGGGG
12,356
MLVAV_P03356_3mutA





GSSPAPEAAAK
12,357
MLVCB_P08361_3mutA





GGSGGSGGS
12,358
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSSGSS
12,359
XMRV6_A1Z651_3mut





GGGPAPGGS
12,360
XMRV6_A1Z651_3mutA





GGGPAPEAAAK
12,361
BAEVM_P10272_3mutA





GGSGGG
12,362
AVIRE_P03360_3mutA





SGSETPGTSESATPES
12,363
PERV_Q4VFZ2_3mutA_WS





EAAAKGSSPAP
12,364
MLVMS_P03355_PLV919





GSSEAAAK
12,365
XMRV6_A1Z651_3mut





GSSGGSGGG
12,366
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,367
WMSV_P03359_3mutA


AKEAAAK







GGGGSEAAAKGGGGS
12,368
MLVMS_P03355_PLV919





PAPGGGGSS
12,369
MLVMS_P03355_3mutA_WS





SGSETPGTSESATPES
12,370
MLVMS_P03355_3mutA_WS





GGSPAPEAAAK
12,371
KORV_Q9TTC1-Pro_3mutA





GSSEAAAKGGG
12,372
MLVMS_P03355_3mutA_WS





GSSEAAAK
12,373
WMSV_P03359_3mutA





GGGGSEAAAKGGGGS
12,374
AVIRE_P03360_3mutA





GSS

WMSV_P03359_3mutA





PAPGGSEAAAK
12,376
MLVFF_P26809_3mutA





GGGGS
12,377
MLVMS_P03355_3mutA_WS





GGGPAP
12,378
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,379
MLVMS_P03355_3mutA_WS


AKEAAAKEAAAK







EAAAKPAPGSS
12,380
PERV_Q4VFZ2_3mut





EAAAKPAPGSS
12,381
MLVCB_P08361_3mutA





GGGGGG
12,382
WMSV_P03359_3mutA





EAAAKPAPGGS
12,383
MLVMS_P03355_PLV919





PAPGGGEAAAK
12,384
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
12,385
AVIRE_P03360_3mutA


AKEAAAK







GSSEAAAKPAP
12,386
XMRV6_A1Z651_3mutA





PAPGGSEAAAK
12,387
MLVBM_Q7SVK7_3mutA_WS





PAPGSS
12,388
MLVCB_P08361_3mutA





EAAAKGGG
12,389
MLVMS_P03355_3mutA_WS





EAAAKPAP
12,390
MLVCB_P08361_3mutA





PAPEAAAKGGS
12,391
MLVBM_Q7SVK7_3mutA_WS





GGSPAPGGG
12,392
MLVCB_P08361_3mutA





PAPGGSGSS
12,393
WMSV_P03359_3mutA





EAAAKEAAAKEAAAKEAA
12,394
MLVMS_P03355_PLV919


AKEAAAKEAAAK







GGSGGGPAP
12,395
MLVMS_P03355_PLV919





AEAAAKEAAAKEAAAKEA
12,396
MLVMS_P03355


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPEAAAKGSS
12,397
MLVCB_P08361_3mutA





EAAAKGSS
12,398
MLVMS_P03355_3mutA_WS





GGSGGS
12,399
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,400
BAEVM_P10272_3mutA


AKEAAAK







GGGGSEAAAKGGGGS
12,401
FLV_P10273_3mutA





GGSEAAAKGGG
12,402
MLVCB_P08361_3mutA





GSSGSSGSSGSSGSS
12,403
BAEVM_P10272_3mutA





GGGGGGGGSGGGGSGGGG
12,404
MLVFF_P26809_3mutA


SGGGGSGGGGS







EAAAKGGG
12,405
PERV_Q4VFZ2_3mut





GGGGGSEAAAK
12,406
MLVCB_P08361_3mutA





EAAAKPAPGGS
12,407
MLVMS_P03355_3mutA_WS





GGGGGSGSS
12,408
XMRV6_A1Z651_3mutA





PAPGSSEAAAK
12,409
MLVMS_P03355_3mutA_WS





GSSEAAAKPAP
12,410
MLVCB_P08361_3mutA





EAAAKGSSPAP
12,411
MLVAV_P03356_3mutA





GGGPAPGGS
12,412
WMSV_P03359_3mutA





GGSPAP
12,413
MLVMS_P03355_3mutA_WS





GGSEAAAKGGG
12,414
MLVMS_P03355_3mutA_WS





GGGGGGGG
12,415
MLVFF_P26809_3mutA





GGGGSGGGGSGGGGSGGG
12,416
MLVMS_P03355_3mutA_WS


GSGGGGSGGGGS







GGGGSGGGGSGGGGSGGG
12,417
MLVBM_Q7SVK7_3mutA_WS


GSGGGGSGGGGS







GSSPAPGGG
12,418
MLVAV_P03356_3mutA





GGGGGG
12,419
AVIRE_P03360_3mutA





GSSGGS
12,420
MLVMS_P03355_3mutA_WS





GGSPAPGSS
12,421
MLVFF_P26809_3mutA





PAPEAAAKGGG
12,422
PERV_Q4VFZ2_3mut





EAAAKGGGPAP
12,423
MLVFF_P26809_3mutA





GGGEAAAKGGS
12,424
MLVMS_P03355_PLV919





GGSGSSPAP
12,425
MLVFF_P26809_3mutA





SGSETPGTSESATPES
12,426
WMSV_P03359_3mutA





PAPGGSEAAAK
12,427
MLVBM_Q7SVK7_3mutA_WS





GGSGGG
12,428
MLVMS_P03355_PLV919





GGGGSSPAP
12,429
PERV_Q4VFZ2_3mut





GGGEAAAKGSS
12,430
MLVAV_P03356_3mutA





PAPAPAPAPAPAP
12,431
MLVMS_P03355_3mutA_WS





EAAAKGGGGSEAAAK
12,432
PERV_Q4VFZ2





EAAAKEAAAKEAAAKEAA
12,433
MLVMS_P03355_PLV919


AKEAAAK







GGGGGSEAAAK
12,434
PERV_Q4VFZ2_3mut





PAPGSSEAAAK
12,435
MLVCB_P08361_3mutA





GSAGSAAGSGEF
12,436
PERV_Q4VFZ2_3mutA_WS





EAAAKGGGGSEAAAK
12,437
MLVFF_P26809_3mutA





GGSPAPGGG
12,438
PERV_Q4VFZ2_3mutA_WS





GSSEAAAKGGG
12,439
AVIRE_P03360_3mutA





GGGEAAAKPAP
12,440
MLVMS_P03355_3mutA_WS





GGGPAP
12,441
AVIRE_P03360_3mutA





GGSEAAAK
12,442
MLVCB_P08361_3mutA





SGGSSGGSSGSETPGTSE
12,443
PERV_Q4VFZ2_3mut


SATPESSGGSSGGSS







EAAAKPAPGGS
12,444
MLVBM_Q7SVK7_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,445
XMRV6_A1Z651_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGGGGG
12,446
MLVCB_P08361_3mutA





PAPGSS
12,447
PERV_Q4VFZ2_3mut





EAAAK
12,448
PERV_Q4VFZ2_3mut





GSAGSAAGSGEF
12,449
MLVMS_P03355_3mutA_WS





PAPGGGEAAAK
12,450
PERV_Q4VFZ2_3mut





EAAAKGSSGGS
12,451
MLVFF_P26809_3mut





GGGGSEAAAKGGGGS
12,452
BAEVM_P10272_3mutA





GGGGSGGGGSGGGGS
12,453
MLVMS_P03355_PLV919





EAAAKGGGGSEAAAK
12,454
BAEVM_P10272_3mut





PAPGGGEAAAK
12,455
MLVMS_P03355_3mutA_WS





GGSEAAAKPAP
12,456
MLVMS_P03355_3mutA_WS





PAPAP
12,457
MLVCB_P08361_3mutA





PAPAP
12,458
MLVFF_P26809_3mutA





GGSPAP
12,459
AVIRE_P03360_3mutA





EAAAKGSSGGS
12,460
MLVCB_P08361_3mutA





PAPGSSGGS
12,461
AVIRE_P03360_3mutA





EAAAKGGGGSEAAAK
12,462
XMRV6_A1Z651_3mutA





PAPAPAP
12,463
BAEVM_P10272_3mutA





GGSGGSGGSGGSGGSGGS
12,464
MLVMS_P03355_PLV919





GGGGGSGSS
12,465
MLVMS_P03355_PLV919





PAPGSSEAAAK
12,466
XMRV6_A1Z651_3mut





GGSEAAAKPAP
12,467
XMRV6_A1Z651_3mutA





EAAAKEAAAKEAAAKEAA
12,468
XMRV6_A1Z651_3mut


AK







AEAAAKEAAAKEAAAKEA
12,469
WMSV_P03359_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSGGGEAAAK
12,470
XMRV6_A1Z651_3mutA





GGGEAAAK
12,471
XMRV6_A1Z651_3mutA





GGGGSGGGGSGGGGS
12,472
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGSGGS
12,473
MLVFF_P26809_3mutA





GSSGGGGGS
12,474
MLVMS_P03355_3mut





PAPGGSEAAAK
12,475
MLVMS_P03355_3mutA_WS





GSSGGSPAP
12,476
MLVMS_P03355_3mutA_WS





SGSETPGTSESATPES
12,477
XMRV6_A1Z651_3mutA





GGGGSGGGGS
12,478
MLVMS_P03355_PLV919





PAPAPAPAPAP
12,479
MLVMS_P03355_3mut





GSSGSS
12,480
XMRV6_A1Z651_3mutA





GSSEAAAKPAP
12,481
PERV_Q4VFZ2_3mut





GGSGSSGGG
12,482
MLVMS_P03355_3mutA_WS





EAAAKEAAAK
12,483
MLVCB_P08361_3mutA





GSSGSSGSSGSS
12,484
MLVMS_P03355_3mutA_WS





GSSPAPGGG
12,485
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAK
12,486
MLVMS_P03355_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,487
SFV1_P23074_2mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGSGGGGSGGGGSGGG
12,488
MLVMS_P03355_PLV919


GSGGGGSGGGGS







GSAGSAAGSGEF
12,489
MLVMS_P03355_PLV919





PAPGSSEAAAK
12,490
MLVMS_P03355_3mutA_WS





GGSEAAAK
12,491
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSSGSS
12,492
PERV_Q4VFZ2_3mutA_WS





GGSEAAAKPAP
12,493
PERV_Q4VFZ2_3mutA_WS





GGSGGSGGS
12,494
MLVCB_P08361_3mutA





EAAAKGGSGSS
12,495
MLVCB_P08361_3mutA





GGGGSGGGGSGGGGSGGG
12,496
FLV_P10273_3mutA


GSGGGGS







EAAAKEAAAKEAAAKEAA
12,497
MLVBM_Q7SVK7_3mutA_WS


AK







GGSGSSPAP
12,498
BAEVM_P10272_3mutA





EAAAKEAAAKEAAAKEAA
12,499
XMRV6_A1Z651_3mutA


AKEAAAK







GGGGSGGGGSGGGGSGGG
12,500
MLVBM_Q7SVK7_3mutA_WS


GSGGGGS







GGSGSS
12,501
WMSV_P03359_3mutA





PAPEAAAK
12,502
MLVCB_P08361_3mutA





EAAAKPAP
12,503
BAEVM_P10272_3mutA





GSSPAP
12,504
PERV_Q4VFZ2_3mutA_WS





GGGPAP
12,505
PERV_Q4VFZ2_3mutA_WS





EAAAKGGSGSS
12,506
MLVMS_P03355_3mutA_WS





EAAAKGGGGSEAAAK
12,507
AVIRE_P03360_3mutA





GGSGGG
12,508
KORV_Q9TTC1-Pro_3mutA





GSSPAP
12,509
MLVFF_P26809_3mutA





GGSGSSEAAAK
12,510
BAEVM_P10272_3mutA





PAPGSSGGS
12,511
BAEVM_P10272_3mutA





GGGGGG
12,512
MLVFF_P26809_3mutA





PAPGGSEAAAK
12,513
MLVMS_P03355_PLV919





PAPGGS
12,514
MLVMS_P03355_PLV919





GGSGGSGGSGGS
12,515
BAEVM_P10272_3mutA





GSSPAP
12,516
MLVCB_P08361_3mutA





PAPAPAPAP
12,517
MLVMS_P03355_3mutA_WS





GGGGGG
12,518
MLVCB_P08361_3mutA





GSSGSSGSSGSSGSSGSS
12,519
KORV_Q9TTC1-Pro_3mutA





GSSEAAAKGGS
12,520
BAEVM_P10272_3mutA





GGSEAAAK
12,521
FLV_P10273_3mutA





GGSGGSGGSGGSGGS
12,522
KORV_Q9TTC1-Pro_3mutA





GSSPAPEAAAK
12,523
PERV_Q4VFZ2_3mut





GSSGSSGSSGSSGSS
12,524
XMRV6_A1Z651_3mutA





EAAAKPAPGGS
12,525
MLVMS_P03355_3mut





SGGSSGGSSGSETPGTSE
12,526
FLV_P10273_3mut


SATPESSGGSSGGSS







GGSPAPEAAAK
12,527
XMRV6_A1Z651_3mut





EAAAKGGSGGG
12,528
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
12,529
MLVFF_P26809_3mutA


AK







GSSPAP
12,530
WMSV_P03359_3mutA





PAPAPAPAP
12,531
MLVAV_P03356_3mutA





PAPGGSEAAAK
12,532
KORV_Q9TTC1_3mut





GGSGSSEAAAK
12,533
MLVBM_Q7SVK7_3mutA_WS





GSSGGG
12,534
MLVCB_P08361_3mutA





GGGEAAAKGSS
12,535
PERV_Q4VFZ2_3mut





PAPGGSGGG
12,536
MLVFF_P26809_3mutA





AEAAAKEAAAKEAAAKEA
12,537
FFV_O93209


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPGGGGSS
12,538
MLVMS_P03355_3mutA_WS





EAAAKGGS
12,539
MLVAV_P03356_3mutA





EAAAKEAAAKEAAAKEAA
12,540
MLVBM_Q7SVK7_3mutA_WS


AKEAAAKEAAAK







GGSGGSGGS
12,541
WMSV_P03359_3mutA





PAPAP
12,542
MLVMS_P03355_3mutA_WS





GSSGGGEAAAK
12,543
MLVAV_P03356_3mutA





GGGGSSEAAAK
12,544
MLVFF_P26809_3mutA





EAAAKGSSGGS
12,545
MLVMS_P03355_PLV919





EAAAKGGGGSEAAAK
12,546
MLVMS_P03355_3mutA_WS





GGGGGGGG
12,547
MLVMS_P03355_PLV919





GSSGSSGSS
12,548
MLVMS_P03355_PLV919





GGGEAAAKPAP
12,549
PERV_Q4VFZ2_3mutA_WS





GGGGGSGSS
12,550
MLVMS_P03355_3mutA_WS





GGGGGGG
12,551
MLVMS_P03355_PLV919





GGS

MLVMS_P03355_PLV919





GSSGGG
12,553
MLVMS_P03355_3mutA_WS





EAAAKGGSGSS
12,554
PERV_Q4VFZ2_3mutA_WS





PAPGSSEAAAK
12,555
MLVMS_P03355_PLV919





GSSEAAAKPAP
12,556
MLVMS_P03355_PLV919





GGSPAPGSS
12,557
BAEVM_P10272_3mutA





GSAGSAAGSGEF
12,558
MLVCB_P08361_3mut





GGSPAPGGG
12,559
PERV_Q4VFZ2_3mut





GGGGSGGGGSGGGGSGGG
12,560
MLVMS_P03355_3mut


GS







GSSGSSGSS
12,561
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,562
PERV_Q4VFZ2_3mut


AKEAAAKEAAAK







GGGGSEAAAKGGGGS
12,563
MLVCB_P08361_3mutA





GGSEAAAKGSS
12,564
MLVAV_P03356_3mutA





EAAAKGGGGSEAAAK
12,565
MLVCB_P08361_3mut





EAAAKEAAAKEAAAKEAA
12,566
XMRV6_A1Z651_3mutA


AKEAAAKEAAAK







PAPGGGEAAAK
12,567
MLVMS_P03355_3mutA_WS





GSSGGGEAAAK
12,568
PERV_Q4VFZ2_3mutA_WS





GSSGSS
12,569
MLVCB_P08361_3mut





PAPAPAPAPAPAP
12,570
PERV_Q4VFZ2_3mut





GGSPAPGGG
12,571
MLVFF_P26809_3mutA





GGSGGSGGSGGSGGS
12,572
MLVCB_P08361_3mutA





EAAAKEAAAK
12,573
MLVFF_P26809_3mutA





AEAAAKEAAAKEAAAKEA
12,574
GALV_P21414_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPAPAPAPAPAP
12,575
WMSV_P03359_3mutA





GGGEAAAKGGS
12,576
KORV_Q9TTC1_3mutA





EAAAKGGGPAP
12,577
KORV_Q9TTC1_3mut





PAPEAAAKGSS
12,578
MLVBM_Q7SVK7_3mutA_WS





PAPEAAAKGSS
12,579
FLV_P10273_3mutA





PAPGGSEAAAK
12,580
MLVMS_P03355_3mut





GSSPAPGGG
12,581
BAEVM_P10272_3mutA





GGGEAAAKPAP
12,582
KORV_Q9TTC1-Pro_3mutA





GGGGSGGGGS
12,583
MLVMS_P03355_PLV919





GGGEAAAKGSS
12,584
MLVFF_P26809_3mutA





PAPGGGGSS
12,585
MLVBM_Q7SVK7_3mutA_WS





GSSEAAAK
12,586
BAEVM_P10272_3mutA





GGGGGGGG
12,587
MLVMS_P03355_PLV919





PAPGSSGGS
12,588
MLVAV_P03356_3mutA





GGGGGGGGSGGGGSGGGG
12,589
BAEVM_P10272_3mutA


S







PAP

MLVMS_P03355_3mut





EAAAKGSSPAP
12,591
XMRV6_A1Z651_3mutA





PAPEAAAKGGS
12,592
MLVFF_P26809_3mutA





GSSGGGEAAAK
12,593
BAEVM_P10272_3mutA





PAPAPAP
12,594
MLVMS_P03355_3mutA_WS





GGSEAAAKGGG
12,595
MLVMS_P03355_PLV919





GSSEAAAK
12,596
PERV_Q4VFZ2_3mut





GGGG
12,597
MLVMS_P03355_3mutA_WS





GGGGGS
12,598
MLVMS_P03355_3mut





GGGGSSEAAAK
12,599
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
12,600
SFV3L_P27401-Pro_2mutA


AKEAAAKEAAAK







GGSEAAAKGSS
12,601
MLVMS_P03355_3mutA_WS





PAPGSSGGS
12,602
XMRV6_A1Z651_3mutA





GGSPAP
12,603
MLVMS_P03355_3mutA_WS





GGGGSSEAAAK
12,604
BAEVM_P10272_3mut





GGSGGSGGSGGS
12,605
AVIRE_P03360_3mutA





PAPGSSGGS
12,606
MLVFF_P26809_3mutA





GSSPAPGGG
12,607
MLVMS_P03355_3mutA_WS





GGGGGGG
12,608
MLVMS_P03355_3mutA_WS





EAAAKGGGGGS
12,609
MLVMS_P03355_3mutA_WS





EAAAKGGSGGG
12,610
MLVMS_P03355_PLV919





GGGGSSEAAAK
12,611
XMRV6_A1Z651_3mutA





GGGGSEAAAKGGGGS
12,612
MLVBM_Q7SVK7_3mutA_WS





GSSGSS
12,613
MLVMS_P03355_PLV919





GGSGGG
12,614
MLVMS_P03355_PLV919





PAPEAAAKGGG
12,615
AVIRE_P03360_3mutA





AEAAAKEAAAKEAAAKEA
12,616
FOAMV_P14350-Pro_2mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGGSGSS
12,617
PERV_Q4VFZ2_3mut





GSSGSSGSSGSSGSS
12,618
KORV_Q9TTC1-Pro_3mut





GGGGSEAAAKGGGGS
12,619
MLVMS_P03355_3mutA_WS





GGGGGSPAP
12,620
FLV_P10273_3mut





GGGEAAAK
12,621
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGS
12,622
FLV_P10273_3mutA





GGG

MLVMS_P03355_PLV919





GGSPAPEAAAK
12,624
BAEVM_P10272_3mutA





EAAAKEAAAK
12,625
FLV_P10273_3mutA





GGGEAAAKPAP
12,626
BAEVM_P10272_3mutA





GGGEAAAKGGS
12,627
PERV_Q4VFZ2_3mut





GGSGGSGGS
12,628
PERV_Q4VFZ2_3mut





EAAAKGGGPAP
12,629
XMRV6_A1Z651_3mutA





EAAAK
12,630
MLVBM_Q7SVK7_3mutA_WS





PAPEAAAKGGG
12,631
PERV_Q4VFZ2_3mut





EAAAKGSS
12,632
MLVCB_P08361_3mutA





GGSEAAAKGGG
12,633
MLVBM_Q7SVK7_3mutA_WS





GGGGSGGGGSGGGGSGGG
12,634
XMRV6_A1Z651_3mutA


GS







GGGGSGGGGSGGGGSGGG
12,635
BAEVM_P10272_3mut


GSGGGGS







GGGGSSPAP
12,636
PERV_Q4VFZ2_3mutA_WS





GGSGGSGGSGGSGGSGGS
12,637
PERV_Q4VFZ2_3mut





GGGEAAAKPAP
12,638
PERV_Q4VFZ2_3mut





EAAAKEAAAK
12,639
BAEVM_P10272_3mutA





GGSGSSEAAAK
12,640
XMRV6_A1Z651_3mutA





PAPEAAAKGSS
12,641
WMSV_P03359_3mutA





PAPAPAPAPAP
12,642
XMRV6_A1Z651_3mutA





GSSGGGEAAAK
12,643
MLVMS_P03355_PLV919





GSSPAPGGG
12,644
MLVFF_P26809_3mutA





GGSPAPEAAAK
12,645
MLVFF_P26809_3mut





PAPGGSEAAAK
12,646
PERV_Q4VFZ2_3mut





GGGGSS
12,647
MLVFF_P26809_3mutA





GGSGSSGGG
12,648
BAEVM_P10272_3mutA





GSSGGGEAAAK
12,649
MLVMS_P03355_3mutA_WS





EAAAKGGS
12,650
MLVBM_Q7SVK7_3mutA_WS





GGGPAPGGS
12,651
MLVMS_P03355_PLV919





EAAAKEAAAK
12,652
MLVMS_P03355_PLV919





GSSGSSGSS
12,653
MLVMS_P03355_PLV919





GGGEAAAKPAP
12,654
MLVAV_P03356_3mutA





SGSETPGTSESATPES
12,655
FLV_P10273_3mutA





PAPAPAPAPAP
12,656
KORV_Q9TTC1-Pro_3mut





AEAAAKEAAAKEAAAKEA
12,657
BAEVM_P10272_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPGSSGGG
12,658
MLVMS_P03355_3mutA_WS





GSSGGGEAAAK
12,659
XMRV6_A1Z651_3mutA





GGGGSGGGGSGGGGSGGG
12,660
XMRV6_A1Z651_3mutA


GSGGGGS







GGGGSSPAP
12,661
MLVFF_P26809_3mutA





GGSGGGPAP
12,662
PERV_Q4VFZ2_3mutA_WS





GSS

PERV_Q4VFZ2_3mut





EAAAKGSSPAP
12,664
MLVMS_P03355_3mut





EAAAKGGG
12,665
XMRV6_A1Z651_3mutA





GSSGSSGSSGSS
12,666
WMSV_P03359_3mutA





PAPEAAAKGSS
12,667
MLVMS_P03355_PLV919





GSSEAAAK
12,668
AVIRE_P03360_3mutA





EAAAKGGSGSS
12,669
AVIRE_P03360_3mutA





GSSEAAAK
12,670
MLVMS_P03355_3mut





GGSGSSEAAAK
12,671
MLVMS_P03355_PLV919





GGSEAAAKGGG
12,672
MLVFF_P26809_3mutA





GGGGSGGGGSGGGGSGGG
12,673
MLVAV_P03356_3mutA


GS







PAPAPAPAPAPAP
12,674
MLVFF_P26809_3mut





EAAAKPAPGSS
12,675
KORV_Q9TTC1-Pro_3mut





PAPGSSEAAAK
12,676
MLVAV_P03356_3mutA





GGGGSSPAP
12,677
WMSV_P03359_3mutA





EAAAKGGGGGS
12,678
MLVMS_P03355_3mutA_WS





GGGEAAAKGGS
12,679
MLVMS_P03355_3mut





GGSGSSGGG
12,680
MLVMS_P03355_3mut





GGGPAPGGS
12,681
MLVAV_P03356_3mutA





PAPGGGGGS
12,682
MLVMS_P03355_PLV919





GGGPAPGSS
12,683
PERV_Q4VFZ2_3mut





GGGGGGG
12,684
MLVFF_P26809_3mutA





GGSGGGGSS
12,685
MLVCB_P08361_3mutA





GGGGGG
12,686
FLV_P10273_3mutA





GGSEAAAKGSS
12,687
PERV_Q4VFZ2_3mut





GGSPAPGGG
12,688
BAEVM_P10272_3mutA





GGSPAPGSS
12,689
AVIRE_P03360_3mutA





GGSGGSGGSGGS
12,690
KORV_Q9TTC1_3mut





EAAAKEAAAKEAAAKEAA
12,691
MLVBM_Q7SVK7_3mut


AKEAAAK







PAPGSSGGS
12,692
XMRV6_A1Z651_3mut





EAAAKGGGGSS
12,693
PERV_Q4VFZ2_3mutA_WS





GGSGGSGGSGGSGGS
12,694
PERV_Q4VFZ2_3mutA_WS





PAPGGSGGG
12,695
MLVMS_P03355_PLV919





PAPGSSGGG
12,696
PERV_Q4VFZ2_3mutA_WS





GSSGSS
12,697
BAEVM_P10272_3mutA





EAAAKGSS
12,698
MLVFF_P26809_3mutA





GGGPAP
12,699
MLVMS_P03355_PLV919





EAAAKGGGGGS
12,700
MLVFF_P26809_3mutA





EAAAKGGSPAP
12,701
MLVBM_Q7SVK7_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,702
WMSV_P03359_3mutA


AKEAAAKEAAAK







GSSPAPGGG
12,703
MLVBM_Q7SVK7_3mutA_WS





GGGEAAAKGSS
12,704
AVIRE_P03360_3mutA





GGGGSSEAAAK
12,705
AVIRE_P03360_3mutA





GGGGGGGG
12,706
PERV_Q4VFZ2_3mutA_WS





PAPGSSEAAAK
12,707
BAEVM_P10272_3mutA





EAAAKGSS
12,708
MLVFF_P26809_3mut





GSSEAAAKGGG
12,709
MLVCB_P08361_3mutA





GGSEAAAK
12,710
MLVBM_Q7SVK7_3mutA_WS





GSSEAAAKGGG
12,711
PERV_Q4VFZ2_3mutA_WS





PAPGGSGGG
12,712
WMSV_P03359_3mutA





GSSGGSGGG
12,713
MLVCB_P08361_3mutA





EAAAKGSSGGG
12,714
FLV_P10273_3mutA





GSSEAAAK
12,715
MLVCB_P08361_3mutA





GSSGGGEAAAK
12,716
MLVMS_P03355_3mut





GGGGSGGGGS
12,717
MLVCB_P08361_3mutA





EAAAKGGGGSEAAAK
12,718
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGG
12,719
PERV_Q4VFZ2_3mutA_WS





EAAAKGGSPAP
12,720
MLVMS_P03355_PLV919





GGGPAPGGS
12,721
AVIRE_P03360_3mutA





GSSEAAAK
12,722
MLVBM_Q7SVK7_3mutA_WS





GSSGGGEAAAK
12,723
PERV_Q4VFZ2_3mut





SGSETPGTSESATPES
12,724
MLVMS_P03355_PLV919





GGSGSSPAP
12,725
MLVMS_P03355_3mut





GGGGGG
12,726
MLVBM_Q7SVK7_3mutA_WS





GGSPAPGGG
12,727
XMRV6_A1Z651_3mutA





GGSGSS
12,728
PERV_Q4VFZ2_3mutA_WS





PAP

MLVBM_Q7SVK7_3mutA_WS





EAAAKPAPGSS
12,730
MLVMS_P03355_PLV919





EAAAKGGG
12,731
MLVMS_P03355_3mut





GSSEAAAKPAP
12,732
PERV_Q4VFZ2_3mutA_WS





GGGGSS
12,733
MLVMS_P03355_3mutA_WS





GGSGSSEAAAK
12,734
PERV_Q4VFZ2_3mut





GGGGSS
12,735
BAEVM_P10272_3mutA





PAPAP
12,736
MLVFF_P26809_3mut





PAPEAAAKGGG
12,737
BAEVM_P10272_3mutA





EAAAKGGS
12,738
MLVMS_P03355_PLV919





PAPAPAPAPAPAP
12,739
PERV_Q4VFZ2_3mutA_WS





GGGGGSEAAAK
12,740
MLVMS_P03355_3mut





PAPGGS
12,741
PERV_Q4VFZ2_3mut





GGGGSS
12,742
MLVCB_P08361_3mutA





GGGGS
12,743
MLVAV_P03356_3mutA





GSSPAPEAAAK
12,744
MLVMS_P03355_PLV919





GGGGSSGGS
12,745
MLVFF_P26809_3mutA





PAPEAAAKGSS
12,746
MLVMS_P03355_PLV919





GGSGSSEAAAK
12,747
MLVMS_P03355_3mutA_WS





EAAAKGGG
12,748
MLVAV_P03356_3mutA





PAPGSSEAAAK
12,749
FLV_P10273_3mutA





EAAAKGSSGGG
12,750
MLVCB_P08361_3mutA





PAPEAAAK
12,751
KORV_Q9TTC1-Pro_3mutA





GGSPAPEAAAK
12,752
KORV_Q9TTC1-Pro_3mut





GGSGGSGGSGGSGGSGGS
12,753
MLVAV_P03356_3mutA





GSSEAAAKPAP
12,754
MLVBM_Q7SVK7_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,755
KORV_Q9TTC1-Pro_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSGGGEAAAK
12,756
XMRV6_A1Z651_3mut





PAPGGSGGG
12,757
AVIRE_P03360_3mutA





PAPGGSEAAAK
12,758
PERV_Q4VFZ2_3mutA_WS





GGGGS
12,759
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGS
12,760
MLVBM_Q7SVK7_3mutA_WS





PAPAPAPAPAP
12,761
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAKEAA
12,762
MLVMS_P03355_3mut


AKEAAAK







GSSGGSEAAAK
12,763
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGS
12,764
WMSV_P03359_3mutA





EAAAKGSSGGG
12,765
WMSV_P03359_3mutA





EAAAKGGG
12,766
PERV_Q4VFZ2_3mutA_WS





SGSETPGTSESATPES
12,767
PERV_Q4VFZ2_3mut





PAPGSSGGS
12,768
MLVMS_P03355_3mutA_WS





PAPEAAAKGSS
12,769
PERV_Q4VFZ2_3mut





PAPEAAAK
12,770
AVIRE_P03360_3mutA





GSSEAAAKGGG
12,771
BAEVM_P10272_3mutA





GSSPAP
12,772
MLVAV_P03356_3mutA





EAAAKEAAAKEAAAKEAA
12,773
MLVFF_P26809_3mut


AK







PAPGGSGSS
12,774
MLVAV_P03356_3mutA





GGGGSGGGGSGGGGS
12,775
PERV_Q4VFZ2_3mutA_WS





GSSGGSEAAAK
12,776
MLVCB_P08361_3mutA





EAAAKGGS
12,777
KORV_Q9TTC1-Pro_3mutA





EAAAKGGS
12,778
MLVFF_P26809_3mutA





GGSPAP
12,779
MLVMS_P03355_PLV919





GGSGSS
12,780
MLVMS_P03355_PLV919





SGSETPGTSESATPES
12,781
WMSV_P03359_3mut





GGGGGGG
12,782
WMSV_P03359_3mut





GGSPAPGSS
12,783
MLVCB_P08361_3mutA





GGGGSSGGS
12,784
WMSV_P03359_3mut





PAPGGS
12,785
MLVMS_P03355_PLV919





PAPGSSGGS
12,786
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
12,787
MLVFF_P26809_3mut


AKEAAAK







SGGSSGGSSGSETPGTSE
12,788
PERV_Q4VFZ2_3mut


SATPESSGGSSGGSS







GGSGGSGGSGGSGGS
12,789
BAEVM_P10272_3mutA





GSSEAAAK
12,790
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
12,791
KORV_Q9TTC1-Pro_3mutA


AK







GGSGGSGGSGGSGGS
12,792
MLVMS_P03355_3mut





PAPAPAPAPAPAP
12,793
MLVMS_P03355_3mut





GGSPAPEAAAK
12,794
MLVMS_P03355_PLV919





EAAAK
12,795
WMSV_P03359_3mutA





EAAAKGSSGGS
12,796
MLVBM_Q7SVK7_3mutA_WS





GGSGGGGSS
12,797
MLVMS_P03355_3mutA_WS





GGGEAAAKPAP
12,798
MLVMS_P03355_3mut





EAAAKGGSGGG
12,799
XMRV6_A1Z651_3mutA





GGGGGSEAAAK
12,800
KORV_Q9TTC1-Pro_3mutA





GGGGGG
12,801
BAEVM_P10272_3mutA





GGGGGG
12,802
MLVMS_P03355_3mut





GGGGGGG
12,803
MLVBM_Q7SVK7_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,804
AVIRE_P03360


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPGSSGGS
12,805
PERV_Q4VFZ2_3mut





GGGGGS
12,806
XMRV6_A1Z651_3mut





EAAAKPAP
12,807
XMRV6_A1Z651_3mutA





GGG

MLVMS_P03355_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,809
FLV_P10273_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKGSSPAP
12,810
MLVMS_P03355_3mut





SGSETPGTSESATPES
12,811
BAEVM_P10272_3mutA





GGSPAPEAAAK
12,812
MLVMS_P03355_3mut





GSSGSSGSSGSS
12,813
MLVAV_P03356_3mutA





AEAAAKEAAAKEAAAKEA
12,814
MLVMS_P03355_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSPAP
12,815
MLVCB_P08361_3mutA





GGGGGSEAAAK
12,816
MLVMS_P03355_3mutA_WS





GGGGG
12,817
MLVFF_P26809_3mutA





GSSEAAAK
12,818
MLVAV_P03356_3mutA





GGS

BAEVM_P10272_3mut





EAAAKGGSPAP
12,820
MLVCB_P08361_3mutA





PAPAPAPAP
12,821
FLV_P10273_3mutA





PAPGGGEAAAK
12,822
MLVCB_P08361_3mutA





GGGGSSEAAAK
12,823
MLVMS_P03355_3mutA_WS





GGGGG
12,824
PERV_Q4VFZ2_3mutA_WS





GGSGGSGGSGGSGGSGGS
12,825
PERV_Q4VFZ2_3mut





GGGGG
12,826
MLVMS_P03355_3mut





PAPEAAAKGGG
12,827
MLVBM_Q7SVK7_3mutA_WS





GSSGGGPAP
12,828
XMRV6_A1Z651_3mutA





GSSGSSGSSGSSGSSGSS
12,829
PERV_Q4VFZ2_3mutA_WS





EAAAKGGSPAP
12,830
PERV_Q4VFZ2_3mut





GSSGGSEAAAK
12,831
MLVMS_P03355_PLV919





GSS

PERV_Q4VFZ2_3mut





EAAAKGGS
12,833
WMSV_P03359_3mutA





GGGGGSPAP
12,834
PERV_Q4VFZ2_3mutA_WS





EAAAKGSS
12,835
MLVMS_P03355_PLV919





EAAAKGGGGSS
12,836
KORV_Q9TTC1-Pro_3mutA





PAPGSSGGG
12,837
PERV_Q4VFZ2_3mut





GGGGSSEAAAK
12,838
MLVFF_P26809_3mut





PAPAPAP
12,839
MLVMS_P03355_3mut





GSSGGSEAAAK
12,840
XMRV6_A1Z651_3mut





PAPEAAAKGSS
12,841
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGSGGS
12,842
MLVMS_P03355_3mutA_WS





GGSGSSPAP
12,843
XMRV6_A1Z651_3mutA





GGGGSSPAP
12,844
MLVMS_P03355_PLV919





GGGGS
12,845
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
12,846
PERV_Q4VFZ2_3mutA_WS


AK







EAAAKEAAAK
12,847
KORV_Q9TTC1_3mutA





PAPGGGEAAAK
12,848
BAEVM_P10272_3mutA





GSSGGSEAAAK
12,849
XMRV6_A1Z651_3mutA





EAAAKEAAAKEAAAKEAA
12,850
FLV_P10273_3mut


AKEAAAKEAAAK







GSSEAAAKPAP
12,851
MLVMS_P03355_3mutA_WS





EAAAKPAPGSS
12,852
PERV_Q4VFZ2_3mutA_WS





GSSGGSPAP
12,853
XMRV6_A1Z651_3mutA





GSSEAAAKGGG
12,854
PERV_Q4VFZ2_3mut





GGGEAAAKGGS
12,855
WMSV_P03359_3mutA





GSSEAAAKGGG
12,856
MLVFF_P26809_3mut





PAPAPAP
12,857
KORV_Q9TTC1-Pro_3mutA





EAAAKGGSPAP
12,858
MLVMS_P03355_3mutA_WS





PAPGGSEAAAK
12,859
PERV_Q4VFZ2_3mut





GGGGS
12,860
MLVBM_Q7SVK7_3mutA_WS





EAAAKGSSGGG
12,861
KORV_Q9TTC1_3mut





EAAAKGGGPAP
12,862
MLVCB_P08361_3mutA





EAAAKGSS
12,863
BAEVM_P10272_3mutA





GGSPAPGGG
12,864
MLVBM_Q7SVK7_3mutA_WS





GGGGSEAAAKGGGGS
12,865
MLVMS_P03355_3mutA_WS





GGGEAAAKGGS
12,866
PERV_Q4VFZ2_3mutA_WS





EAAAKGGGGSS
12,867
MLVMS_P03355_3mutA_WS





EAAAKGGGPAP
12,868
MLVFF_P26809_3mut





GSSPAP
12,869
PERV_Q4VFZ2_3mutA_WS





EAAAKGGS
12,870
MLVMS_P03355_3mut





GGGGSS
12,871
KORV_Q9TTC1-Pro_3mutA





EAAAKGSSPAP
12,872
MLVMS_P03355_3mutA_WS





GGGPAP
12,873
PERV_Q4VFZ2_3mut





EAAAKGSSGGS
12,874
XMRV6_A1Z651_3mutA





PAPGGG
12,875
MLVAV_P03356_3mutA





GSSPAPEAAAK
12,876
BAEVM_P10272_3mutA





GGGPAP
12,877
MLVBM_Q7SVK7_3mutA_WS





GSSGGGGGS
12,878
AVIRE_P03360_3mutA





SGSETPGTSESATPES
12,879
MLVMS_P03355_PLV919





GGGPAP
12,880
MLVFF_P26809_3mut





EAAAKGGGGSS
12,881
XMRV6_A1Z651_3mutA





GGGGSSPAP
12,882
XMRV6_A1Z651_3mut





GGGGSEAAAKGGGGS
12,883
MLVMS_P03355_3mut





GSSPAP
12,884
MLVBM_Q7SVK7_3mutA_WS





GGSGSSEAAAK
12,885
FLV_P10273_3mutA





SGSETPGTSESATPES
12,886
MLVBM_Q7SVK7_3mutA_WS





PAPGGG
12,887
AVIRE_P03360_3mutA





GGGEAAAKPAP
12,888
MLVMS_P03355_3mutA_WS





EAAAKGGSGSS
12,889
PERV_Q4VFZ2_3mut





GGSPAPGGG
12,890
MLVAV_P03356_3mutA





PAPGGSGSS
12,891
BAEVM_P10272_3mutA





GSSGGSPAP
12,892
MLVFF_P26809_3mutA





EAAAKGSSGGG
12,893
PERV_Q4VFZ2_3mut





GGGGSGGGGS
12,894
PERV_Q4VFZ2_3mutA_WS





GSSGGGGGS
12,895
BAEVM_P10272_3mutA





GGGGSSGGS
12,896
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGS
12,897
PERV_Q4VFZ2_3mutA_WS





GSSGSSGSSGSS
12,898
MLVMS_P03355_3mut





GGS

MLVMS_P03355_3mutA_WS





GSSGGSEAAAK
12,900
MLVBM_Q7SVK7_3mutA_WS





SGGSSGGSSGSETPGTSE
12,901
XMRV6_A1Z651


SATPESSGGSSGGSS







GGGGG
12,902
FLV_P10273_3mutA





PAPEAAAKGSS
12,903
PERV_Q4VFZ2_3mut





GGGGGG
12,904
WMSV_P03359_3mut





EAAAKGGG
12,905
BAEVM_P10272_3mutA





GGGGSS
12,906
MLVMS_P03355_3mutA_WS





GSSGGGEAAAK
12,907
KORV_Q9TTC1_3mut





GGSGSS
12,908
AVIRE_P03360_3mutA





EAAAKPAP
12,909
MLVMS_P03355_3mut





EAAAKEAAAKEAAAK
12,910
FLV_P10273_3mutA





GGGG
12,911
XMRV6_A1Z651_3mutA





GSSPAPGGS
12,912
BAEVM_P10272_3mutA





GSSGGGGGS
12,913
MLVFF_P26809_3mutA





GGGGSSGGS
12,914
MLVAV_P03356_3mutA





GGS

PERV_Q4VFZ2_3mut





GGGGG
12,916
WMSV_P03359_3mutA





GSSGSSGSSGSSGSSGSS
12,917
FLV_P10273_3mutA





PAPGGGGSS
12,918
MLVAV_P03356_3mutA





GGGGGGGG
12,919
BAEVM_P10272_3mutA





SGSETPGTSESATPES
12,920
MLVCB_P08361_3mutA





PAPGGG
12,921
BAEVM_P10272_3mutA





GSSGSSGSS
12,922
MLVCB_P08361_3mutA





GGSGSS
12,923
MLVMS_P03355_3mutA_WS





EAAAKGGGGSEAAAK
12,924
WMSV_P03359_3mutA





GGGGGGGG
12,925
FLV_P10273_3mutA





GSSGSS
12,926
MLVMS_P03355_3mutA_WS





PAPEAAAKGGS
12,927
XMRV6_A1Z651_3mutA





EAAAKEAAAK
12,928
MLVMS_P03355_3mut





GGGGSGGGGSGGGGS
12,929
BAEVM_P10272_3mutA





EAAAKGSSPAP
12,930
MLVMS_P03355_PLV919





GGGGSSEAAAK
12,931
MLVMS_P03355_3mut





GGGGSSEAAAK
12,932
BAEVM_P10272_3mutA





PAPGGSGSS
12,933
PERV_Q4VFZ2_3mut





GGSGGGEAAAK
12,934
MLVFF_P26809_3mut





PAPEAAAKGGS
12,935
PERV_Q4VFZ2_3mut





GGGPAPGSS
12,936
AVIRE_P03360_3mut





PAPGGSGGG
12,937
PERV_Q4VFZ2_3mutA_WS





GGGGGGGG
12,938
PERV_Q4VFZ2_3mutA_WS





GSSEAAAK
12,939
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGS
12,940
PERV_Q4VFZ2_3mutA_WS





EAAAKGGS
12,941
MLVMS_P03355_3mut





GGGGGSGSS
12,942
MLVCB_P08361_3mut





GGGPAP
12,943
KORV_Q9TTC1-Pro_3mutA





EAAAKPAPGGG
12,944
MLVCB_P08361_3mut





GSSGGSPAP
12,945
MLVCB_P08361_3mutA





SGGSSGGSSGSETPGTSE
12,946
MLVMS_P03355_3mut


SATPESSGGSSGGSS







PAPAPAPAP
12,947
MLVMS_P03355_3mut





GSSGGS
12,948
XMRV6_A1Z651_3mutA





GSSEAAAKGGG
12,949
MLVMS_P03355_3mut





GGSGSSPAP
12,950
MLVMS_P03355_3mutA_WS





GSSEAAAKGGS
12,951
MLVMS_P03355_PLV919





EAAAKEAAAKEAAAKEAA
12,952
BAEVM_P10272_3mut


AKEAAAK







PAPGGGGSS
12,953
KORV_Q9TTC1_3mutA





EAAAKGSS
12,954
MLVMS_P03355_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,955
FFV_O93209_2mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSGGSGGSGGSGGSGGS
12,956
BAEVM_P10272_3mutA





GGGGGG
12,957
MLVMS_P03355_PLV919





PAPEAAAK
12,958
BAEVM_P10272_3mutA





GGSGSSEAAAK
12,959
MLVAV_P03356_3mutA





GGG

MLVCB_P08361_3mutA





GGGGG
12,961
MLVCB_P08361_3mutA





GGSGGSGGSGGS
12,962
KORV_Q9TTC1-Pro_3mutA





GSSGSSGSSGSSGSSGSS
12,963
XMRV6_A1Z651_3mutA





GSSEAAAKPAP
12,964
FLV_P10273_3mutA





GGGEAAAKPAP
12,965
MLVCB_P08361_3mutA





GSSGSSGSS
12,966
MLVMS_P03355_3mutA_WS





PAPAPAPAP
12,967
MLVMS_P03355_PLV919





EAAAKGGG
12,968
MLVMS_P03355_PLV919





PAPAPAPAPAPAP
12,969
FLV_P10273_3mutA





EAAAKGGSGSS
12,970
MLVMS_P03355_3mut





GGGGGG
12,971
PERV_Q4VFZ2_3mutA_WS





PAPGGG
12,972
MLVCB_P08361_3mutA





GGGGGSGSS
12,973
KORV_Q9TTC1_3mutA





GGGGSGGGGGGGGSGGGG
12,974
XMRV6_A1Z651_3mut


S







GGSGGSGGS
12,975
KORV_Q9TTC1-Pro_3mutA





EAAAKPAPGGG
12,976
MLVMS_P03355_3mutA_WS





AEAAAKEAAAKEAAAKEA
12,977
XMRV6_A1Z651


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGSGGGGSGGGGSGGG
12,978
FLV_P10273_3mutA


GSGGGGSGGGGS







EAAAKGGGGSEAAAK
12,979
PERV_Q4VFZ2_3mutA_WS





GGGPAPGSS
12,980
AVIRE_P03360_3mutA





GGGGG
12,981
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGSGGG
12,982
MLVMS_P03355_3mut


GSGGGGSGGGGS







GGGGSGGGGS
12,983
MLVMS_P03355_3mutA_WS





EAAAKGGSPAP
12,984
XMRV6_A1Z651_3mutA





EAAAKGSSPAP
12,985
AVIRE_P03360_3mutA





PAPGGSGSS
12,986
KORV_Q9TTC1-Pro_3mutA





GSS

MLVBM_Q7SVK7_3mutA_WS





GSS

WMSV_P03359_3mut





GGGPAPGSS
12,989
MLVFF_P26809_3mutA





EAAAKPAP
12,990
MLVMS_P03355_3mut





GSSPAPEAAAK
12,991
FLV_P10273_3mutA





GGSPAPGSS
12,992
MLVBM_Q7SVK7_3mutA_WS





GGGGGSEAAAK
12,993
XMRV6_A1Z651_3mut





PAPEAAAKGGG
12,994
WMSV_P03359_3mutA





PAPGGG
12,995
PERV_Q4VFZ2_3mut





GGSPAPEAAAK
12,996
WMSV_P03359_3mutA





GGSGGGGSS
12,997
PERV_Q4VFZ2_3mut





EAAAKGGGGSS
12,998
PERV_Q4VFZ2_3mut





EAAAKGGSPAP
12,999
AVIRE_P03360_3mut





GGSGGGGSS
13,000
WMSV_P03359_3mutA





PAPGSSEAAAK
13,001
MLVFF_P26809_3mut





GSSEAAAK
13,002
MLVMS_P03355_PLV919





GSAGSAAGSGEF
13,003
AVIRE_P03360_3mutA





EAAAKGGSGSS
13,004
MLVMS_P03355_3mut





GGSEAAAKPAP
13,005
MLVMS_P03355_PLV919





GGGGSGGGGSGGGGSGGG
13,006
MLVFF_P26809_3mutA


GSGGGGS







PAPGSSEAAAK
13,007
PERV_Q4VFZ2_3mutA_WS





GGGGSSPAP
13,008
MLVMS_P03355_3mutA_WS





PAPAPAP
13,009
MLVCB_P08361_3mutA





EAAAKPAPGGG
13,010
MLVBM_Q7SVK7_3mutA_WS





GGGPAPGSS
13,011
BAEVM_P10272_3mutA





PAP

MLVMS_P03355_3mutA_WS





PAPGGSGGG
13,013
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGSGGS
13,014
MLVBM_Q7SVK7_3mutA_WS





PAPAPAPAP
13,015
XMRV6_A1Z651_3mut





GSSPAPGGG
13,016
MLVMS_P03355_3mutA_WS





GSSPAPGGG
13,017
MLVMS_P03355_3mut





PAPGGG
13,018
MLVMS_P03355_PLV919





GGGEAAAKGSS
13,019
WMSV_P03359_3mut





EAAAKGSS
13,020
KORV_Q9TTC1-Pro_3mutA





EAAAKGGS
13,021
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
13,022
PERV_Q4VFZ2_3mut


AKEAAAK







PAPEAAAKGGG
13,023
MLVMS_P03355_PLV919





EAAAKGSSGGG
13,024
MLVFF_P26809_3mut





AEAAAKEAAAKEAAAKEA
13,025
PERV_Q4VFZ2


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKEAAAKEAAAKEAA
13,026
MLVAV_P03356_3mutA


AKEAAAKEAAAK







GSSGGSGGG
13,027
MLVFF_P26809_3mut





GSSGSSGSSGSS
13,028
PERV_Q4VFZ2_3mutA_WS





GGSPAPGGG
13,029
MLVMS_P03355_PLV919





GSS

BAEVM_P10272_3mut





GGGPAPGSS
13,031
MLVMS_P03355_3mutA_WS





GGGGSS
13,032
KORV_Q9TTC1_3mutA





GSSGGSGGG
13,033
BAEVM_P10272_3mutA





EAAAKEAAAKEAAAK
13,034
MLVCB_P08361_3mutA





SGGSSGGSSGSETPGTSE
13,035
FLV_P10273_3mutA


SATPESSGGSSGGSS







PAPGGGGGS
13,036
PERV_Q4VFZ2_3mut





PAPAPAPAPAP
13,037
KORV_Q9TTC1-Pro_3mutA





EAAAK
13,038
MLVMS_P03355_3mutA_WS





GGG

MLVCB_P08361_3mut





GGSEAAAKGGG
13,040
BAEVM_P10272_3mutA





GGGGGSGSS
13,041
MLVAV_P03356_3mutA





EAAAKGSSPAP
13,042
MLVBM_Q7SVK7_3mutA_WS





GGSGGSGGS
13,043
XMRV6_A1Z651_3mut





EAAAKPAPGGG
13,044
KORV_Q9TTC1-Pro_3mutA





GGGPAPEAAAK
13,045
FLV_P10273_3mutA





GGSPAPEAAAK
13,046
MLVMS_P03355_3mutA_WS





GGSGGSGGSGGSGGS
13,047
MLVFF_P26809_3mut





EAAAKGGSGSS
13,048
MLVMS_P03355_PLV919





GGGEAAAKGGS
13,049
MLVBM_Q7SVK7_3mutA_WS





PAPAPAPAP
13,050
BAEVM_P10272_3mutA





EAAAKEAAAKEAAAKEAA
13,051
MLVMS_P03355_3mut


AK







EAAAKPAP
13,052
XMRV6_A1Z651_3mut





EAAAKEAAAK
13,053
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGG
13,054
BAEVM_P10272_3mut





EAAAKGSS
13,055
MLVAV_P03356_3mutA





EAAAKEAAAKEAAAKEAA
13,056
MLVFF_P26809_3mut


AKEAAAKEAAAK







GGGPAPGSS
13,057
PERV_Q4VFZ2_3mutA_WS





GGGG
13,058
PERV_Q4VFZ2_3mut





EAAAKGGSGSS
13,059
MLVMS_P03355_PLV919





GGGGSGGGGSGGGGS
13,060
MLVMS_P03355_3mutA_WS





EAAAK
13,061
MLVMS_P03355_3mutA_WS





GGGGSS
13,062
PERV_Q4VFZ2





PAPEAAAKGGS
13,063
MLVCB_P08361_3mut





GSS

MLVMS_P03355_3mut





GSAGSAAGSGEF
13,065
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
13,066
KORV_Q9TTC1-Pro_3mut


AKEAAAKEAAAK







GGGGSGGGGS
13,067
AVIRE_P03360_3mutA





EAAAK
13,068
MLVMS_P03355_3mut





GGGPAPGGS
13,069
PERV_Q4VFZ2_3mut





GGGGSGGGGSGGGGS
13,070
MLVMS_P03355_PLV919





PAPGGG
13,071
MLVMS_P03355_3mutA_WS





GGGEAAAKPAP
13,072
PERV_Q4VFZ2_3mutA_WS





EAAAKPAPGSS
13,073
KORV_Q9TTC1-Pro_3mutA





PAPGSS
13,074
KORV_Q9TTC1_3mutA





GSAGSAAGSGEF
13,075
PERV_Q4VFZ2_3mut





PAPGGGGSS
13,076
KORV_Q9TTC1-Pro_3mutA





GSSGGGEAAAK
13,077
MLVCB_P08361_3mutA





GSS

AVIRE_P03360_3mutA





GSSGSSGSSGSS
13,079
XMRV6_A1Z651_3mutA





PAPEAAAKGGG
13,080
MLVMS_P03355_PLV919





GGGPAPEAAAK
13,081
MLVCB_P08361_3mutA





PAPGGGGGS
13,082
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
13,083
PERV_Q4VFZ2_3mutA_WS


AK







GGGGGSPAP
13,084
MLVFF_P26809_3mutA





GSSGSSGSSGSSGSS
13,085
PERV_Q4VFZ2





GSSPAPEAAAK
13,086
MLVMS_P03355_PLV919





GSSGSSGSSGSSGSSGSS
13,087
MLVBM_Q7SVK7_3mutA_WS





GSSGSSGSSGSSGSSGSS
13,088
MLVMS_P03355_3mutA_WS





GGSPAPEAAAK
13,089
MLVAV_P03356_3mutA





GSSGGG
13,090
BAEVM_P10272_3mut





EAAAKGSSGGS
13,091
KORV_Q9TTC1-Pro_3mutA





GGSGSSEAAAK
13,092
MLVMS_P03355_3mutA_WS





GGGPAPEAAAK
13,093
MLVFF_P26809_3mutA





GGGPAPGGS
13,094
MLVMS_P03355_3mutA_WS





GGGGG
13,095
MLVMS_P03355_PLV919





GGGEAAAKPAP
13,096
MLVBM_Q7SVK7_3mutA_WS





GGGGSGGGGS
13,097
WMSV_P03359_3mut





GGGPAPEAAAK
13,098
PERV_Q4VFZ2_3mut





GGSGSSEAAAK
13,099
MLVMS_P03355_PLV919





EAAAKGGGPAP
13,100
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSSGSS
13,101
KORV_Q9TTC1-Pro_3mutA





PAPAP
13,102
WMSV_P03359_3mutA





GGSPAPGSS
13,103
MLVAV_P03356_3mutA





GGSGGGPAP
13,104
MLVMS_P03355_3mut





GGSPAP
13,105
MLVMS_P03355_PLV919





EAAAKGGSPAP
13,106
PERV_Q4VFZ2_3mut





GSSPAPGGG
13,107
KORV_Q9TTC1-Pro_3mutA





GSAGSAAGSGEF
13,108
MLVMS_P03355_3mut





GGSPAP
13,109
PERV_Q4VFZ2_3mut





GSSGSS
13,110
KORV_Q9TTC1-Pro_3mut





GGGPAPGSS
13,111
MLVMS_P03355_3mutA_WS





AEAAAKEAAAKEAAAKEA
13,112
FOAMV_P14350


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPGSSGGG
13,113
MLVMS_P03355_PLV919





GGSEAAAKPAP
13,114
BAEVM_P10272_3mutA





GGGGGS
13,115
MLVCB_P08361_3mutA





PAPEAAAKGGS
13,116
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
13,117
BAEVM_P10272_3mutA


AKEAAAKEAAAK







GGSEAAAK
13,118
BAEVM_P10272_3mutA





GSSPAPEAAAK
13,119
MLVMS_P03355_3mutA_WS





PAPGGG
13,120
WMSV_P03359_3mut





EAAAKPAP
13,121
PERV_Q4VFZ2_3mut





GSSGSSGSSGSSGSS
13,122
WMSV_P03359_3mut





PAPGGG
13,123
MLVBM_Q7SVK7_3mutA_WS





GGSGGGEAAAK
13,124
BAEVM_P10272_3mutA





PAPGGS
13,125
MLVMS_P03355_3mut





GGSGGSGGSGGS
13,126
MLVBM_Q7SVK7_3mutA_WS





EAAAKEAAAKEAAAKEAA
13,127
PERV_Q4VFZ2_3mut


AK







GGSEAAAKGGG
13,128
WMSV_P03359_3mutA





GGGPAP
13,129
BAEVM_P10272_3mutA





GGGGSGGGGGGGGSGGGG
13,130
XMRV6_A1Z651_3mut


SGGGGSGGGGS







GGSPAPGSS
13,131
KORV_Q9TTC1_3mut





GGGPAPGSS
13,132
MLVMS_P03355_3mut





GGGGSSGGS
13,133
BAEVM_P10272_3mutA





GGGEAAAKGSS
13,134
KORV_Q9TTC1-Pro_3mutA





PAPAP
13,135
MLVBM_Q7SVK7_3mutA_WS





GGSPAPGGG
13,136
PERV_Q4VFZ2_3mut





PAPGSS
13,137
PERV_Q4VFZ2_3mutA_WS





GSSGGSPAP
13,138
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGGGSEAAAK
13,139
PERV_Q4VFZ2_3mut





GSSEAAAKGGS
13,140
KORV_Q9TTC1-Pro_3mut





PAPAPAPAP
13,141
KORV_Q9TTC1-Pro_3mutA





GGSEAAAKPAP
13,142
WMSV_P03359_3mutA





PAPGGS
13,143
FLV_P10273_3mutA





EAAAKGGGPAP
13,144
PERV_Q4VFZ2_3mut





GGSGSSGGG
13,145
AVIRE_P03360_3mutA





EAAAKGGSGSS
13,146
BAEVM_P10272_3mutA





SGGSSGGSSGSETPGTSE
13,147
MLVCB_P08361_3mutA


SATPESSGGSSGGSS







GSSEAAAKGGS
13,148
XMRV6_A1Z651_3mutA





GGGGG
13,149
BAEVM_P10272_3mutA





GGGGSGGGGSGGGGSGGG
13,150
SFV3L_P27401_2mutA


GSGGGGSGGGGS







GGGEAAAKGSS
13,151
MLVMS_P03355_PLV919





EAAAKGGGGSEAAAK
13,152
KORV_Q9TTC1_3mutA





EAAAKGGG
13,153
AVIRE_P03360_3mut





GGSGGG
13,154
MLVMS_P03355_3mutA_WS





GGSGSSGGG
13,155
MLVMS_P03355_PLV919





GGGGSGGGGSGGGGSGGG
13,156
KORV_Q9TTC1_3mut


GSGGGGSGGGGS







GGGGSEAAAKGGGGS
13,157
KORV_Q9TTC1_3mutA





PAPAPAPAPAP
13,158
FLV_P10273_3mutA





GGS

MLVBM_Q7SVK7_3mutA_WS





GGGGGSEAAAK
13,160
MLVBM_Q7SVK7_3mutA_WS





GSSGSSGSSGSSGSS
13,161
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
13,162
MLVMS_P03355_3mut


AKEAAAK







GGSGSSGGG
13,163
PERV_Q4VFZ2_3mut





PAP

MLVFF_P26809_3mut





GSSPAPEAAAK
13,165
MLVAV_P03356_3mutA





EAAAKGGGGSS
13,166
MLVMS_P03355_3mut





GGGEAAAKGGS
13,167
XMRV6_A1Z651_3mut





GGSGGGPAP
13,168
MLVBM_Q7SVK7_3mutA_WS





GSAGSAAGSGEF
13,169
BAEVM_P10272_3mutA





GSSEAAAK
13,170
MLVCB_P08361_3mut





PAPGSS
13,171
MLVMS_P03355_3mut





EAAAKEAAAKEAAAK
13,172
MLVAV_P03356_3mutA





GSAGSAAGSGEF
13,173
XMRV6_A1Z651_3mutA





GSSGSSGSSGSS
13,174
BAEVM_P10272_3mutA





AEAAAKEAAAKEAAAKEA
13,175
KORV_Q9TTC1-Pro_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGSSEAAAK
13,176
WMSV_P03359_3mut





GSSGGGEAAAK
13,177
MLVBM_Q7SVK7_3mutA_WS





EAAAKPAP
13,178
MLVFF_P26809_3mutA





GGSPAPGGG
13,179
KORV_Q9TTC1_3mutA





PAPEAAAK
13,180
FLV_P10273_3mutA





GSSGSSGSS
13,181
MLVBM_Q7SVK7_3mutA_WS





GSSGGGEAAAK
13,182
FLV_P10273_3mutA





GGSPAP
13,183
MLVBM_Q7SVK7_3mutA_WS





GSAGSAAGSGEF
13,184
KORV_Q9TTC1-Pro_3mutA





PAPGGSEAAAK
13,185
MLVMS_P03355_PLV919





GGSPAPEAAAK
13,186
MLVBM_Q7SVK7_3mutA_WS





GGGGGSPAP
13,187
MLVBM_Q7SVK7_3mutA_WS





EAAAKGSSPAP
13,188
WMSV_P03359_3mut





EAAAKGGGPAP
13,189
MLVBM_Q7SVK7_3mutA_WS





PAPGSS
13,190
KORV_Q9TTC1-Pro_3mutA





GGSGSSGGG
13,191
BAEVM_P10272_3mut





SGGSSGGSSGSETPGTSE
13,192
FFV_O93209-Pro_2mut


SATPESSGGSSGGSS







GGSGGSGGSGGSGGSGGS
13,193
WMSV_P03359_3mutA





GGSGGSGGS
13,194
PERV_Q4VFZ2_3mutA_WS





GGGGG
13,195
PERV_Q4VFZ2_3mutA_WS





GGGPAP
13,196
FLV_P10273_3mutA





PAPGGSGGG
13,197
XMRV6_A1Z651_3mutA





GGGGSEAAAKGGGGS
13,198
XMRV6_A1Z651_3mut





EAAAKGSSGGG
13,199
KORV_Q9TTC1-Pro_3mutA





GSSGGSEAAAK
13,200
WMSV_P03359_3mut





EAAAKGGSGSS
13,201
PERV_Q4VFZ2_3mut





PAPAPAPAPAP
13,202
PERV_Q4VFZ2_3mut





GGGGSGGGGSGGGGSGGG
13,203
MLVMS_P03355_3mutA_WS


GGGGGSGGGGS







GGGGGGG
13,204
KORV_Q9TTC1_3mutA





EAAAK
13,205
KORV_Q9TTC1-Pro_3mutA





GGGEAAAKGGS
13,206
KORV_Q9TTC1-Pro_3mutA





GGGEAAAKGGS
13,207
PERV_Q4VFZ2_3mutA_WS





GGGGGSPAP
13,208
XMRV6_A1Z651_3mut





GGGGSGGGGSGGGGSGGG
13,209
MLVFF_P26809_3mut


GS







GGGGGGG
13,210
MLVFF_P26809_3mut





PAPAPAPAPAPAP
13,211
AVIRE_P03360_3mutA





GSSPAPGGG
13,212
FLV_P10273_3mutA





GGGGGSPAP
13,213
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGS
13,214
MLVMS_P03355_3mut





GGGGSGGGGSGGGGS
13,215
KORV_Q9TTC1_3mut





GSSEAAAKGGS
13,216
MLVAV_P03356_3mutA





GSSGSSGSSGSSGSS
13,217
MLVMS_P03355_3mut





EAAAKGGGGGS
13,218
PERV_Q4VFZ2_3mutA_WS





GSSGGGGGS
13,219
PERV_Q4VFZ2_3mut





GGGEAAAKPAP
13,220
MLVMS_P03355_3mut





GSSGGSPAP
13,221
PERV_Q4VFZ2_3mutA_WS





GSSGGGPAP
13,222
BAEVM_P10272_3mutA





GGGGGSGSS
13,223
MLVMS_P03355_PLV919





AEAAAKEAAAKEAAAKEA
13,224
BAEVM_P10272_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPEAAAK
13,225
MLVMS_P03355_3mut





GGGGSGGGGSGGGGS
13,226
FLV_P10273_3mutA





GGSGSSGGG
13,227
WMSV_P03359_3mutA





EAAAKGGS
13,228
PERV_Q4VFZ2_3mut





EAAAKGSSPAP
13,229
MLVCB_P08361_3mut





EAAAKGGSGSS
13,230
WMSV_P03359_3mutA





GSSGSS
13,231
PERV_Q4VFZ2_3mutA_WS





PAPAPAPAP
13,232
MLVMS_P03355_PLV919





GGSGGG
13,233
PERV_Q4VFZ2_3mutA_WS





GSS

MLVBM_Q7SVK7_3mutA_WS





PAP

KORV_Q9TTC1-Pro_3mutA





GGSGSSEAAAK
13,236
MLVFF_P26809_3mut





PAPEAAAKGSS
13,237
KORV_Q9TTC1-Pro_3mutA





GGSGGS
13,238
MLVCB_P08361_3mutA





GGGGGGG
13,239
PERV_Q4VFZ2_3mutA_WS





GGSPAPEAAAK
13,240
MLVBM_Q7SVK7_3mut





EAAAKEAAAKEAAAKEAA
13,241
KORV_Q9TTC1_3mutA


AKEAAAKEAAAK







GGSPAP
13,242
MLVMS_P03355_3mut





GGSEAAAKGGG
13,243
PERV_Q4VFZ2_3mut





GGGGSGGGGS
13,244
FLV_P10273_3mutA





GGGEAAAK
13,245
BAEVM_P10272_3mutA





GGGGSGGGGSGGGGSGGG
13,246
SFV3L_P27401_2mut


GGGGGSGGGGS







GGSEAAAKPAP
13,247
KORV_Q9TTC1-Pro_3mutA





GSSGGGEAAAK
13,248
MLVMS_P03355_PLV919





GGGGGSEAAAK
13,249
MLVMS_P03355_PLV919





EAAAKGGSGGG
13,250
MLVMS_P03355_3mutA_WS





GGGGSSPAP
13,251
MLVAV_P03356_3mutA





EAAAKEAAAK
13,252
MLVMS_P03355_3mutA_WS





AEAAAKEAAAKEAAAKEA
13,253
SFV3L_P27401_2mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSGSSGSSGSSGSS
13,254
MLVMS_P03355_PLV919





GSSGGG
13,255
KORV_Q9TTC1-Pro_3mutA





GSSGGS
13,256
MLVFF_P26809_3mutA





GGGGSGGGGS
13,257
XMRV6_A1Z651_3mutA





PAPGSS
13,258
MLVBM_Q7SVK7_3mutA_WS





GGGPAPEAAAK
13,259
XMRV6_A1Z651_3mutA





EAAAKGGS
13,260
MLVFF_P26809_3mut





GSS

KORV_Q9TTC1_3mutA





GGGG
13,262
PERV_Q4VFZ2_3mut





GGGGGSEAAAK
13,263
AVIRE_P03360_3mutA





GSSGSSGSSGSSGSS
13,264
MLVMS_P03355_PLV919





PAPGGSGGG
13,265
PERV_Q4VFZ2_3mut





GGGPAP
13,266
PERV_Q4VFZ2_3mut





GGGPAPEAAAK
13,267
AVIRE_P03360_3mutA





GGGEAAAK
13,268
MLVCB_P08361_3mut





GGG

MLVFF_P26809_3mutA





EAAAKPAPGSS
13,270
XMRV6_A1Z651_3mutA





GGSGSSEAAAK
13,271
PERV_Q4VFZ2_3mutA_WS





EAAAKGSS
13,272
MLVMS_P03355_3mut





GGSGSSEAAAK
13,273
BAEVM_P10272_3mut





GGSGGG
13,274
MLVBM_Q7SVK7_3mutA_WS





GGGPAP
13,275
MLVMS_P03355_PLV919





GGSPAPGGG
13,276
PERV_Q4VFZ2_3mutA_WS





GGGGGSEAAAK
13,277
MLVFF_P26809_3mutA





EAAAKGSSGGS
13,278
MLVBM_Q7SVK7_3mut





PAPAP
13,279
XMRV6_A1Z651_3mut





GSSPAPGGS
13,280
MLVBM_Q7SVK7_3mutA_WS





GSSEAAAKGGG
13,281
WMSV_P03359_3mutA





EAAAKGGGGGS
13,282
PERV_Q4VFZ2_3mut





GSSGSSGSSGSSGSS
13,283
MLVCB_P08361_3mutA





EAAAKGGGGSS
13,284
PERV_Q4VFZ2_3mut





EAAAKGSS
13,285
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
13,286
AVIRE_P03360_3mutA


AKEAAAKEAAAK







EAAAKGGS
13,287
MLVCB_P08361_3mut





GSSGGSEAAAK
13,288
MLVAV_P03356_3mutA





EAAAKPAPGGS
13,289
PERV_Q4VFZ2_3mut





GGSGGS
13,290
MLVAV_P03356_3mutA





EAAAKGSSGGG
13,291
AVIRE_P03360_3mutA





GGSGGSGGSGGS
13,292
PERV_Q4VFZ2_3mut





GGGGGGGG
13,293
KORV_Q9TTC1_3mutA





GGSGSSEAAAK
13,294
MLVCB_P08361_3mutA





EAAAKGGG
13,295
MLVBM_Q7SVK7_3mutA_WS





GGGGGGGGSGGGGS
13,296
MLVCB_P08361_3mut





GGSGGSGGSGGS
13,297
PERV_Q4VFZ2_3mutA_WS





PAPAPAPAPAP
13,298
WMSV_P03359_3mut





EAAAKEAAAKEAAAKEAA
13,299
PERV_Q4VFZ2_3mut


AK







GGSGGSGGS
13,300
XMRV6_A1Z651_3mutA





PAPGGGGSS
13,301
BAEVM_P10272_3mutA





GSSEAAAKGGS
13,302
MLVCB_P08361_3mut





GSSGGGPAP
13,303
MLVCB_P08361_3mutA





GGSGSS
13,304
MLVBM_Q7SVK7_3mutA_WS





GGGGGSEAAAK
13,305
MLVAV_P03356_3mutA





GSSEAAAK
13,306
PERV_Q4VFZ2_3mutA_WS





GGGGGSGSS
13,307
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGSGSS
13,308
MLVFF_P26809_3mut





PAP

FLV_P10273_3mutA





GGGGG
13,310
MLVMS_P03355_3mutA_WS





EAAAK
13,311
PERV_Q4VFZ2_3mut





GSS

FLV_P10273_3mutA





PAPAPAPAPAPAP
13,313
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAKEAAAKEAA
13,314
MLVCB_P08361_3mut


AK







EAAAKGGGGSEAAAK
13,315
XMRV6_A1Z651_3mut





PAPGGSGGG
13,316
MLVBM_Q7SVK7_3mutA_WS





GGSGGGPAP
13,317
WMSV_P03359_3mutA





GGGGSSEAAAK
13,318
MLVBM_Q7SVK7_3mutA_WS





PAPGGGGSS
13,319
MLVCB_P08361_3mut





GGSGGSGGSGGS
13,320
PERV_Q4VFZ2_3mutA_WS





PAPGGSGGG
13,321
MLVMS_P03355_3mutA_WS





GSSPAPGGS
13,322
MLVCB_P08361_3mutA





GSSGSSGSS
13,323
MLVFF_P26809_3mut





PAPGGGGGS
13,324
MLVBM_Q7SVK7_3mutA_WS





GSSPAP
13,325
PERV_Q4VFZ2_3mut





GGSGGG
13,326
KORV_Q9TTC1-Pro_3mut





EAAAKGGGGSEAAAK
13,327
PERV_Q4VFZ2_3mutA_WS





GGSPAPEAAAK
13,328
PERV_Q4VFZ2_3mutA_WS





EAAAKPAP
13,329
BAEVM_P10272_3mut





GGGGSGGGGSGGGGSGGG
13,330
MLVMS_P03355_3mut


GSGGGGSGGGGS







EAAAKGGGGSS
13,331
MLVFF_P26809_3mut





EAAAKEAAAK
13,332
MLVCB_P08361_3mut





GSSEAAAKGGS
13,333
PERV_Q4VFZ2_3mut





GGSPAP
13,334
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAKEAAAKEAA
13,335
MLVMS_P03355_3mutA_WS


AK







GSSGSSGSSGSSGSS
13,336
BAEVM_P10272_3mut





PAPEAAAK
13,337
MLVMS_P03355_3mut





GSSGGSPAP
13,338
PERV_Q4VFZ2





GGGPAPGGS
13,339
BAEVM_P10272_3mutA





EAAAKPAPGGS
13,340
MLVMS_P03355_PLV919





GGGGSGGGGS
13,341
PERV_Q4VFZ2





GGGEAAAK
13,342
KORV_Q9TTC1-Pro_3mut





EAAAKGGGGGS
13,343
FLV_P10273_3mutA





GGSPAPGSS
13,344
MLVMS_P03355_3mut





GSSPAPEAAAK
13,345
MLVMS_P03355_3mutA_WS





GSAGSAAGSGEF
13,346
MLVBM_Q7SVK7_3mutA_WS





EAAAK
13,347
BAEVM_P10272_3mutA





EAAAKGGGGSS
13,348
BAEVM_P10272_3mutA





GGG

WMSV_P03359_3mut





GGSGSSPAP
13,350
BAEVM_P10272_3mut





GGSEAAAKPAP
13,351
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGSGSS
13,352
MLVCB_P08361_3mut





PAPGSS
13,353
MLVAV_P03356_3mutA





PAPEAAAKGGG
13,354
MLVCB_P08361_3mutA





AEAAAKEAAAKEAAAKEA
13,355
FOAMV_P14350-Pro_2mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSGSSGSS
13,356
PERV_Q4VFZ2_3mut





PAPGGG
13,357
MLVMS_P03355_3mut





PAPGGS
13,358
PERV_Q4VFZ2_3mut





GSSGGG
13,359
MLVMS_P03355_PLV919





GSSGSSGSSGSSGSSGSS
13,360
WMSV_P03359_3mut





PAP

AVIRE_P03360_3mutA





EAAAKGSSPAP
13,362
MLVBM_Q7SVK7_3mutA_WS





GSSGSSGSSGSS
13,363
MLVMS_P03355_PLV919





GGGGSGGGGSGGGGSGGG
13,364
AVIRE_P03360


GSGGGGS







GGGGS
13,365
PERV_Q4VFZ2_3mut





EAAAKGSSGGG
13,366
MLVBM_Q7SVK7_3mutA_WS





GGGGGG
13,367
KORV_Q9TTC1-Pro_3mut





GGSGSSEAAAK
13,368
PERV_Q4VFZ2_3mut





GSSPAPEAAAK
13,369
MLVBM_Q7SVK7_3mutA_WS





GGGGSGGGGS
13,370
MLVBM_Q7SVK7_3mutA_WS





GSSGGGGGS
13,371
MLVAV_P03356_3mutA





GSAGSAAGSGEF
13,372
WMSV_P03359_3mutA





GGGEAAAKGSS
13,373
BAEVM_P10272_3mutA





AEAAAKEAAAKEAAAKEA
13,374
FFV_O93209-Pro_2mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPGGSGGG
13,375
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
13,376
SFV3L_P27401_2mut


AKEAAAK







GGSGSSPAP
13,377
MLVMS_P03355_PLV919





GGGGGG
13,378
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
13,379
PERV_Q4VFZ2_3mut


AKEAAAK







EAAAKGSSPAP
13,380
MLVFF_P26809_3mut





GGGPAPGGS
13,381
MLVBM_Q7SVK7_3mutA_WS





AEAAAKEAAAKEAAAKEA
13,382
SFV3L_P27401


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAP

PERV_Q4VFZ2_3mut





EAAAKGGS
13,384
MLVMS_P03355_PLV919





GSSGGSEAAAK
13,385
WMSV_P03359_3mutA





GGSGSSEAAAK
13,386
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAKEAAAK
13,387
PERV_Q4VFZ2





GGSGGGEAAAK
13,388
MLVMS_P03355_3mutA_WS





GGGGSGGGGSGGGGSGGG
13,389
BAEVM_P10272_3mut


GS







EAAAKGSS
13,390
XMRV6_A1Z651_3mutA





GSSGGGGGS
13,391
WMSV_P03359_3mutA





GSSGSSGSSGSSGSSGSS
13,392
MLVFF_P26809_3mutA





GGSGSS
13,393
MLVAV_P03356_3mutA





EAAAKGGGGSEAAAK
13,394
MLVMS_P03355_PLV919





EAAAKGGGPAP
13,395
PERV_Q4VFZ2





GGSEAAAKGGG
13,396
MLVAV_P03356_3mutA





EAAAKEAAAKEAAAKEAA
13,397
MLVBM_Q7SVK7_3mut


AKEAAAKEAAAK







EAAAKEAAAKEAAAKEAA
13,398
KORV_Q9TTC1-Pro_3mutA


AKEAAAKEAAAK







GSSPAPEAAAK
13,399
MLVFF_P26809_3mutA





GGGGSEAAAKGGGGS
13,400
PERV_Q4VFZ2_3mut





GSSGSSGSSGSS
13,401
PERV_Q4VFZ2_3mut





GGSEAAAK
13,402
MLVFF_P26809_3mutA





GGGGGGGG
13,403
MLVMS_P03355_3mut





GSSGGG
13,404
XMRV6_A1Z651_3mutA





EAAAKGGS
13,405
BAEVM_P10272_3mutA





GGGGS
13,406
BAEVM_P10272_3mutA





GGSEAAAKGGG
13,407
KORV_Q9TTC1-Pro_3mutA





GGSGSSGGG
13,408
KORV_Q9TTC1_3mutA





GGSGSSEAAAK
13,409
WMSV_P03359_3mut





EAAAKGGSGSS
13,410
MLVBM_Q7SVK7_3mutA_WS





GGS

BAEVM_P10272_3mutA





GGGPAPGSS
13,412
WMSV_P03359_3mutA





GSSGSSGSSGSSGSS
13,413
AVIRE_P03360_3mut





GGGEAAAKPAP
13,414
XMRV6_A1Z651_3mut





GSSGGG
13,415
MLVFF_P26809_3mutA





GGSPAPGSS
13,416
PERV_Q4VFZ2_3mut





PAPGGS
13,417
MLVCB_P08361_3mut





PAPAPAPAPAP
13,418
KORV_Q9TTC1_3mutA





GSSGGS
13,419
MLVCB_P08361_3mutA





GSSGGSEAAAK
13,420
PERV_Q4VFZ2_3mut





EAAAKGSSGGS
13,421
MLVMS_P03355_PLV919





EAAAKGGG
13,422
WMSV_P03359_3mut





PAPGGGGGS
13,423
BAEVM_P10272_3mutA





GGGGSEAAAKGGGGS
13,424
WMSV_P03359_3mutA





EAAAKEAAAKEAAAKEAA
13,425
MLVMS_P03355_3mutA_WS


AKEAAAKEAAAK







GGS

KORV_Q9TTC1-Pro_3mutA





GSSGGSPAP
13,427
BAEVM_P10272_3mutA





GGG

MLVMS_P03355_PLV919





PAPGSS
13,429
KORV_Q9TTC1-Pro_3mut





GGSEAAAKGGG
13,430
FLV_P10273_3mutA





GGSEAAAKPAP
13,431
PERV_Q4VFZ2_3mutA_WS





GGGGSSPAP
13,432
XMRV6_A1Z651_3mutA





EAAAKEAAAKEAAAKEAA
13,433
PERV_Q4VFZ2_3mutA_WS


AKEAAAK







GGGG
13,434
PERV_Q4VFZ2_3mutA_WS





GGSEAAAKPAP
13,435
MLVMS_P03355_3mut





PAPGSSGGG
13,436
MLVMS_P03355_3mutA_WS





PAPEAAAKGGS
13,437
AVIRE_P03360_3mut





GGGGSSPAP
13,438
MLVMS_P03355_3mutA_WS





GGGGSGGGGGGGGSGGGG
13,439
PERV_Q4VFZ2_3mut


S







GGGEAAAK
13,440
MLVMS_P03355_3mut





GGGGSS
13,441
MLVFF_P26809_3mut





GGSPAPGSS
13,442
XMRV6_A1Z651_3mut





GGGGS
13,443
KORV_Q9TTC1-Pro_3mutA





EAAAKGSSGGS
13,444
FLV_P10273_3mutA





GSS

MLVMS_P03355_PLV919





GGGG
13,446
MLVMS_P03355_PLV919





GSSGGS
13,447
MLVMS_P03355_PLV919





GGSGGSGGSGGS
13,448
MLVMS_P03355_3mut





PAPEAAAKGGS
13,449
MLVMS_P03355_3mut





EAAAKGSSGGG
13,450
BAEVM_P10272_3mutA





GSSEAAAK
13,451
KORV_Q9TTC1-Pro_3mutA





GSAGSAAGSGEF
13,452
KORV_Q9TTC1_3mutA





GGGGGSEAAAK
13,453
MLVCB_P08361_3mut





GGGG
13,454
WMSV_P03359_3mut





GGGGSSEAAAK
13,455
MLVMS_P03355_PLV919





PAPGGG
13,456
WMSV_P03359_3mutA





EAAAKGGSGGG
13,457
MLVAV_P03356_3mutA





GGGPAPGGS
13,458
MLVMS_P03355_3mut





EAAAKPAP
13,459
PERV_Q4VFZ2_3mutA_WS





GSSGSSGSS
13,460
KORV_Q9TTC1-Pro_3mutA





GSSPAPGGS
13,461
XMRV6_A1Z651_3mut





GGGGGSPAP
13,462
BAEVM_P10272_3mutA





GGSGSSGGG
13,463
PERV_Q4VFZ2_3mutA_WS





GGGEAAAKGSS
13,464
AVIRE_P03360_3mut





GSSEAAAK
13,465
FLV_P10273_3mutA





EAAAK
13,466
MLVMS_P03355_3mut





EAAAKGGSGSS
13,467
WMSV_P03359_3mut





GSSEAAAKGGG
13,468
PERV_Q4VFZ2_3mut





PAPGSSGGG
13,469
BAEVM_P10272_3mutA





EAAAKGGGGGS
13,470
MLVMS_P03355_3mut





GGSEAAAKPAP
13,471
AVIRE_P03360_3mut





GGGPAPGGS
13,472
XMRV6_A1Z651_3mut





GGGGS
13,473
KORV_Q9TTC1_3mutA





GGSGGSGGSGGSGGS
13,474
XMRV6_A1Z651_3mut





GGGPAP
13,475
KORV_Q9TTC1-Pro_3mut





EAAAKPAP
13,476
MLVBM_Q7SVK7_3mutA_WS





GGSEAAAK
13,477
MLVMS_P03355_PLV919





GSSEAAAKPAP
13,478
KORV_Q9TTC1-Pro_3mutA





GGSGSS
13,479
MLVMS_P03355_3mut





EAAAKPAPGGG
13,480
PERV_Q4VFZ2_3mut





GGSPAPEAAAK
13,481
KORV_Q9TTC1_3mutA





GGSEAAAKGGG
13,482
AVIRE_P03360_3mutA





GGGGSEAAAKGGGGS
13,483
MLVMS_P03355_PLV919





GSSGGGEAAAK
13,484
KORV_Q9TTC1-Pro_3mutA





EAAAKGGGPAP
13,485
WMSV_P03359_3mut





GSSPAP
13,486
XMRV6_A1Z651_3mutA





AEAAAKEAAAKEAAAKEA
13,487
SFV3L_P27401-Pro


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSEAAAKGSS
13,488
MLVMS_P03355_PLV919





GSSGGSEAAAK
13,489
KORV_Q9TTC1-Pro_3mutA





GGSEAAAKGSS
13,490
KORV_Q9TTC1-Pro_3mutA





EAAAKGGG
13,491
AVIRE_P03360_3mutA





GSSGGSEAAAK
13,492
BAEVM_P10272_3mutA





GGGGSEAAAKGGGGS
13,493
KORV_Q9TTC1-Pro_3mut





PAPGSSEAAAK
13,494
MLVMS_P03355_3mut





PAPEAAAK
13,495
WMSV_P03359_3mut





PAPGGSGSS
13,496
PERV_Q4VFZ2_3mutA_WS





PAPGSS
13,497
BAEVM_P10272_3mut





PAPGGGGGS
13,498
MLVMS_P03355_3mut





EAAAKPAPGSS
13,499
MLVBM_Q7SVK7_3mutA_WS





GSSPAPGGS
13,500
MLVMS_P03355_PLV919





GGSGSSEAAAK
13,501
MLVMS_P03355_3mut





GGGGGG
13,502
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAKEAAAKEAA
13,503
MLVBM_Q7SVK7_3mut


AK







GGSPAPGSS
13,504
MLVMS_P03355_PLV919





PAPAPAPAPAP
13,505
MLVCB_P08361_3mut





GGSGSSPAP
13,506
WMSV_P03359_3mutA





EAAAKGGSGGG
13,507
PERV_Q4VFZ2_3mutA_WS





GSSGSSGSSGSSGSS
13,508
PERV_Q4VFZ2_3mut





AEAAAKEAAAKEAAAKEA
13,509
KORV_Q9TTC1_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSGGGEAAAK
13,510
WMSV_P03359_3mutA





GSSGGSEAAAK
13,511
FLV_P10273_3mutA





GGGGGGGG
13,512
PERV_Q4VFZ2_3mut





PAPGGSEAAAK
13,513
FLV_P10273_3mutA





GGGGSSPAP
13,514
BAEVM_P10272_3mutA





PAPAPAPAP
13,515
WMSV_P03359_3mut





GGSEAAAKPAP
13,516
PERV_Q4VFZ2_3mut





PAPGGSGGG
13,517
BAEVM_P10272_3mutA





EAAAKEAAAKEAAAKEAA
13,518
MLVMS_P03355_3mut


AKEAAAKEAAAK







GGGGSGGGGSGGGGS
13,519
PERV_Q4VFZ2_3mut





GGSGGGPAP
13,520
PERV_Q4VFZ2_3mut





GGGPAPEAAAK
13,521
MLVFF_P26809_3mut





GGGGGSGSS
13,522
MLVMS_P03355_3mutA_WS





GSS

MLVCB_P08361_3mut





GGGGGSPAP
13,524
MLVMS_P03355_PLV919





GGSPAP
13,525
MLVAV_P03356_3mutA





GGGPAPGGS
13,526
KORV_Q9TTC1-Pro_3mutA





PAPGSSGGG
13,527
FLV_P10273_3mutA





PAPGSSGGG
13,528
WMSV_P03359_3mutA





PAPGGS
13,529
MLVBM_Q7SVK7_3mutA_WS





GGGEAAAKGSS
13,530
PERV_Q4VFZ2_3mutA_WS





GGSEAAAKGSS
13,531
MLVBM_Q7SVK7_3mutA_WS





PAPGGSEAAAK
13,532
MLVCB_P08361_3mut





GGSEAAAKGGG
13,533
XMRV6_A1Z651_3mutA





GGSGGGGSS
13,534
WMSV_P03359_3mut





GGGEAAAKPAP
13,535
KORV_Q9TTC1_3mutA





EAAAKGSS
13,536
KORV_Q9TTC1-Pro_3mut





PAPEAAAKGSS
13,537
MLVFF_P26809_3mut





GSAGSAAGSGEF
13,538
PERV_Q4VFZ2_3mut





EAAAKGGGGGS
13,539
WMSV_P03359_3mut





EAAAKGSSPAP
13,540
WMSV_P03359_3mutA





GGGGSEAAAKGGGGS
13,541
XMRV6_A1Z651_3mutA





GSSEAAAKPAP
13,542
SFV3L_P27401-Pro_2mutA





GGGGGG
13,543
PERV_Q4VFZ2_3mutA_WS





PAPGGS
13,544
BAEVM_P10272_3mut





PAP

AVIRE_P03360_3mut





PAPAPAP
13,546
MLVBM_Q7SVK7_3mutA_WS





GGGG
13,547
PERV_Q4VFZ2_3mutA_WS





GSSGGSEAAAK
13,548
MLVBM_Q7SVK7_3mut





GGSGGGGSS
13,549
MLVFF_P26809_3mut





GGGGSSGGS
13,550
AVIRE_P03360_3mutA





GSSPAPGGG
13,551
PERV_Q4VFZ2_3mutA_WS





GGSEAAAKPAP
13,552
MLVMS_P03355_PLV919





PAP

KORV_Q9TTC1-Pro_3mut





GSSGGS
13,554
PERV_Q4VFZ2_3mut





GGGGG
13,555
PERV_Q4VFZ2_3mut





GSSGGGPAP
13,556
FLV_P10273_3mutA





GSSEAAAKGGG
13,557
KORV_Q9TTC1-Pro_3mut





EAAAKEAAAKEAAAKEAA
13,558
MLVCB_P08361_3mut


AKEAAAKEAAAK







GGSEAAAKPAP
13,559
MLVCB_P08361_3mut





PAPAPAPAPAPAP
13,560
BAEVM_P10272_3mutA





GGGGSEAAAKGGGGS
13,561
MLVMS_P03355_3mut





EAAAKPAPGSS
13,562
MLVMS_P03355_3mut





GSSGSSGSSGSSGSS
13,563
MLVBM_Q7SVK7_3mutA_WS





PAPEAAAKGSS
13,564
MLVAV_P03356_3mut





AEAAAKEAAAKEAAAKEA
13,565
AVIRE_P03360_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







AEAAAKEAAAKEAAAKEA
13,566
PERV_Q4VFZ2_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSEAAAKGGG
13,567
PERV_Q4VFZ2_3mutA_WS





GGSGGGGSS
13,568
MLVFF_P26809_3mutA





PAPEAAAKGSS
13,569
MLVCB_P08361_3mut





GGG

PERV_Q4VFZ2_3mutA_WS





GGSGGGEAAAK
13,571
MLVMS_P03355_3mut





EAAAKGGGGSS
13,572
WMSV_P03359_3mut





GSSPAPGGG
13,573
WMSV_P03359_3mutA





EAAAKGSSGGG
13,574
PERV_Q4VFZ2_3mut





GGSGGGEAAAK
13,575
PERV_Q4VFZ2_3mutA_WS





GGSGGSGGSGGSGGS
13,576
PERV_Q4VFZ2_3mutA_WS





EAAAKPAPGGS
13,577
PERV_Q4VFZ2_3mutA_WS





GGGGGSEAAAK
13,578
PERV_Q4VFZ2_3mutA_WS





GSSPAP
13,579
MLVFF_P26809_3mut





GGGEAAAKPAP
13,580
AVIRE_P03360_3mut





GSSGGSEAAAK
13,581
MLVMS_P03355_PLV919





EAAAKPAPGGS
13,582
WMSV_P03359_3mutA





PAPGGG
13,583
KORV_Q9TTC1_3mutA





EAAAKGSSPAP
13,584
KORV_Q9TTC1-Pro_3mut





GSSPAPEAAAK
13,585
MLVFF_P26809_3mut





GGSGGGEAAAK
13,586
MLVFF_P26809_3mutA





GSSGSSGSS
13,587
WMSV_P03359_3mutA





EAAAKGGS
13,588
BAEVM_P10272_3mut





EAAAKPAPGGS
13,589
KORV_Q9TTC1_3mutA





EAAAKPAPGGS
13,590
BAEVM_P10272_3mutA





GSSGGGGGS
13,591
PERV_Q4VFZ2_3mut





PAPGGGGSS
13,592
PERV_Q4VFZ2_3mut





GSSGSSGSS
13,593
WMSV_P03359_3mut





EAAAKEAAAKEAAAKEAA
13,594
WMSV_P03359_3mut


AK







GGS

AVIRE_P03360_3mut





EAAAKPAPGSS
13,596
MLVFF_P26809_3mut





EAAAKGGG
13,597
KORV_Q9TTC1_3mut





PAPGSSEAAAK
13,598
MLVMS_P03355_3mut





PAPGSSGGS
13,599
MLVMS_P03355_PLV919





GSSPAPEAAAK
13,600
MLVMS_P03355_3mut





GSSGSSGSSGSSGSSGSS
13,601
WMSV_P03359_3mutA





GGGGS
13,602
BAEVM_P10272_3mut





GSSPAP
13,603
MLVMS_P03355_3mut





EAAAKGGGGSEAAAK
13,604
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAK
13,605
WMSV_P03359_3mutA





GGGGSSGGS
13,606
MLVCB_P08361_3mutA





PAPGGSEAAAK
13,607
BAEVM_P10272_3mut





EAAAKGGSPAP
13,608
MLVFF_P26809_3mut





GSSGGSGGG
13,609
MLVBM_Q7SVK7_3mutA_WS





GSSGGS
13,610
PERV_Q4VFZ2_3mut





PAPGGSGSS
13,611
PERV_Q4VFZ2_3mutA_WS





EAAAKGGSGSS
13,612
KORV_Q9TTC1-Pro_3mutA





PAPAP
13,613
MLVCB_P08361_3mut





EAAAKGSSPAP
13,614
PERV_Q4VFZ2_3mutA_WS





EAAAKPAPGGG
13,615
MLVMS_P03355_PLV919





GGGGSGGGGSGGGGSGGG
13,616
MLVBM_Q7SVK7_3mut


GSGGGGSGGGGS







EAAAKGGGGSS
13,617
MLVMS_P03355_PLV919





PAPEAAAK
13,618
PERV_Q4VFZ2_3mut





EAAAKPAPGSS
13,619
BAEVM_P10272_3mutA





GGSPAP
13,620
PERV_Q4VFZ2_3mutA_WS





GGSGGS
13,621
BAEVM_P10272_3mutA





PAPEAAAKGSS
13,622
KORV_Q9TTC1_3mut





PAPGSS
13,623
MLVMS_P03355_PLV919





PAPAPAPAPAP
13,624
MLVAV_P03356_3mutA





GGG

XMRV6_A1Z651_3mutA





GGGPAP
13,626
PERV_Q4VFZ2_3mutA_WS





GSSPAPEAAAK
13,627
KORV_Q9TTC1_3mutA





PAP

BAEVM_P10272_3mutA





GGSPAP
13,629
BAEVM_P10272_3mutA





PAPEAAAKGGS
13,630
MLVMS_P03355_PLV919





PAPGSSGGS
13,631
PERV_Q4VFZ2_3mutA_WS





PAPAPAPAPAPAP
13,632
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAK
13,633
MLVCB_P08361_3mut





GGSGGSGGSGGSGGS
13,634
MLVMS_P03355_PLV919





EAAAKPAPGGS
13,635
MLVMS_P03355_3mut





GGSGGS
13,636
MLVMS_P03355_PLV919





EAAAKPAP
13,637
MLVMS_P03355_3mutA_WS





GGSEAAAK
13,638
XMRV6_A1Z651_3mutA





GGSGGG
13,639
KORV_Q9TTC1_3mut





GGSGGGEAAAK
13,640
PERV_Q4VFZ2_3mut





PAPEAAAKGGG
13,641
AVIRE_P03360





PAPAP
13,642
PERV_Q4VFZ2_3mut





GSS

KORV_Q9TTC1-Pro_3mutA





EAAAKGSSGGG
13,644
MLVAV_P03356_3mutA





GGSPAPGSS
13,645
MLVBM_Q7SVK7_3mutA_WS





PAPEAAAK
13,646
MLVAV_P03356_3mut





EAAAKGGSPAP
13,647
BAEVM_P10272_3mutA





PAPAPAPAP
13,648
WMSV_P03359_3mutA





PAPGGSEAAAK
13,649
MLVMS_P03355_3mut





GGSGGSGGSGGS
13,650
WMSV_P03359_3mut





GGGGGSGSS
13,651
XMRV6_A1Z651_3mut





PAPGGSGGG
13,652
KORV_Q9TTC1_3mutA





GGS

MLVMS_P03355_3mut





EAAAK
13,654
WMSV_P03359_3mut





GGGEAAAKGSS
13,655
MLVBM_Q7SVK7_3mutA_WS





GGSPAPGSS
13,656
MLVCB_P08361_3mut





GGSEAAAKPAP
13,657
PERV_Q4VFZ2_3mut





GGGGSGGGGGGGGSGGGG
13,658
MLVCB_P08361_3mutA


SGGGGS







GGSGSS
13,659
BAEVM_P10272_3mutA





GGGEAAAKGSS
13,660
WMSV_P03359_3mutA





EAAAKGGSPAP
13,661
WMSV_P03359_3mut





GSSPAPEAAAK
13,662
MLVMS_P03355_3mut





GGSGGSGGSGGS
13,663
MLVMS_P03355_PLV919





GSSPAPEAAAK
13,664
WMSV_P03359_3mut





GSSGSSGSSGSS
13,665
PERV_Q4VFZ2





GGSGSSEAAAK
13,666
WMSV_P03359_3mutA





GGSGGG
13,667
MLVFF_P26809_3mut





GGSPAPGGG
13,668
MLVFF_P26809_3mut





GGSGGSGGS
13,669
BAEVM_P10272_3mutA





GGGGSSEAAAK
13,670
MLVBM_Q7SVK7_3mut





GGSPAPGSS
13,671
MLVMS_P03355_3mut





EAAAKPAPGSS
13,672
AVIRE_P03360_3mut





GGGGSSGGS
13,673
FLV_P10273_3mutA





GGSPAPEAAAK
13,674
PERV_Q4VFZ2_3mut





GGSEAAAK
13,675
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSS
13,676
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAKEAA
13,677
MLVMS_P03355_PLV919


AKEAAAK







GGGGG
13,678
PERV_Q4VFZ2_3mut





GGSEAAAKGSS
13,679
MLVCB_P08361_3mutA





GSSGGG
13,680
MLVBM_Q7SVK7_3mutA_WS





PAPGSSGGG
13,681
KORV_Q9TTC1-Pro_3mutA





GGSGGS
13,682
BAEVM_P10272_3mut





EAAAKGGGGGS
13,683
MLVBM_Q7SVK7_3mutA_WS





GGSGSSPAP
13,684
MLVCB_P08361_3mut





PAPGSSGGG
13,685
KORV_Q9TTC1





PAPGGSGGG
13,686
MLVMS_P03355_3mut





GGGG
13,687
WMSV_P03359_3mutA





EAAAKGGSPAP
13,688
MLVCB_P08361_3mut





GSSGSS
13,689
FLV_P10273_3mutA





GGSEAAAKPAP
13,690
SFV3L_P27401_2mut





EAAAKGSSGGS
13,691
MLVAV_P03356_3mutA





AEAAAKEAAAKEAAAKEA
13,692
MLVAV_P03356_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKGGSGSS
13,693
PERV_Q4VFZ2_3mutA_WS





GGGGG
13,694
MLVCB_P08361_3mut





GGGEAAAK
13,695
BAEVM_P10272_3mut





GGSGGSGGSGGS
13,696
MLVCB_P08361_3mut





EAAAKEAAAKEAAAKEAA
13,697
PERV_Q4VFZ2


AKEAAAKEAAAK







PAPAPAPAPAP
13,698
MLVMS_P03355_3mutA_WS





EAAAKEAAAK
13,699
XMRV6_A1Z651_3mut





GSSGGSEAAAK
13,700
PERV_Q4VFZ2_3mutA_WS





PAPGGSEAAAK
13,701
KORV_Q9TTC1-Pro_3mutA





EAAAKGGGPAP
13,702
MLVBM_Q7SVK7_3mutA_WS





PAPGGSGSS
13,703
PERV_Q4VFZ2





SGSETPGTSESATPES
13,704
MLVMS_P03355_3mut





GGSGGS
13,705
MLVMS_P03355_PLV919





EAAAKGGS
13,706
FLV_P10273_3mut





GGSPAPGSS
13,707
MLVMS_P03355_3mutA_WS





EAAAKEAAAKEAAAKEAA
13,708
FFV_O93209_2mut


AK







GSSGGSGGG
13,709
MLVMS_P03355_3mutA_WS





PAPGSSEAAAK
13,710
WMSV_P03359_3mut





PAPAPAPAPAPAP
13,711
KORV_Q9TTC1_3mutA





GGGGSS
13,712
BAEVM_P10272_3mut





GGGGSEAAAKGGGGS
13,713
AVIRE_P03360_3mut





GSSPAPEAAAK
13,714
KORV_Q9TTC1-Pro_3mutA





PAPEAAAKGGG
13,715
MLVBM_Q7SVK7_3mut





EAAAKEAAAK
13,716
WMSV_P03359_3mut





EAAAK
13,717
SFV3L_P27401-Pro_2mutA





GSSGGSGGG
13,718
XMRV6_A1Z651_3mutA





GGGEAAAKPAP
13,719
WMSV_P03359_3mutA





GGSGGS
13,720
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAKEAA
13,721
FOAMV_P14350_2mutA


AKEAAAKEAAAK







GGGGG
13,722
MLVAV_P03356_3mutA





GSSGGSEAAAK
13,723
BAEVM_P10272_3mut





SGGSSGGSSGSETPGTSE
13,724
SFV1_P23074


SATPESSGGSSGGSS







GGSGGGPAP
13,725
MLVCB_P08361_3mut





GGSGSS
13,726
PERV_Q4VFZ2_3mut





SGSETPGTSESATPES
13,727
MLVFF_P26809_3mut





EAAAKGGSPAP
13,728
MLVMS_P03355_3mut





PAPAP
13,729
PERV_Q4VFZ2_3mut





AEAAAKEAAAKEAAAKEA
13,730
MLVBM_Q7SVK7_3mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGGS
13,731
BAEVM_P10272_3mutA





EAAAKEAAAK
13,732
AVIRE_P03360_3mut





GSSGGSEAAAK
13,733
PERV_Q4VFZ2_3mut





GGGEAAAK
13,734
WMSV_P03359_3mut





GSSGGGEAAAK
13,735
AVIRE_P03360_3mutA





GGG

XMRV6_A1Z651_3mut





GGGGSEAAAKGGGGS
13,737
BAEVM_P10272_3mut





GGGG
13,738
MLVMS_P03355_3mut





GGSGGS
13,739
MLVMS_P03355_3mutA_WS





GGSGGGGSS
13,740
MLVBM_Q7SVK7_3mutA_WS





GSSPAPGGS
13,741
PERV_Q4VFZ2_3mut





GSSPAPEAAAK
13,742
PERV_Q4VFZ2_3mutA_WS





EAAAKGGS
13,743
WMSV_P03359_3mut





GGSGGSGGSGGS
13,744
PERV_Q4VFZ2_3mut





GGGGSSEAAAK
13,745
KORV_Q9TTC1-Pro_3mut





PAPAPAPAPAPAP
13,746
MLVAV_P03356_3mut





EAAAKGSSGGG
13,747
MLVMS_P03355_PLV919





GGGGG
13,748
MLVBM_Q7SVK7_3mutA_WS





AEAAAKEAAAKEAAAKEA
13,749
FFV_O93209_2mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







SGGSSGGSSGSETPGTSE
13,750
KORV_Q9TTC1-Pro_3mut


SATPESSGGSSGGSS







GGSPAPGGG
13,751
MLVMS_P03355_3mutA_WS





GGGEAAAKGGS
13,752
MLVMS_P03355_3mut





GGGEAAAK
13,753
PERV_Q4VFZ2_3mut





PAPEAAAKGGG
13,754
MLVMS_P03355_3mut





GSSGSSGSSGSSGSSGSS
13,755
BAEVM_P10272_3mutA





AEAAAKEAAAKEAAAKEA
13,756
GALV_P21414_3mutA


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKGGSPAP
13,757
FFV_O93209-Pro





EAAAKEAAAK
13,758
MLVFF_P26809_3mut





GGGGSGGGGSGGGGSGGG
13,759
PERV_Q4VFZ2_3mutA_WS


GSGGGGSGGGGS







GGSGGSGGSGGS
13,760
MLVAV_P03356_3mutA





EAAAKEAAAKEAAAKEAA
13,761
SFV3L_P27401_2mutA


AKEAAAK







GSSGSSGSSGSSGSSGSS
13,762
BAEVM_P10272_3mut





GGGGS
13,763
MLVMS_P03355_PLV919





AEAAAKEAAAKEAAAKEA
13,764
SFV1_P23074


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGGGSGGGGS
13,765
KORV_Q9TTC1-Pro_3mutA





GGGGSGGGGS
13,766
MLVMS_P03355_3mut





GGSGSS
13,767
KORV_Q9TTC1_3mutA





GSSPAPGGG
13,768
PERV_Q4VFZ2_3mut





GSSGGSPAP
13,769
PERV_Q4VFZ2_3mutA_WS





PAPGGS
13,770
PERV_Q4VFZ2_3mutA_WS





GGSPAPEAAAK
13,771
FOAMV_P14350_2mutA





GGGPAPGGS
13,772
SFV3L_P27401_2mut





PAPGSSGGG
13,773
MLVCB_P08361_3mut





GSSGGGEAAAK
13,774
AVIRE_P03360_3mut





GSSGGG
13,775
XMRV6_A1Z651_3mut





GSSGSS
13,776
PERV_Q4VFZ2_3mut





GSSGGG
13,777
MLVAV_P03356_3mutA





PAPGGGGGS
13,778
PERV_Q4VFZ2_3mut





GSSEAAAK
13,779
MLVMS_P03355_3mut





PAPGGG
13,780
FLV_P10273_3mutA





GGGGSGGGGS
13,781
PERV_Q4VFZ2_3mut





GSSGGS
13,782
MLVMS_P03355_PLV919





GGGGSGGGGS
13,783
SFV3L_P27401_2mut





EAAAKGGSGSS
13,784
FLV_P10273_3mutA





GSSEAAAKGGS
13,785
MLVMS_P03355_3mutA_WS





PAPGSSEAAAK
13,786
SFV3L_P27401_2mutA





GGGGSGGGGS
13,787
SFV3L_P27401-Pro_2mutA





PAPGSSEAAAK
13,788
PERV_Q4VFZ2_3mut





PAPGSSEAAAK
13,789
PERV_Q4VFZ2





GGSPAPGGG
13,790
AVIRE_P03360_3mut





GGGGGS
13,791
PERV_Q4VFZ2_3mutA_WS





GGGGSSGGS
13,792
PERV_Q4VFZ2_3mut





PAPAPAPAP
13,793
AVIRE_P03360_3mutA





GGSGGS
13,794
WMSV_P03359_3mutA





GGGPAPGGS
13,795
PERV_Q4VFZ2_3mut





GGSGGSGGSGGSGGS
13,796
MLVMS_P03355_PLV919





GGSGGG
13,797
PERV_Q4VFZ2_3mut





EAAAKEAAAK
13,798
SFV3L_P27401_2mut





PAPGSS
13,799
XMRV6_A1Z651_3mut





GSSEAAAK
13,800
MLVFF_P26809_3mut





GGSPAPGGG
13,801
MLVMS_P03355_3mut





EAAAKGGG
13,802
WMSV_P03359_3mutA





GSSEAAAKGGS
13,803
PERV_Q4VFZ2_3mutA_WS





GSSGGSPAP
13,804
FFV_O93209





GGGGGS
13,805
KORV_Q9TTC1-Pro_3mut





GSSGGG
13,806
MLVCB_P08361_3mut





GSSGSS
13,807
MLVCB_P08361_3mutA





GGSEAAAKPAP
13,808
BAEVM_P10272_3mut





EAAAKGGGGSS
13,809
MLVCB_P08361_3mut





EAAAKPAPGGS
13,810
KORV_Q9TTC1-Pro_3mutA





GSSGSSGSSGSSGSS
13,811
MLVAV_P03356_3mutA





GGGGSEAAAKGGGGS
13,812
PERV_Q4VFZ2_3mutA_WS





GGSGSS
13,813
KORV_Q9TTC1-Pro_3mut





GSS

SFV3L_P27401-Pro_2mutA





PAPAP
13,815
BAEVM_P10272_3mut





EAAAKPAP
13,816
BAEVM_P10272





EAAAKEAAAKEAAAKEAA
13,817
KORV_Q9TTC1-Pro_3mut


AKEAAAK







GGGGGGG
13,818
PERV_Q4VFZ2_3mutA_WS





GGGGS
13,819
MLVMS_P03355_3mut





GSSGGG
13,820
FLV_P10273_3mutA





PAPAPAPAPAP
13,821
FLV_P10273_3mut





EAAAKEAAAKEAAAK
13,822
WMSV_P03359_3mutA





GSSGGS
13,823
MLVBM_Q7SVK7_3mutA_WS





EAAAKPAPGGG
13,824
MLVMS_P03355_3mut





GSSPAPGGS
13,825
WMSV_P03359_3mut





PAPGSSGGG
13,826
PERV_Q4VFZ2_3mutA_WS





GSSGGG
13,827
AVIRE_P03360_3mutA





PAPGGSGSS
13,828
MLVFF_P26809_3mut





PAPGSS
13,829
PERV_Q4VFZ2_3mut





GGGGGSGSS
13,830
WMSV_P03359_3mutA





EAAAKGGGGSS
13,831
MLVBM_Q7SVK7_3mutA_WS





GGGGGGG
13,832
BAEVM_P10272_3mut





PAPEAAAKGSS
13,833
MLVMS_P03355_3mut





GGSGGGEAAAK
13,834
MLVMS_P03355_PLV919





EAAAKGGGGGS
13,835
MLVCB_P08361_3mut





PAPGGS
13,836
KORV_Q9TTC1-Pro_3mut





GGGG
13,837
FLV_P10273_3mutA





EAAAKGGSGSS
13,838
MLVBM_Q7SVK7_3mutA_WS





GGGGSSGGS
13,839
MLVMS_P03355_3mutA_WS





GGGGGGGG
13,840
WMSV_P03359_3mut





GGSGSSGGG
13,841
MLVMS_P03355_PLV919





GSSEAAAKGGS
13,842
KORV_Q9TTC1-Pro_3mutA





EAAAKPAPGSS
13,843
MLVCB_P08361_3mut





GGSPAPGSS
13,844
KORV_Q9TTC1_3mutA





PAPGSSGGG
13,845
BAEVM_P10272_3mut





EAAAKPAPGSS
13,846
WMSV_P03359_3mut





GGSPAPEAAAK
13,847
XMRV6_A1Z651_3mutA





GSSPAP
13,848
FLV_P10273_3mutA





GSS

BAEVM_P10272_3mutA





EAAAKPAPGGS
13,850
FLV_P10273_3mutA





GGSGSSPAP
13,851
FLV_P10273_3mutA





PAPGSSGGS
13,852
MLVMS_P03355_3mut





GSAGSAAGSGEF
13,853
PERV_Q4VFZ2_3mutA_WS





GSSGGSEAAAK
13,854
KORV_Q9TTC1_3mutA





GSSGGS
13,855
MLVMS_P03355_3mutA_WS





EAAAKGGGGSEAAAK
13,856
SFV3L_P27401_2mut





GSSGGS
13,857
PERV_Q4VFZ2_3mutA_WS





GGSPAPEAAAK
13,858
FLV_P10273_3mut





GGSEAAAKGSS
13,859
PERV_Q4VFZ2_3mutA_WS





GSSPAPEAAAK
13,860
PERV_Q4VFZ2_3mutA_WS





GGSGSSGGG
13,861
PERV_Q4VFZ2_3mut





GGGG
13,862
AVIRE_P03360_3mutA





GGSEAAAKPAP
13,863
WMSV_P03359_3mut





GSSGGSPAP
13,864
MLVAV_P03356_3mutA





GSSGGSEAAAK
13,865
MLVMS_P03355_3mut





PAPEAAAKGGS
13,866
KORV_Q9TTC1-Pro_3mut





GGSPAP
13,867
PERV_Q4VFZ2_3mutA_WS





GGSEAAAK
13,868
MLVAV_P03356_3mutA





EAAAKGGGGSEAAAK
13,869
KORV_Q9TTC1-Pro_3mut





SGGSSGGSSGSETPGTSE
13,870
MLVMS_P03355_PLV919


SATPESSGGSSGGSS







GSSEAAAK
13,871
KORV_Q9TTC1_3mutA





GGG

AVIRE_P03360





GGSEAAAKGSS
13,873
MLVBM_Q7SVK7_3mut





GGSEAAAKGSS
13,874
MLVMS_P03355_3mut





GGSPAPEAAAK
13,875
MLVCB_P08361_3mut





GGSGGGEAAAK
13,876
MLVCB_P08361_3mut





GGSEAAAKPAP
13,877
MLVMS_P03355_3mutA_WS





EAAAKGGSGSS
13,878
KORV_Q9TTC1-Pro_3mut





GGGEAAAKGGS
13,879
MLVCB_P08361_3mut





EAAAKGGGGSEAAAK
13,880
FLV_P10273_3mutA





GGSPAP
13,881
MLVFF_P26809_3mut





GGGGSSGGS
13,882
XMRV6_A1Z651_3mutA





PAP

MLVCB_P08361_3mut





GGS

SFV3L_P27401-Pro_2mutA





GGGGSGGGGS
13,885
MLVMS_P03355_3mut





GGGEAAAKGGS
13,886
MLVAV_P03356_3mutA





GSSGSSGSSGSSGSSGSS
13,887
MLVMS_P03355_PLV919





PAPGSS
13,888
MLVCB_P08361_3mut





GGSGGSGGS
13,889
MLVMS_P03355_PLV919





PAPGGSGGG
13,890
FLV_P10273_3mutA





GGGGSGGGGSGGGGS
13,891
FLV_P10273_3mut





GGSGSSGGG
13,892
KORV_Q9TTC1-Pro_3mutA





GGSGGSGGS
13,893
GALV_P21414_3mutA





GGGEAAAKGGS
13,894
WMSV_P03359_3mut





SGSETPGTSESATPES
13,895
KORV_Q9TTC1_3mutA





EAAAKGGGGGS
13,896
KORV_Q9TTC1-Pro_3mut





EAAAKGSSPAP
13,897
BAEVM_P10272_3mut





GGGG
13,898
MLVCB_P08361_3mut





GGGGSGGGGSGGGGSGGG
13,899
MLVBM_Q7SVK7_3mut


GSGGGGS







GSSGGSGGG
13,900
MLVMS_P03355_PLV919





GGSGSS
13,901
MLVFF_P26809_3mut





EAAAKGGS
13,902
AVIRE_P03360_3mutA





GSSEAAAKGGS
13,903
MLVBM_Q7SVK7_3mutA_WS





EAAAKPAPGGG
13,904
WMSV_P03359_3mut





PAPGSSGGG
13,905
MLVCB_P08361_3mutA





GGGGSSEAAAK
13,906
KORV_Q9TTC1-Pro_3mutA





GSSEAAAKPAP
13,907
BAEVM_P10272_3mutA





PAPGGGEAAAK
13,908
MLVBM_Q7SVK7_3mutA_WS





GGSGGGEAAAK
13,909
MLVCB_P08361_3mutA





GGGGSGGGGGGGGSGGGG
13,910
FFV_O93209


SGGGGSGGGGS







EAAAKGGGGGS
13,911
GALV_P21414_3mutA





GGSPAPGGG
13,912
MLVMS_P03355_3mut





GSSGSSGSS
13,913
FLV_P10273_3mutA





EAAAK
13,914
MLVBM_Q7SVK7_3mut





GGGGSSGGS
13,915
MLVMS_P03355_3mut





GGSGSSPAP
13,916
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
13,917
BAEVM_P10272_3mut


AK







GGGPAPGSS
13,918
MLVMS_P03355_3mut





GSSPAPGGS
13,919
PERV_Q4VFZ2_3mutA_WS





PAPAP
13,920
FLV_P10273_3mutA





PAPAPAPAP
13,921
PERV_Q4VFZ2_3mut





GGGGGSEAAAK
13,922
GALV_P21414_3mutA





GGGGGSGSS
13,923
BAEVM_P10272_3mutA





GGGEAAAKGSS
13,924
KORV_Q9TTC1_3mutA





GGGGGSPAP
13,925
AVIRE_P03360_3mut





GGGGGSEAAAK
13,926
SFV3L_P27401_2mutA





GGS

KORV_Q9TTC1_3mutA





GGGGGGG
13,928
PERV_Q4VFZ2_3mut





SGSETPGTSESATPES
13,929
SFV3L_P27401_2mutA





EAAAKGGSGGG
13,930
MLVMS_P03355_3mut





GGGGS
13,931
MLVFF_P26809_3mut





EAAAKGSSGGG
13,932
BAEVM_P10272_3mut





EAAAKPAPGGS
13,933
MLVF5_P26810_3mutA





SGGSSGGSSGSETPGTSE
13,934
SFV3L_P27401_2mutA


SATPESSGGSSGGSS







GGSPAPGGG
13,935
WMSV_P03359_3mutA





GSAGSAAGSGEF
13,936
MLVFF_P26809_3mut





GGGGSSGGS
13,937
MLVMS_P03355_3mutA_WS





GGGGGGG
13,938
MLVCB_P08361_3mut





GSSEAAAK
13,939
WMSV_P03359_3mut





PAPGSS
13,940
FLV_P10273_3mutA





GSSGGG
13,941
PERV_Q4VFZ2_3mutA_WS





PAPGGG
13,942
MLVFF_P26809_3mut





GGGGGSPAP
13,943
MLVMS_P03355_3mut





GGSEAAAK
13,944
XMRV6_A1Z651_3mut





GSSGGG
13,945
PERV_Q4VFZ2_3mut





GGSGGSGGSGGS
13,946
MLVMS_P03355_3mut





PAPAP
13,947
AVIRE_P03360_3mut





GGSEAAAK
13,948
PERV_Q4VFZ2_3mut





GGGGS
13,949
MLVMS_P03355_PLV919





GGGG
13,950
BAEVM_P10272_3mutA





EAAAKGGGGSS
13,951
MLVCB_P08361_3mutA





EAAAKEAAAKEAAAK
13,952
GALV_P21414_3mutA





PAPGGGEAAAK
13,953
KORV_Q9TTC1





EAAAKGGSPAP
13,954
MLVMS_P03355_3mut





GGSGSSEAAAK
13,955
MLVMS_P03355_3mut





GGSPAPEAAAK
13,956
FLV_P10273_3mutA





GGGGGGG
13,957
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
13,958
SFV1_P23074_2mutA


AKEAAAKEAAAK







EAAAKGSSGGS
13,959
MLVMS_P03355_3mut





GSSEAAAKPAP
13,960
MLVFF_P26809_3mut





GGGGSS
13,961
FLV_P10273_3mutA





EAAAKGGSGGG
13,962
AVIRE_P03360_3mutA





GGSGGS
13,963
PERV_Q4VFZ2_3mutA_WS





GGGGGSPAP
13,964
AVIRE_P03360_3mutA





EAAAKEAAAKEAAAK
13,965
XMRV6_A1Z651_3mut





PAPEAAAKGGS
13,966
FLV_P10273_3mutA





GSSGGSEAAAK
13,967
MLVCB_P08361_3mut





EAAAKGGSGGG
13,968
MLVMS_P03355





GGSGGGPAP
13,969
MLVMS_P03355_3mut





GGS

XMRV6_A1Z651_3mut





GGSEAAAKPAP
13,971
MLVFF_P26809_3mut





EAAAKGGG
13,972
MLVMS_P03355_PLV919





GSSGSSGSSGSS
13,973
WMSV_P03359_3mut





GGSGSSPAP
13,974
PERV_Q4VFZ2_3mut





GGGEAAAK
13,975
MLVMS_P03355_3mutA_WS





GSSPAPGGS
13,976
KORV_Q9TTC1-Pro_3mutA





GSSEAAAKGGG
13,977
SFV3L_P27401_2mut





EAAAKPAPGGS
13,978
MLVCB_P08361_3mut





GGSGGGEAAAK
13,979
PERV_Q4VFZ2





GGSGSS
13,980
MLVCB_P08361_3mut





GGSGGGEAAAK
13,981
MLVBM_Q7SVK7_3mutA_WS





GGSGGSGGSGGSGGSGGS
13,982
FLV_P10273_3mut





PAPEAAAKGSS
13,983
MLVMS_P03355_3mut





EAAAKGSSGGS
13,984
WMSV_P03359_3mutA





GGSGSSEAAAK
13,985
MLVCB_P08361_3mut





GGSGSSEAAAK
13,986
KORV_Q9TTC1_3mutA





GSSGGSGGG
13,987
MLVMS_P03355_PLV919





EAAAKGGSGGG
13,988
SFV3L_P27401-Pro_2mutA





GGSGGS
13,989
AVIRE_P03360_3mutA





GSAGSAAGSGEF
13,990
MLVMS_P03355_PLV919





GGSGSS
13,991
GALV_P21414_3mutA





GGGG
13,992
MLVFF_P26809_3mutA





GGGGSGGGGSGGGGSGGG
13,993
WMSV_P03359_3mut


GS







SGSETPGTSESATPES
13,994
BAEVM_P10272_3mut





EAAAKEAAAKEAAAKEAA
13,995
FOAMV_P14350_2mutA


AK







GGGEAAAKGGS
13,996
FLV_P10273_3mutA





GSSGGSEAAAK
13,997
MLVFF_P26809_3mut





EAAAKGGGGSS
13,998
MLVAV_P03356_3mut





PAPGGSEAAAK
13,999
KORV_Q9TTC1-Pro_3mut





EAAAK
14,000
XMRV6_A1Z651_3mut





GSSGSSGSSGSSGSSGSS
14,001
PERV_Q4VFZ2_3mut





GGGG
14,002
MLVCB_P08361_3mutA





GSSGSS
14,003
WMSV_P03359_3mutA





GSSGGSPAP
14,004
AVIRE_P03360_3mut





GGSGGSGGS
14,005
MLVCB_P08361_3mut





EAAAKGGGPAP
14,006
FLV_P10273_3mutA





GGGGSGGGGS
14,007
MLVCB_P08361_3mut





GGSEAAAKGSS
14,008
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAKEAA
14,009
SFV3L_P27401_2mutA


AKEAAAKEAAAK







GGSGSSEAAAK
14,010
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAKEAA
14,011
SFV3L_P27401-Pro_2mutA


AK







GSSEAAAKGGS
14,012
FLV_P10273_3mutA





GGSGSS
14,013
PERV_Q4VFZ2





GGSGSSEAAAK
14,014
SFV3L_P27401-Pro_2mutA





GSSGSSGSS
14,015
XMRV6_A1Z651_3mutA





EAAAKGSSPAP
14,016
KORV_Q9TTC1_3mutA





EAAAKPAP
14,017
FLV_P10273_3mutA





GGSGSSEAAAK
14,018
KORV_Q9TTC1-Pro_3mut





GGGGSGGGGSGGGGSGGG
14,019
KORV_Q9TTC1_3mutA


GSGGGGSGGGGS







GGGGSGGGGSGGGGS
14,020
KORV_Q9TTC1-Pro_3mutA





GGGGGGG
14,021
FLV_P10273_3mut





EAAAKGSS
14,022
WMSV_P03359_3mut





EAAAKGGGPAP
14,023
MLVCB_P08361_3mut





GSSGSS
14,024
MLVBM_Q7SVK7_3mutA_WS





EAAAKGGGGGS
14,025
MLVFF_P26809_3mut





GGSGGGEAAAK
14,026
FLV_P10273_3mutA





PAPGSS
14,027
MLVFF_P26809_3mutA





PAPGSS
14,028
BAEVM_P10272_3mutA





GGSPAPGSS
14,029
AVIRE_P03360_3mut





GGGGSSEAAAK
14,030
MLVMS_P03355_3mut





GSSGGGGGS
14,031
FFV_O93209-Pro





EAAAKGSSPAP
14,032
PERV_Q4VFZ2_3mut





GSSPAPGGS
14,033
PERV_Q4VFZ2_3mut





GGGGGG
14,034
BAEVM_P10272_3mut





EAAAKGGGGSS
14,035
PERV_Q4VFZ2_3mutA_WS





PAPGGSEAAAK
14,036
KORV_Q9TTC1_3mutA





SGGSSGGSSGSETPGTSE
14,037
MLVMS_P03355_3mutA_WS


SATPESSGGSSGGSS







GSSGSSGSSGSS
14,038
MLVMS_P03355_3mut





EAAAKGSSGGG
14,039
MLVMS_P03355_PLV919





GGSEAAAKPAP
14,040
AVIRE_P03360_3mutA





GSSGSSGSSGSSGSS
14,041
WMSV_P03359_3mutA





GGGEAAAKPAP
14,042
FLV_P10273_3mutA





PAPGSSGGG
14,043
KORV_Q9TTC1_3mutA





GSSGSS
14,044
MLVMS_P03355_3mutA_WS





PAPEAAAK
14,045
BAEVM_P10272_3mut





GGGPAPGSS
14,046
PERV_Q4VFZ2





GSSGGSPAP
14,047
MLVFF_P26809_3mut





GGGGSS
14,048
SFV3L_P27401_2mut





PAPEAAAKGSS
14,049
SFV3L_P27401_2mut





GGSGGGPAP
14,050
XMRV6_A1Z651_3mutA





PAPGGS
14,051
BAEVM_P10272_3mutA





EAAAKGGGGGS
14,052
AVIRE_P03360_3mut





GSSGGSPAP
14,053
KORV_Q9TTC1-Pro_3mutA





GSSGGGGGS
14,054
WMSV_P03359_3mut





GGGEAAAKGGS
14,055
AVIRE_P03360_3mut





GGGEAAAKGSS
14,056
BAEVM_P10272_3mut





PAPEAAAKGSS
14,057
MLVAV_P03356_3mutA





GSSGSSGSSGSSGSS
14,058
MLVCB_P08361_3mut





GGSPAPGSS
14,059
FLV_P10273_3mutA





EAAAKGSSPAP
14,060
BAEVM_P10272_3mutA





GGSGGSGGSGGSGGSGGS
14,061
PERV_Q4VFZ2





GGGGSSEAAAK
14,062
FLV_P10273_3mutA





GGGGSSPAP
14,063
FFV_O93209





GSSGGSPAP
14,064
MLVMS_P03355_3mut





GGGPAPGSS
14,065
MLVMS_P03355_PLV919





PAPGSSGGS
14,066
PERV_Q4VFZ2_3mut





GGGGGSPAP
14,067
MLVFF_P26809_3mut





SGSETPGTSESATPES
14,068
MLVMS_P03355_3mutA_WS





GSSGSSGSSGSSGSS
14,069
KORV_Q9TTC1_3mutA





GSSPAPGGG
14,070
WMSV_P03359_3mut





PAPAPAPAPAPAP
14,071
SFV3L_P27401_2mutA





GGGPAPGGS
14,072
MLVMS_P03355_3mut





PAPGGSEAAAK
14,073
WMSV_P03359_3mut





GGGGSSEAAAK
14,074
FFV_O93209-Pro





GGSPAPGGG
14,075
FLV_P10273_3mutA





GSSPAPEAAAK
14,076
AVIRE_P03360_3mut





GGGEAAAK
14,077
FLV_P10273_3mutA





PAPEAAAKGGG
14,078
MLVCB_P08361_3mut





GGSPAPGGG
14,079
MLVCB_P08361_3mut





GGSGGGGSS
14,080
BAEVM_P10272_3mutA





GSSPAPEAAAK
14,081
MLVCB_P08361_3mut





GGSPAPGGG
14,082
KORV_Q9TTC1-Pro_3mutA





PAPGGSGSS
14,083
KORV_Q9TTC1_3mutA





GSSPAP
14,084
KORV_Q9TTC1-Pro_3mutA





SGSETPGTSESATPES
14,085
MLVMS_P03355





GSSGSSGSS
14,086
MLVAV_P03356_3mutA





PAPGSSGGS
14,087
PERV_Q4VFZ2_3mutA_WS





PAPGGS
14,088
KORV_Q9TTC1-Pro_3mutA





PAPEAAAKGGG
14,089
SFV3L_P27401-Pro_2mutA





GGSGGSGGS
14,090
BAEVM_P10272_3mut





PAPGGS
14,091
MLVFF_P26809_3mut





GSSGGSPAP
14,092
MLVMS_P03355_PLV919





GSSGGGGGS
14,093
FLV_P10273_3mutA





GGGGGSPAP
14,094
KORV_Q9TTC1-Pro_3mut





EAAAKPAPGSS
14,095
SFV3L_P27401-Pro_2mutA





EAAAKGGSPAP
14,096
KORV_Q9TTC1-Pro





GGGPAPEAAAK
14,097
MLVMS_P03355_PLV919





GGSEAAAKGSS
14,098
MLVMS_P03355





PAPEAAAKGSS
14,099
KORV_Q9TTC1_3mutA





PAPEAAAKGGS
14,100
WMSV_P03359_3mutA





GSSGGG
14,101
PERV_Q4VFZ2_3mutA_WS





EAAAKGGGGSS
14,102
MLVMS_P03355_PLV919





EAAAKGGSPAP
14,103
AVIRE_P03360_3mutA





GGGGSSGGS
14,104
MLVMS_P03355_PLV919





PAPEAAAKGSS
14,105
PERV_Q4VFZ2_3mutA_WS





EAAAKGGGGGS
14,106
BAEVM_P10272_3mut





GSSGGGGGS
14,107
MLVMS_P03355_3mut





PAPAPAPAP
14,108
KORV_Q9TTC1_3mutA





GGSGGSGGSGGS
14,109
MLVAV_P03356_3mut





PAPAPAPAP
14,110
SFV3L_P27401_2mut





GSSEAAAKPAP
14,111
MLVMS_P03355_3mut





GGSGGGEAAAK
14,112
SFV3L_P27401_2mutA





GSSGGSGGG
14,113
MLVMS_P03355_3mutA_WS





GGGGGSPAP
14,114
MLVCB_P08361_3mutA





GGGEAAAKGSS
14,115
XMRV6_A1Z651_3mutA





GGGGSSPAP
14,116
BAEVM_P10272_3mut





GGSGGG
14,117
PERV_Q4VFZ2_3mut





GGGGSS
14,118
MLVBM_Q7SVK7_3mutA_WS





EAAAKGSSGGS
14,119
PERV_Q4VFZ2_3mutA_WS





GSSGGGGGS
14,120
PERV_Q4VFZ2





EAAAKGSSGGS
14,121
PERV_Q4VFZ2_3mut





EAAAKEAAAK
14,122
MLVAV_P03356_3mut





GSSGGGEAAAK
14,123
MLVAV_P03356_3mut





GSSPAPGGG
14,124
XMRV6_A1Z651_3mut





GGGGSGGGGSGGGGS
14,125
PERV_Q4VFZ2_3mut





EAAAKEAAAKEAAAKEAA
14,126
KORV_Q9TTC1_3mutA


AK







EAAAKGGSGSS
14,127
MLVBM_Q7SVK7_3mut





PAPEAAAK
14,128
BLVJ_P03361





GSSGGG
14,129
FFV_O93209-Pro





GGSGGGEAAAK
14,130
KORV_Q9TTC1-Pro_3mutA





EAAAK
14,131
FLV_P10273_3mutA





GGGGSSPAP
14,132
MLVMS_P03355_3mut





GSS

SFV3L_P27401-Pro_2mut





PAPEAAAKGSS
14,134
BAEVM_P10272_3mut





GGGGGSPAP
14,135
PERV_Q4VFZ2_3mut





GSSGSSGSS
14,136
BAEVM_P10272_3mutA





GGGGSGGGGSGGGGSGGG
14,137
SFV1_P23074_2mut


GS







GGGGSSEAAAK
14,138
SFV3L_P27401_2mutA





GGGGSGGGGSGGGGSGGG
14,139
FOAMV_P14350-Pro_2mut


GS







PAPGSSEAAAK
14,140
MLVBM_Q7SVK7_3mutA_WS





GGGGGSGSS
14,141
MLVFF_P26809_3mutA





GGSEAAAKGGG
14,142
MLVBM_Q7SVK7_3mut





PAPGSSGGG
14,143
PERV_Q4VFZ2





GGS

PERV_Q4VFZ2_3mutA_WS





EAAAKGGSGSS
14,145
FLV_P10273_3mut





GGGEAAAK
14,146
WMSV_P03359_3mutA





GGSEAAAKPAP
14,147
MLVBM_Q7SVK7_3mut





SGSETPGTSESATPES
14,148
FOAMV_P14350-Pro_2mutA





EAAAKPAPGGS
14,149
AVIRE_P03360_3mut





EAAAKGGGGGS
14,150
KORV_Q9TTC1-Pro_3mutA





GGGGS
14,151
PERV_Q4VFZ2_3mut





GGSEAAAKGSS
14,152
MLVFF_P26809_3mutA





GGSEAAAKGGG
14,153
AVIRE_P03360





GGSGGSGGSGGSGGSGGS
14,154
SFV3L_P27401_2mut





GGSEAAAKGSS
14,155
SFV3L_P27401-Pro_2mutA





GGGEAAAKPAP
14,156
MLVCB_P08361_3mut





GGSEAAAK
14,157
MLVMS_P03355_PLV919





GGSPAPGSS
14,158
KORV_Q9TTC1-Pro_3mutA





GSSPAPEAAAK
14,159
WMSV_P03359_3mutA





GGSGSS
14,160
KORV_Q9TTC1-Pro_3mutA





PAPGGGGGS
14,161
AVIRE_P03360_3mut





PAPEAAAKGSS
14,162
FFV_O93209-Pro





GGSGGGEAAAK
14,163
WMSV_P03359_3mut





PAPGGG
14,164
MLVMS_P03355_3mut





EAAAKGGG
14,165
FLV_P10273_3mutA





GSSGSSGSSGSS
14,166
MLVCB_P08361_3mut





EAAAKGGSGGG
14,167
FFV_O93209





GSSPAPGGS
14,168
PERV_Q4VFZ2_3mutA_WS





GSSPAPGGS
14,169
MLVCB_P08361_3mut





GGGPAP
14,170
WMSV_P03359_3mutA





GGGPAP
14,171
KORV_Q9TTC1_3mutA





GGSPAPGSS
14,172
KORV_Q9TTC1-Pro_3mut





PAPAP
14,173
MLVMS_P03355_3mut





GGGGGGG
14,174
MLVMS_P03355_3mut





GGGGG
14,175
KORV_Q9TTC1-Pro_3mut





GSAGSAAGSGEF
14,176
FOAMV_P14350_2mutA





PAPAP
14,177
KORV_Q9TTC1-Pro_3mutA





GGSEAAAKGGG
14,178
SFV3L_P27401-Pro_2mutA





PAPAP
14,179
WMSV_P03359_3mut





GGGGSGGGGSGGGGS
14,180
SFV3L_P27401_2mut





PAPGGS
14,181
KORV_Q9TTC1_3mutA





GGGEAAAKPAP
14,182
FLV_P10273_3mut





GGGGGS
14,183
MLVAV_P03356_3mutA





GSSEAAAKGGG
14,184
WMSV_P03359_3mut





EAAAKGGGGSS
14,185
GALV_P21414_3mutA





GSSGGS
14,186
MLVAV_P03356_3mutA





GSSGGG
14,187
MLVBM_Q7SVK7_3mut





PAPAPAP
14,188
SFV3L_P27401-Pro_2mutA





GGGG
14,189
KORV_Q9TTC1_3mutA





EAAAKPAPGGS
14,190
MLVFF_P26809_3mut





GGGGGGGGS
14,191
XMRV6_A1Z651_3mut





EAAAKGGG
14,192
MLVCB_P08361_3mut





GGGGSSPAP
14,193
KORV_Q9TTC1_3mutA





GSSEAAAKGGG
14,194
KORV_Q9TTC1-Pro_3mutA





GGGGG
14,195
BLVJ_P03361_2mutB





GGGEAAAKGSS
14,196
FFV_O93209-Pro





GSSGSSGSS
14,197
BAEVM_P10272_3mut





GSSGGSPAP
14,198
PERV_Q4VFZ2_3mut





EAAAKGGS
14,199
KORV_Q9TTC1_3mut





GGSPAPEAAAK
14,200
AVIRE_P03360_3mut





GGSEAAAK
14,201
WMSV_P03359_3mut





GSSGGS
14,202
KORV_Q9TTC1-Pro_3mutA





GGGPAPEAAAK
14,203
KORV_Q9TTC1_3mutA





PAPGSS
14,204
WMSV_P03359_3mutA





GGSEAAAKGSS
14,205
FLV_P10273_3mutA





EAAAKEAAAKEAAAKEAA
14,206
SFV3L_P27401


AKEAAAK







GSSEAAAKGGG
14,207
SFV3L_P27401-Pro_2mutA





GGGGSEAAAKGGGGS
14,208
KORV_Q9TTC1-Pro_3mutA





GGSGGSGGS
14,209
WMSV_P03359_3mut





GGGGGSGSS
14,210
KORV_Q9TTC1-Pro





GGGGSGGGGGGGGSGGGG
14,211
MLVMS_P03355_3mut


S







EAAAKGGG
14,212
PERV_Q4VFZ2





GGSEAAAKGGG
14,213
KORV_Q9TTC1-Pro_3mut





GSSGGSGGG
14,214
PERV_Q4VFZ2_3mutA_WS





GGGGGS
14,215
PERV_Q4VFZ2_3mut





GSAGSAAGSGEF
14,216
PERV_Q4VFZ2





PAPEAAAKGSS
14,217
BAEVM_P10272_3mutA





GSSPAPGGG
14,218
MLVCB_P08361_3mut





GGGGSSPAP
14,219
KORV_Q9TTC1-Pro_3mutA





PAPGGSGGG
14,220
MLVFF_P26809_3mut





GSSPAP
14,221
KORV_Q9TTC1_3mutA





PAPGSS
14,222
SFV3L_P27401-Pro_2mut





GGSGGGGSS
14,223
MLVMS_P03355_PLV919





GSSGGS
14,224
WMSV_P03359_3mutA





EAAAKGGGGGS
14,225
PERV_Q4VFZ2





GGGGG
14,226
KORV_Q9TTC1_3mutA





EAAAKGSS
14,227
MLVMS_P03355_PLV919





EAAAKEAAAKEAAAKEAA
14,228
FLV_P10273_3mut


AKEAAAK







EAAAKEAAAKEAAAKEAA
14,229
SFV3L_P27401-Pro_2mut


AK







GSAGSAAGSGEF
14,230
SFV3L_P27401_2mutA





GGGPAPGGS
14,231
FLV_P10273_3mutA





GGSEAAAKGGG
14,232
MLVCB_P08361_3mut





PAPGGGEAAAK
14,233
BAEVM_P10272_3mut





EAAAKPAPGSS
14,234
FOAMV_P14350_2mut





GGSEAAAK
14,235
KORV_Q9TTC1_3mutA





GGSGSS
14,236
AVIRE_P03360





GGSPAPEAAAK
14,237
MLVMS_P03355_PLV919





GGGGS
14,238
XMRV6_A1Z651_3mut





GGSPAPGGG
14,239
XMRV6_A1Z651_3mut





EAAAKPAPGGS
14,240
PERV_Q4VFZ2





GSSPAP
14,241
BAEVM_P10272_3mut





GGSGSSGGG
14,242
FLV_P10273_3mutA





PAPGGG
14,243
PERV_Q4VFZ2_3mutA_WS





GSSGGSEAAAK
14,244
MLVBM_Q7SVK7_3mut





GGSEAAAK
14,245
MLVMS_P03355_3mut





GGGPAPGGS
14,246
MLVFF_P26809_3mut





GSAGSAAGSGEF
14,247
MLVBM_Q7SVK7_3mutA_WS





EAAAKPAPGGS
14,248
SFVCP_Q87040





PAPGGG
14,249
PERV_Q4VFZ2_3mutA_WS





GSSPAPEAAAK
14,250
MLVBM_Q7SVK7





PAPEAAAK
14,251
MLVBM_Q7SVK7_3mut





PAPGGGGGS
14,252
AVIRE_P03360_3mutA





GGSEAAAKPAP
14,253
MLVBM_Q7SVK7_3mut





EAAAKGSS
14,254
WMSV_P03359_3mutA





GGGEAAAK
14,255
MLVFF_P26809_3mutA





EAAAKEAAAKEAAAK
14,256
MLVMS_P03355_3mut





PAPEAAAKGGG
14,257
BAEVM_P10272_3mut





PAPAPAP
14,258
MLVCB_P08361_3mut





EAAAKPAPGGS
14,259
BAEVM_P10272_3mut





GGGGSGGGGS
14,260
FLV_P10273_3mut





GGGGSEAAAKGGGGS
14,261
KORV_Q9TTC1_3mut





EAAAK
14,262
FLV_P10273_3mut





PAPAPAP
14,263
WMSV_P03359_3mut





GGGGSEAAAKGGGGS
14,264
FFV_O93209-Pro





GGSPAPEAAAK
14,265
MLVMS_P03355_3mut





GGSGSSGGG
14,266
XMRV6_A1Z651_3mut





GGSPAPGSS
14,267
PERV_Q4VFZ2_3mut





SGGSSGGSSGSETPGTSE
14,268
SFV3L_P27401-Pro_2mutA


SATPESSGGSSGGSS







EAAAKGGGPAP
14,269
BAEVM_P10272_3mutA





GSSGGSEAAAK
14,270
MLVMS_P03355_3mutA_WS





SGSETPGTSESATPES
14,271
PERV_Q4VFZ2_3mutA_WS





EAAAKEAAAKEAAAKEAA
14,272
KORV_Q9TTC1-Pro_3mutA


AKEAAAK







GSSGSSGSS
14,273
KORV_Q9TTC1_3mutA





GSSPAPGGG
14,274
SFV3L_P27401-Pro_2mutA





GSSGGGEAAAK
14,275
KORV_Q9TTC1_3mutA





GGSGGGGSS
14,276
PERV_Q4VFZ2_3mutA_WS





GSSGGGEAAAK
14,277
MLVCB_P08361_3mut





GSSEAAAKGGG
14,278
MLVCB_P08361_3mut





GGSGGGGSS
14,279
KORV_Q9TTC1_3mutA





GGSGSSPAP
14,280
PERV_Q4VFZ2_3mutA_WS





GSSPAP
14,281
MLVMS_P03355_3mut





GGGGSSEAAAK
14,282
AVIRE_P03360





GGS

WMSV_P03359_3mut





EAAAKEAAAK
14,284
PERV_Q4VFZ2_3mut





PAPAPAPAP
14,285
MLVAV_P03356_3mut





GGSEAAAKGGG
14,286
KORV_Q9TTC1_3mutA





PAPGGG
14,287
MLVAV_P03356_3mut





EAAAKGSS
14,288
BAEVM_P10272_3mut





GGGGSGGGGS
14,289
WMSV_P03359_3mutA





GGSGGSGGS
14,290
SFV3L_P27401_2mut





EAAAK
14,291
MLVCB_P08361_3mut





GGGGSSGGS
14,292
WMSV_P03359_3mutA





GGGPAPEAAAK
14,293
MLVAV_P03356_3mutA





EAAAKEAAAKEAAAK
14,294
FFV_O93209





GSSEAAAKGGG
14,295
MLVBM_Q7SVK7_3mut





GGGPAPGGS
14,296
FLV_P10273_3mut





GGSEAAAKGGG
14,297
WMSV_P03359_3mut





EAAAKGGGGGS
14,298
XMRV6_A1Z651_3mutA





EAAAKGGSGGG
14,299
FLV_P10273_3mutA





GGSEAAAKGGG
14,300
SFV3L_P27401_2mutA





GGGGS
14,301
PERV_Q4VFZ2_3mutA_WS





GSSGGS
14,302
MLVMS_P03355_3mut





GSSGSS
14,303
MLVAV_P03356_3mutA





GGSPAPGGG
14,304
MLVBM_Q7SVK7_3mutA_WS





GSSGGGGGS
14,305
MLVF5_P26810_3mut





PAPAPAPAP
14,306
MLVCB_P08361_3mut





PAPAP
14,307
PERV_Q4VFZ2_3mutA_WS





PAPGSSGGS
14,308
KORV_Q9TTC1_3mut





PAPGSSGGG
14,309
PERV_Q4VFZ2_3mut





GGGEAAAK
14,310
MLVMS_P03355_PLV919





GGSGGSGGSGGSGGS
14,311
SFV3L_P27401-Pro_2mutA





GGSGGG
14,312
FLV_P10273_3mut





PAPEAAAKGGG
14,313
MLVFF_P26809_3mut





PAP

PERV_Q4VFZ2_3mutA_WS





PAPGGSGSS
14,315
FFV_O93209_2mut





EAAAKEAAAKEAAAKEAA
14,316
FFV_O93209-Pro_2mut


AKEAAAKEAAAK







GSSGSSGSSGSS
14,317
FFV_O93209-Pro





GSSGSSGSSGSSGSS
14,318
FLV_P10273_3mutA





GGGEAAAKPAP
14,319
PERV_Q4VFZ2





PAPGSSGGG
14,320
SFV3L_P27401_2mut





PAPGGSGSS
14,321
KORV_Q9TTC1-Pro_3mut





PAPAPAPAPAP
14,322
GALV_P21414_3mutA





GGSGGGEAAAK
14,323
PERV_Q4VFZ2_3mut





GSSPAP
14,324
MLVCB_P08361_3mut





EAAAKPAP
14,325
MLVF5_P26810_3mut





GGGGGGGGSGGGGSGGGG
14,326
MLVBM_Q7SVK7_3mut


S







GGSGGG
14,327
WMSV_P03359_3mut





GGSGGSGGS
14,328
KORV_Q9TTC1_3mut





GGGGGGGG
14,329
MLVFF_P26809_3mut





GGGGSS
14,330
MLVAV_P03356_3mut





GSSGGGGGS
14,331
SFV3L_P27401_2mut





EAAAKEAAAKEAAAKEAA
14,332
GALV_P21414_3mutA


AKEAAAKEAAAK







GSSGSSGSS
14,333
PERV_Q4VFZ2_3mut





GSSPAPGGS
14,334
MLVFF_P26809_3mut





PAPAPAP
14,335
AVIRE_P03360_3mutA





EAAAKEAAAKEAAAKEAA
14,336
WMSV_P03359_3mutA


AK







PAPAPAPAP
14,337
SFV3L_P27401_2mutA





GGGGSS
14,338
MLVAV_P03356_3mutA





GSSGSSGSSGSSGSS
14,339
SFV3L_P27401_2mutA





PAPGGS
14,340
WMSV_P03359_3mutA





GSSEAAAKGGG
14,341
PERV_Q4VFZ2





GSSGGSPAP
14,342
MLVMS_P03355_PLV919





GSSGSSGSSGSSGSSGSS
14,343
SFV3L_P27401_2mutA





GGSGSSGGG
14,344
MLVCB_P08361_3mut





GGGPAPGSS
14,345
SFV3L_P27401-Pro_2mutA





GSSEAAAKGGS
14,346
WMSV_P03359_3mut





GSSEAAAKGGG
14,347
MLVAV_P03356_3mut





GGSGGGPAP
14,348
FFV_O93209-Pro





GSSGSS
14,349
PERV_Q4VFZ2_3mut





PAPGGGGGS
14,350
GALV_P21414_3mutA





EAAAKPAPGGS
14,351
MLVAV_P03356_3mut





GSSGSS
14,352
MLVMS_P03355_3mut





EAAAKPAPGGS
14,353
FFV_O93209-Pro





GGGPAPEAAAK
14,354
MLVMS_P03355_3mutA_WS





GSSEAAAKGGG
14,355
MLVBM_Q7SVK7_3mut





GGGEAAAKGGS
14,356
BAEVM_P10272_3mut





GSSGSS
14,357
KORV_Q9TTC1-Pro_3mutA





EAAAKEAAAKEAAAK
14,358
SFV1_P23074





PAPGSSGGS
14,359
KORV_Q9TTC1-Pro_3mut





PAPAPAPAPAP
14,360
MLVMS_P03355





GSSEAAAK
14,361
SFV3L_P27401_2mut





PAP

PERV_Q4VFZ2_3mut





GGSEAAAKGGG
14,363
MLVBM_Q7SVK7_3mut





GGSGGGPAP
14,364
MLVBM_Q7SVK7_3mutA_WS





GSSGSS
14,365
MLVMS_P03355_3mut





GGSEAAAK
14,366
MLVMS_P03355





GSSEAAAKGGS
14,367
MLVMS_P03355_PLV919





PAPGGGGGS
14,368
MLVFF_P26809_3mut





GSSGGG
14,369
PERV_Q4VFZ2_3mut





GSSGGS
14,370
PERV_Q4VFZ2_3mutA_WS





PAPGGG
14,371
BAEVM_P10272_3mut





PAPGSSGGG
14,372
MLVBM_Q7SVK7_3mut





GGSEAAAK
14,373
SFV3L_P27401_2mut





GSSPAPEAAAK
14,374
SFV3L_P27401-Pro_2mut





GSSGGSPAP
14,375
BAEVM_P10272_3mut





GGSPAPGSS
14,376
PERV_Q4VFZ2_3mutA_WS





GGSGGSGGS
14,377
PERV_Q4VFZ2





GGSGGGPAP
14,378
FLV_P10273_3mut





GGGPAPEAAAK
14,379
SFV3L_P27401_2mutA





GGGGS
14,380
FLV_P10273_3mutA





GSSGGSGGG
14,381
XMRV6_A1Z651_3mut





EAAAKGGGGSS
14,382
PERV_Q4VFZ2





GGSGSSGGG
14,383
SFV3L_P27401-Pro_2mutA





GGSGGSGGS
14,384
MLVFF_P26809_3mut





GGGPAPEAAAK
14,385
FLV_P10273_3mut





GSSGGGEAAAK
14,386
MLVMS_P03355_3mut





GGG

SFV3L_P27401_2mut





GSAGSAAGSGEF
14,388
WMSV_P03359_3mut





GSSGGGPAP
14,389
MLVMS_P03355_PLV919





GGGGSS
14,390
KORV_Q9TTC1-Pro_3mut





GGGGSSEAAAK
14,391
KORV_Q9TTC1





PAPGGSGGG
14,392
SFV3L_P27401_2mut





GSSGSSGSSGSSGSS
14,393
FFV_O93209





GSSGGSPAP
14,394
MLVMS_P03355_3mut





GGSEAAAK
14,395
KORV_Q9TTC1-Pro_3mutA





GGGGSGGGGS
14,396
BAEVM_P10272_3mut





GSSEAAAKGGG
14,397
AVIRE_P03360_3mut





EAAAKPAPGGG
14,398
FLV_P10273_3mut





EAAAKGGSPAP
14,399
SFV3L_P27401-Pro_2mutA





GSSEAAAKPAP
14,400
MLVBM_Q7SVK7_3mut





GGGPAPGGS
14,401
MLVCB_P08361_3mut





GGG

SFV3L_P27401_2mutA





EAAAKGGGGSEAAAK
14,403
SFV3L_P27401_2mutA





GGSGSSGGG
14,404
MLVBM_Q7SVK7_3mut





GSAGSAAGSGEF
14,405
BAEVM_P10272_3mut





GGGEAAAK
14,406
FOAMV_P14350_2mutA





PAPEAAAKGGS
14,407
WMSV_P03359_3mut





PAPAPAPAPAPAP
14,408
MLVF5_P26810_3mutA





GGSGGGGSS
14,409
FLV_P10273_3mutA





PAPGSSGGS
14,410
BAEVM_P10272_3mut





PAPEAAAK
14,411
WMSV_P03359_3mutA





GSSGSSGSSGSSGSSGSS
14,412
FFV_O93209-Pro_2mut





GGGGGSGSS
14,413
FFV_O93209-Pro





GGGGGGGG
14,414
SFV3L_P27401-Pro_2mutA





GGGGGG
14,415
FLV_P10273_3mut





GSSGGSGGG
14,416
MLVAV_P03356_3mutA





GGGGSS
14,417
SFV3L_P27401-Pro_2mutA





GGSGGGPAP
14,418
FOAMV_P14350_2mut





GSSGSS
14,419
AVIRE_P03360_3mutA





EAAAKEAAAKEAAAKEAA
14,420
SFV3L_P27401-Pro_2mutA


AKEAAAK







EAAAKEAAAK
14,421
BAEVM_P10272_3mut





GSSPAPEAAAK
14,422
GALV_P21414_3mutA





GGSEAAAKPAP
14,423
SFV3L_P27401_2mutA





GGSGGGEAAAK
14,424
SFV3L_P27401-Pro_2mutA





EAAAKGSSPAP
14,425
FOAMV_P14350_2mut





GGSGSSEAAAK
14,426
SFV3L_P27401_2mut





GGG

PERV_Q4VFZ2





GGGGGSGSS
14,428
FOAMV_P14350_2mut





GGSGGGEAAAK
14,429
KORV_Q9TTC1-Pro_3mut





GSSGGSGGG
14,430
AVIRE_P03360_3mutA





EAAAKPAPGGG
14,431
SFV3L_P27401_2mutA





PAPGGSGGG
14,432
KORV_Q9TTC1-Pro_3mut





PAPAPAP
14,433
WMSV_P03359_3mutA





GSSEAAAKPAP
14,434
SFV1_P23074





SGGSSGGSSGSETPGTSE
14,435
SRV2_P51517


SATPESSGGSSGGSS







GSSGGSGGG
14,436
PERV_Q4VFZ2_3mutA_WS





GSSGSSGSSGSSGSSGSS
14,437
FFV_O93209





GSSGGGPAP
14,438
WMSV_P03359_3mut





PAPAPAPAPAPAP
14,439
MLVBM_Q7SVK7_3mut





GGGGGSPAP
14,440
KORV_Q9TTC1-Pro_3mutA





PAPGSS
14,441
MLVBM_Q7SVK7_3mutA_WS





PAPEAAAKGGS
14,442
SFV3L_P27401-Pro_2mut





GGGGSSPAP
14,443
MLVMS_P03355_3mut





GGSEAAAK
14,444
FFV_O93209-Pro





EAAAKPAPGGS
14,445
AVIRE_P03360_3mutA





PAPGSS
14,446
WMSV_P03359_3mut





PAPGSSGGG
14,447
SFV3L_P27401-Pro_2mutA





EAAAKEAAAKEAAAK
14,448
SFV3L_P27401_2mut





GGS

MLVRD_P11227_3mut





GGGGS
14,450
KORV_Q9TTC1-Pro_3mut





GGSGGGGSS
14,451
KORV_Q9TTC1





GGSGGG
14,452
MLVMS_P03355_3mutA_WS





GGGEAAAKPAP
14,453
BAEVM_P10272_3mut





EAAAKEAAAKEAAAKEAA
14,454
FLV_P10273


AKEAAAK







PAPGGSGGG
14,455
KORV_Q9TTC1-Pro_3mutA





GSSGSSGSSGSSGSSGSS
14,456
HTL1L_P0C211





GGGEAAAKPAP
14,457
WMSV_P03359





GSSGGSPAP
14,458
FFV_O93209-Pro





PAPAPAPAPAP
14,459
SFV3L_P27401-Pro_2mutA





GSSGGSEAAAK
14,460
SFV3L_P27401_2mutA





GGSPAPGSS
14,461
SFV3L_P27401_2mut





GGSGGSGGS
14,462
KORV_Q9TTC1-Pro_3mut





PAPEAAAKGSS
14,463
KORV_Q9TTC1-Pro_3mut





EAAAKGGS
14,464
KORV_Q9TTC1_3mutA





EAAAKGGGGSEAAAK
14,465
SFV3L_P27401-Pro_2mut





GGGGSSPAP
14,466
FFV_O93209-Pro





EAAAK
14,467
SFV3L_P27401_2mut





EAAAKGGGGSS
14,468
BAEVM_P10272_3mut





GGGGGSEAAAK
14,469
MLVBM_Q7SVK7_3mut





GGGG
14,470
PERV_Q4VFZ2





GGGGGSEAAAK
14,471
FLV_P10273_3mut





EAAAKGGGPAP
14,472
KORV_Q9TTC1-Pro





GGGGSGGGGSGGGGSGGG
14,473
FFV_O93209_2mutA


GS







GSSGGSGGG
14,474
PERV_Q4VFZ2_3mut





GGGGSGGGGSGGGGS
14,475
GALV_P21414_3mutA





GGSGGGEAAAK
14,476
AVIRE_P03360_3mutA





PAPEAAAKGGG
14,477
SFV3L_P27401_2mut





GGGGSGGGGS
14,478
AVIRE_P03360





GSSGGGEAAAK
14,479
SFV3L_P27401_2mutA





GGGGG
14,480
AVIRE_P03360_3mutA





GGSGSS
14,481
KORV_Q9TTC1_3mut





PAPAPAPAPAPAP
14,482
FOAMV_P14350_2mut





GGSEAAAKPAP
14,483
KORV_Q9TTC1-Pro_3mut





GGGGGG
14,484
PERV_Q4VFZ2_3mut





GSSGGGEAAAK
14,485
MLVBM_Q7SVK7





SGGSSGGSSGSETPGTSE
14,486
MLVAV_P03356


SATPESSGGSSGGSS







GGSPAPGSS
14,487
BAEVM_P10272_3mut





GGGGSSPAP
14,488
BAEVM_P10272





GGGGSEAAAKGGGGS
14,489
SFV3L_P27401_2mut





GGGGGGGG
14,490
GALV_P21414_3mutA





PAPAP
14,491
MLVAV_P03356_3mut





GGGEAAAK
14,492
PERV_Q4VFZ2_3mutA_WS





GSSPAPGGG
14,493
FFV_O93209_2mut





GGSGGSGGSGGSGGS
14,494
BAEVM_P10272





GGGGGS
14,495
MLVF5_P26810_3mutA





PAPGGGGSS
14,496
FLV_P10273_3mutA





GGGEAAAK
14,497
MLVBM_Q7SVK7_3mut





PAPEAAAKGGG
14,498
WMSV_P03359_3mut





GSSEAAAK
14,499
MLVBM_Q7SVK7_3mut





EAAAKEAAAK
14,500
AVIRE_P03360





EAAAKGGGGGS
14,501
MLVBM_Q7SVK7_3mut





GGGEAAAKGGS
14,502
SFV3L_P27401-Pro_2mutA





PAPAPAPAPAP
14,503
MLVF5_P26810_3mut





PAPGSSEAAAK
14,504
SFV3L_P27401-Pro_2mutA





EAAAKEAAAKEAAAK
14,505
BAEVM_P10272_3mutA





GGSPAPGSS
14,506
MLVMS_P03355





PAPGSSGGS
14,507
FLV_P10273_3mutA





EAAAKEAAAKEAAAKEAA
14,508
FOAMV_P14350-Pro_2mut


AK







EAAAKGGG
14,509
KORV_Q9TTC1_3mutA





EAAAKGGSGGG
14,510
MLVBM_Q7SVK7_3mut





GGGGGS
14,511
KORV_Q9TTC1-Pro_3mutA





PAPGGSGGG
14,512
WMSV_P03359_3mut





GGGPAPGGS
14,513
KORV_Q9TTC1_3mutA





GSS

FFV_O93209





GGSGGSGGS
14,515
PERV_Q4VFZ2_3mut





GGGGS
14,516
GALV_P21414_3mutA





GGGG
14,517
MLVF5_P26810_3mut





GGSEAAAKPAP
14,518
FFV_O93209-Pro_2mut





PAPAPAPAP
14,519
FFV_O93209-Pro





PAP

MLVF5_P26810_3mut





EAAAKEAAAKEAAAK
14,521
FFV_O93209_2mut





EAAAKGSS
14,522
MLVCB_P08361_3mut





EAAAKGGG
14,523
MLVBM_Q7SVK7_3mut





PAPEAAAKGGG
14,524
FFV_O93209_2mut





GSSGGGEAAAK
14,525
SFV1_P23074-Pro_2mut





PAPGGGEAAAK
14,526
GALV_P21414_3mutA





GGGGSGGGGSGGGGSGGG
14,527
FOAMV_P14350-Pro_2mutA


GS







GSSGGG
14,528
FOAMV_P14350_2mut





GGGGSGGGGGGGGSGGGG
14,529
SFV3L_P27401_2mutA


S







GGSGSS
14,530
AVIRE_P03360_3mut





GGSGSSEAAAK
14,531
MMTVB_P03365_WS





PAPAPAP
14,532
MLVAV_P03356_3mutA





GSSGGSPAP
14,533
SFV3L_P27401-Pro_2mut





GGSPAP
14,534
AVIRE_P03360





GGSGGGPAP
14,535
FFV_O93209





GSSEAAAK
14,536
PERV_Q4VFZ2





GSSGGGPAP
14,537
PERV_Q4VFZ2_3mutA_WS





GGGGSSEAAAK
14,538
KORV_Q9TTC1_3mutA





GGSEAAAKPAP
14,539
SFVCP_Q87040





GGSGGGPAP
14,540
FOAMV_P14350_2mutA





GGGGSGGGGSGGGGSGGG
14,541
BLVJ_P03361_2mutB


GS







GGGGSSPAP
14,542
SFV3L_P27401_2mutA





EAAAKGGS
14,543
MLVF5_P26810_3mut





GGSEAAAKGSS
14,544
MLVCB_P08361_3mut





GGGGSSEAAAK
14,545
SFV3L_P27401_2mut





EAAAKGGSGGG
14,546
FOAMV_P14350_2mut





GGSGGS
14,547
FLV_P10273_3mut





EAAAKGGG
14,548
FFV_O93209-Pro





GSSGSSGSSGSSGSS
14,549
SFV3L_P27401





GSSGGGPAP
14,550
PERV_Q4VFZ2_3mutA_WS





PAPGGSEAAAK
14,551
SFV3L_P27401-Pro_2mutA





GGSPAP
14,552
KORV_Q9TTC1





EAAAKPAPGSS
14,553
KORV_Q9TTC1_3mutA





SGSETPGTSESATPES
14,554
SFV1_P23074





GSSPAP
14,555
SFV3L_P27401-Pro_2mutA





GSSPAPGGG
14,556
SFV3L_P27401_2mut





GGGEAAAKGSS
14,557
SFV1_P23074_2mut





GGGPAPGGS
14,558
BAEVM_P10272_3mut





EAAAKGGG
14,559
KORV_Q9TTC1-Pro_3mutA





GSSGGG
14,560
SFV3L_P27401-Pro_2mut





GGSPAPEAAAK
14,561
BAEVM_P10272_3mut





EAAAKGSSPAP
14,562
FFV_O93209





EAAAKGGGGSEAAAK
14,563
SFV3L_P27401-Pro_2mutA





GSSGSSGSSGSSGSS
14,564
SFV1_P23074_2mut





EAAAKGGSPAP
14,565
FOAMV_P14350_2mut





GGSGGS
14,566
KORV_Q9TTC1-Pro_3mutA





EAAAKGSSGGS
14,567
GALV_P21414





GSSGGGPAP
14,568
MLVAV_P03356





PAPEAAAKGGS
14,569
FOAMV_P14350_2mut





EAAAKPAPGGG
14,570
AVIRE_P03360_3mut





GGSPAP
14,571
SFV3L_P27401_2mutA





GGGGSGGGGS
14,572
SFV3L_P27401_2mutA





GGGGSS
14,573
AVIRE_P03360_3mutA





GGSPAPGGG
14,574
SFV3L_P27401-Pro_2mutA





EAAAKPAPGSS
14,575
SFV3L_P27401





EAAAKPAP
14,576
FOAMV_P14350-Pro_2mut





PAPEAAAKGSS
14,577
PERV_Q4VFZ2_3mutA_WS





EAAAKGGSGSS
14,578
SFV3L_P27401_2mutA





GGGEAAAKGSS
14,579
GALV_P21414_3mutA





GGGGSEAAAKGGGGS
14,580
PERV_Q4VFZ2_3mut





PAPGGSGSS
14,581
FFV_O93209-Pro_2mutA





GGSEAAAKPAP
14,582
GALV_P21414_3mutA





GGSGGSGGSGGSGGS
14,583
FFV_O93209-Pro





GSSGGSEAAAK
14,584
SFV3L_P27401-Pro_2mut





GGS

GALV_P21414_3mutA





PAPGGSEAAAK
14,586
MLVMS_P03355





PAPEAAAKGGS
14,587
BAEVM_P10272_3mutA





GGSGSSPAP
14,588
SFV3L_P27401-Pro_2mutA





GSSPAP
14,589
WMSV_P03359_3mut





GGGEAAAK
14,590
MMTVB_P03365





GGGGSS
14,591
PERV_Q4VFZ2_3mut





GGSPAPGSS
14,592
SFV3L_P27401-Pro_2mut





PAPGGS
14,593
MLVBM_Q7SVK7_3mut





EAAAKGSSPAP
14,594
MLVBM_Q7SVK7_3mut





GGGGSSGGS
14,595
PERV_Q4VFZ2_3mut





PAPAPAPAPAPAP
14,596
SFV1_P23074





GGSEAAAKGGG
14,597
SFV3L_P27401-Pro_2mut





GGSGGS
14,598
SFV1_P23074_2mut





GSSGGGGGS
14,599
MLVF5_P26810_3mutA





EAAAKGGGPAP
14,600
SFV3L_P27401





EAAAKEAAAKEAAAKEAA
14,601
FOAMV_P14350-Pro_2mutA


AK







GGGPAPGSS
14,602
SFV3L_P27401_2mutA





GGGGSGGGGSGGGGSGGG
14,603
SFV3L_P27401_2mut


GS







EAAAKEAAAKEAAAKEAA
14,604
MMTVB_P03365_WS


AK







PAPGSSGGS
14,605
KORV_Q9TTC1-Pro_3mutA





PAPGSSEAAAK
14,606
FOAMV_P14350-Pro_2mut





GSSPAPEAAAK
14,607
BAEVM_P10272_3mut





EAAAKGGGGSEAAAK
14,608
FFV_O93209-Pro





GGSPAP
14,609
PERV_Q4VFZ2





GGSGSSEAAAK
14,610
XMRV6_A1Z651_3mut





GGSEAAAKGGG
14,611
GALV_P21414_3mutA





PAPGGGGSS
14,612
AVIRE_P03360_3mutA





GGSGGSGGSGGS
14,613
PERV_Q4VFZ2





GGGGSSGGS
14,614
PERV_Q4VFZ2_3mutA_WS





SGGSSGGSSGSETPGTSE
14,615
BAEVM_P10272_3mutA


SATPESSGGSSGGSS







GGGPAP
14,616
MLVAV_P03356_3mut





GGGGSGGGGSGGGGSGGG
14,617
FFV_O93209_2mut


GS







GSSEAAAK
14,618
FFV_O93209





GGSPAPEAAAK
14,619
FOAMV_P14350_2mut





GGGGGSEAAAK
14,620
FOAMV_P14350_2mut





GSSPAPGGS
14,621
MLVBM_Q7SVK7_3mut





GSS

SFVCP_Q87040_2mut





EAAAKPAP
14,623
FOAMV_P14350-Pro





EAAAKGGG
14,624
SFV3L_P27401_2mut





GGGEAAAK
14,625
AVIRE_P03360_3mutA





PAPGSSGGG
14,626
WMSV_P03359_3mut





EAAAKGGSPAP
14,627
SFV3L_P27401





GSSGGSGGG
14,628
SFV3L_P27401-Pro_2mutA





GSSGGGEAAAK
14,629
GALV_P21414_3mutA





GGGPAPGSS
14,630
MLVBM_Q7SVK7_3mutA_WS





PAPGGGEAAAK
14,631
FFV_O93209-Pro_2mut





GSSGSSGSSGSS
14,632
SFV1_P23074_2mut





GGSEAAAK
14,633
PERV_Q4VFZ2_3mutA_WS





GGGEAAAKPAP
14,634
SFV3L_P27401_2mut





EAAAKGGGPAP
14,635
SFV3L_P27401_2mut





GGGGSSPAP
14,636
FLV_P10273_3mut





EAAAKPAPGSS
14,637
FFV_O93209_2mut





GGGGSSPAP
14,638
SFV3L_P27401_2mut





GSSGSS
14,639
KORV_Q9TTC1_3mutA





GGGGSGGGGSGGGGSGGG
14,640
BLVJ_P03361_2mut


GSGGGGS







GGGGSSGGS
14,641
GALV_P21414_3mutA





EAAAKGGSGSS
14,642
FFV_O93209-Pro





EAAAKPAP
14,643
PERV_Q4VFZ2





GSSGGGEAAAK
14,644
MLVBM_Q7SVK7_3mut





PAPGGSGGG
14,645
BAEVM_P10272





EAAAKGGGPAP
14,646
MLVF5_P26810





GSSGSSGSS
14,647
MLVBM_Q7SVK7_3mut





GSSGGS
14,648
AVIRE_P03360_3mutA





GGSEAAAKGGG
14,649
FOAMV_P14350_2mut





EAAAKGGS
14,650
MLVF5_P26810_3mutA





GGSGSSGGG
14,651
WMSV_P03359_3mut





EAAAK
14,652
SFV1_P23074_2mut





GSSGGSPAP
14,653
SFV3L_P27401-Pro_2mutA





GGGGSSGGS
14,654
KORV_Q9TTC1_3mut





PAPGGSGGG
14,655
FFV_O93209-Pro_2mut





GGGPAPGGS
14,656
SFV3L_P27401_2mutA





GSSPAPEAAAK
14,657
FLV_P10273_3mut





GGSGSSPAP
14,658
SFV3L_P27401_2mut





GSSEAAAKGGS
14,659
SFV3L_P27401_2mut





PAPGGG
14,660
SFV3L_P27401_2mutA





SGSETPGTSESATPES
14,661
KORV_Q9TTC1-Pro_3mut





GGGGS
14,662
SFV1_P23074-Pro_2mutA





GSSGGGEAAAK
14,663
WMSV_P03359





EAAAKGGGGSEAAAK
14,664
MLVF5_P26810_3mutA





GSSEAAAKPAP
14,665
FFV_O93209





GGGGGG
14,666
SFV1_P23074_2mutA





EAAAKEAAAKEAAAK
14,667
MMTVB_P03365-Pro





EAAAKPAPGSS
14,668
MLVBM_Q7SVK7_3mut





GGSGSSEAAAK
14,669
SFV3L_P27401_2mutA





GGSEAAAK
14,670
MLVMS_P03355_3mut





GGSPAPEAAAK
14,671
SFV3L_P27401_2mut





GGGPAPGSS
14,672
SFV1_P23074





GGGGGSEAAAK
14,673
MLVBM_Q7SVK7_3mutA_WS





EAAAKPAPGSS
14,674
KORV_Q9TTC1-Pro





GSSGSSGSSGSS
14,675
SFV3L_P27401_2mut





EAAAKPAP
14,676
SFV3L_P27401_2mut





GGGEAAAK
14,677
PERV_Q4VFZ2_3mut





GGSGGS
14,678
SFV3L_P27401_2mutA





EAAAKGSSGGS
14,679
MMTVB_P03365





SGSETPGTSESATPES
14,680
SFV3L_P27401





EAAAKGSSGGG
14,681
PERV_Q4VFZ2





EAAAKEAAAKEAAAKEAA
14,682
MMTVB_P03365


AKEAAAKEAAAK







GGSGGGPAP
14,683
KORV_Q9TTC1_3mutA





PAPAPAPAP
14,684
SFV3L_P27401





GGGEAAAKGGS
14,685
SFV1_P23074_2mut





GSSGGSGGG
14,686
PERV_Q4VFZ2_3mut





PAPEAAAKGGS
14,687
FOAMV_P14350_2mutA





GGGEAAAKGSS
14,688
SFV3L_P27401_2mut





GGGGGGGGSGGGGSGGGG
14,689
MLVBM_Q7SVK7


S







PAPGSSGGG
14,690
FLV_P10273





GGSGSSGGG
14,691
FFV_O93209





EAAAKPAPGSS
14,692
MLVBM_Q7SVK7





GSSEAAAKGGG
14,693
SFV3L_P27401_2mutA





GGSGGSGGSGGSGGS
14,694
MLVF5_P26810





GGSEAAAKPAP
14,695
SFV3L_P27401-Pro_2mutA





EAAAKGGSPAP
14,696
SFV3L_P27401_2mutA





EAAAKGGGGGS
14,697
SFV3L_P27401_2mut





GSSPAPEAAAK
14,698
SFV3L_P27401_2mutA





PAPAP
14,699
MLVBM_Q7SVK7_3mut





PAPGGSEAAAK
14,700
KORV_Q9TTC1-Pro





GGSGSS
14,701
MLVF5_P26810_3mutA





GGSEAAAKPAP
14,702
FFV_O93209_2mut





GSS

MLVMS_P03355





SGGSSGGSSGSETPGTSE
14,704
SFV3L_P27401-Pro


SATPESSGGSSGGSS







PAPGGGEAAAK
14,705
SFV3L_P27401_2mut





PAPGGGGGS
14,706
SFV3L_P27401-Pro_2mut





PAPGGSGSS
14,707
BAEVM_P10272_3mut





GSSGGGEAAAK
14,708
FFV_O93209





GGSEAAAKPAP
14,709
SFV1_P23074_2mut





GGGGG
14,710
FLV_P10273_3mut





GGGEAAAKGSS
14,711
SFV3L_P27401





GSSGSSGSSGSSGSS
14,712
SFV1_P23074-Pro





SGSETPGTSESATPES
14,713
AVIRE_P03360





PAPGSSGGG
14,714
MLVBM_Q7SVK7_3mut





GGGGSSPAP
14,715
HTL3P_Q4U0X6_2mut





GGGEAAAK
14,716
SFV1_P23074





GGSGGG
14,717
AVIRE_P03360





EAAAKGSSGGG
14,718
SFV3L_P27401_2mutA





GSSPAPEAAAK
14,719
FOAMV_P14350-Pro_2mutA





GGGPAPGSS
14,720
WMSV_P03359





EAAAKGSSGGG
14,721
MLVMS_P03355





GGGGGSEAAAK
14,722
MLVMS_P03355





EAAAKPAPGGS
14,723
SFV3L_P27401





EAAAKGSSPAP
14,724
SFV3L_P27401





GGGGGGG
14,725
FOAMV_P14350_2mutA





EAAAKEAAAKEAAAK
14,726
SFV3L_P27401





GSSPAPGGS
14,727
FFV_O93209_2mutA





GGGGSSEAAAK
14,728
SFV3L_P27401-Pro_2mutA





GGSEAAAKGSS
14,729
GALV_P21414_3mutA





GGSEAAAKGSS
14,730
BAEVM_P10272_3mutA





EAAAKPAPGGG
14,731
MLVCB_P08361





GSSGSSGSSGSSGSSGSS
14,732
SFV1_P23074-Pro





GGGGSEAAAKGGGGS
14,733
FOAMV_P14350_2mut





GSSPAPGGS
14,734
MLVMS_P03355_PLV919





GGGGSGGGGS
14,735
FFV_O93209-Pro





GSSGGSPAP
14,736
KORV_Q9TTC1_3mutA





GGSGGS
14,737
GALV_P21414_3mutA





PAPGSSEAAAK
14,738
WMSV_P03359





PAPGGGGSS
14,739
MMTVB_P03365-Pro





GGGGSSGGS
14,740
PERV_Q4VFZ2_3mutA_WS





GGGGSGGGGS
14,741
FFV_O93209_2mut





GGGGSGGGGSGGGGSGGG
14,742
XMRV6_A1Z651


GS







GGSGSSEAAAK
14,743
SFV1_P23074_2mut





GGSGGGGSS
14,744
GALV_P21414_3mutA





GGSEAAAKPAP
14,745
MLVBM_Q7SVK7





EAAAKGGSPAP
14,746
SFV1_P23074_2mutA





PAPAPAPAP
14,747
FFV_O93209





GSSGGSPAP
14,748
MMTVB_P03365-Pro





GGGGGSPAP
14,749
KORV_Q9TTC1_3mutA





EAAAKGGGPAP
14,750
PERV_Q4VFZ2





GSSGGSPAP
14,751
BAEVM_P10272





GGGGG
14,752
FFV_O93209





GGGGGS
14,753
FLV_P10273_3mutA





EAAAKEAAAKEAAAK
14,754
FOAMV_P14350





PAPGGG
14,755
MLVCB_P08361_3mut





GSSGGSEAAAK
14,756
FOAMV_P14350_2mutA





GGSPAPGGG
14,757
FLV_P10273_3mut





GSSGSSGSSGSSGSSGSS
14,758
SFV1_P23074-Pro_2mutA





GGSPAPEAAAK
14,759
SFV3L_P27401





PAPGGGGSS
14,760
HTL3P_Q4U0X6_2mutB





GGGGSSEAAAK
14,761
MMTVB_P03365_2mut_WS





PAPGGS
14,762
MLVRD_P11227_3mut





GGSGGSGGSGGSGGS
14,763
MMTVB_P03365





GSAGSAAGSGEF
14,764
AVIRE_P03360





GSSGGS
14,765
BAEVM_P10272_3mutA





GGSGGGGSS
14,766
MMTVB_P03365





GGSGGGGSS
14,767
WMSV_P03359





PAPEAAAKGSS
14,768
SFV1_P23074





GSSGSSGSSGSS
14,769
SFV1_P23074-Pro_2mutA





PAPAPAPAPAPAP
14,770
SFV3L_P27401





PAPGSSGGG
14,771
FLV_P10273_3mut





GGSGSSPAP
14,772
MLVMS_P03355





GGSGGGPAP
14,773
FOAMV_P14350





PAPGGGGGS
14,774
KORV_Q9TTC1_3mutA





EAAAKGSSPAP
14,775
GALV_P21414_3mutA





GGSGSSPAP
14,776
MLVBM_Q7SVK7_3mut





EAAAKGSS
14,777
SFV3L_P27401_2mut





GGGGGSEAAAK
14,778
WMSV_P03359





GGGGGGGG
14,779
SFV1_P23074-Pro





EAAAKEAAAK
14,780
MLVBM_Q7SVK7





GGGEAAAKGGS
14,781
MLVBM_Q7SVK7





EAAAKGGSPAP
14,782
SFV3L_P27401_2mut





GSSEAAAK
14,783
XMRV6_A1Z651





PAPGGGEAAAK
14,784
MMTVB_P03365_WS





GGSPAP
14,785
GALV_P21414_3mutA





GSSPAPGGG
14,786
MLVBM_Q7SVK7_3mutA_WS





GGSGSSPAP
14,787
SFV1_P23074_2mutA





GGS

HTL32_Q0R5R2_2mut





GGSGGGGSS
14,789
MMTVB_P03365-Pro





GGGGSGGGGSGGGGSGGG
14,790
SFVCP_Q87040_2mutA


GS







EAAAKGGGPAP
14,791
FOAMV_P14350_2mut





GSSGGGEAAAK
14,792
MMTVB_P03365





SGGSSGGSSGSETPGTSE
14,793
MLVBM_Q7SVK7_3mutA_WS


SATPESSGGSSGGSS







AEAAAKEAAAKEAAAKEA
14,794
MMTVB_P03365_WS


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







EAAAKEAAAK
14,795
FOAMV_P14350-Pro_2mut





GSSPAPEAAAK
14,796
FOAMV_P14350_2mutA





EAAAKPAPGGS
14,797
GALV_P21414_3mutA





GSSGGSPAP
14,798
KORV_Q9TTC1-Pro_3mut





GGGPAPEAAAK
14,799
MLVAV_P03356





GGGEAAAKPAP
14,800
SFV1_P23074-Pro_2mut





GGGGGSEAAAK
14,801
SFV3L_P27401_2mut





GGGPAPGSS
14,802
SFV3L_P27401_2mut





GGSEAAAKPAP
14,803
AVIRE_P03360





GSSGSSGSSGSSGSSGSS
14,804
SFV1_P23074-Pro_2mut





EAAAKGSSGGS
14,805
FOAMV_P14350_2mutA





GGGGGG
14,806
MLVBM_Q7SVK7_3mut





GSSPAPGGS
14,807
PERV_Q4VFZ2





GGSGSSPAP
14,808
GALV_P21414_3mutA





GGGPAPEAAAK
14,809
SFV3L_P27401





GGSGGGEAAAK
14,810
WMSV_P03359





GSAGSAAGSGEF
14,811
SFV1_P23074_2mut





GSSGGGEAAAK
14,812
MLVMS_P03355





GGG

MMTVB_P03365-Pro





PAPGSSGGS
14,814
FOAMV_P14350_2mut





GGGGSSPAP
14,815
FFV_O93209_2mut





SGGSSGGSSGSETPGTSE
14,816
MMTVB_P03365_WS


SATPESSGGSSGGSS







GGGGGGG
14,817
XMRV6_A1Z651





PAPAPAPAPAP
14,818
FOAMV_P14350





GGGGSGGGGSGGGGSGGG
14,819
MMTVB_P03365_2mut_WS


GS







GGSGGGPAP
14,820
SFV3L_P27401_2mut





GGGGGG
14,821
SFV1_P23074-Pro





EAAAKPAPGSS
14,822
SFV3L_P27401_2mut





GGGGSSGGS
14,823
HTL3P_Q4U0X6_2mut





PAPGSSEAAAK
14,824
MMTVB_P03365-Pro





GGGGSSPAP
14,825
FOAMV_P14350-Pro_2mut





PAPGSSGGS
14,826
MMTVB_P03365





AEAAAKEAAAKEAAAKEA
14,827
SRV2_P51517


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPAPAP
14,828
MMTVB_P03365_2mut_WS





PAPGGGGGS
14,829
MMTVB_P03365_2mutB





GGGGSS
14,830
SFV1_P23074-Pro_2mutA





EAAAKEAAAKEAAAKEAA
14,831
SFV3L_P27401-Pro


AK







GGSGGSGGSGGSGGS
14,832
MMTVB_P03365-Pro





GGGGGGG
14,833
SFV3L_P27401_2mut





PAPGGGEAAAK
14,834
SFV3L_P27401





PAPGSS
14,835
FOAMV_P14350_2mutA





GGGGSGGGGS
14,836
SFVCP_Q87040_2mutA





GSSGGSGGG
14,837
XMRV6_A1Z651





GGGGSGGGGSGGGGSGGG
14,838
MLVBM_Q7SVK7


GSGGGGSGGGGS







GSSEAAAKGGG
14,839
FFV_O93209-Pro_2mut





GGSEAAAKPAP
14,840
SFV3L_P27401-Pro





GSSGGSGGG
14,841
SFV1_P23074_2mut





EAAAKGGGGSS
14,842
FOAMV_P14350_2mutA





GGGGGG
14,843
SFV3L_P27401_2mut





GGGGG
14,844
MLVBM_Q7SVK7_3mut





PAPEAAAKGGG
14,845
SFV3L_P27401





EAAAKGGSPAP
14,846
KORV_Q9TTC1_3mutA





GGGEAAAKPAP
14,847
SFV1_P23074_2mut





GSSGSSGSSGSSGSSGSS
14,848
KORV_Q9TTC1-Pro





EAAAKEAAAKEAAAKEAA
14,849
SFVCP_Q87040


AK







PAPGSSEAAAK
14,850
MLVBM_Q7SVK7





GSSGSSGSS
14,851
FFV_O93209-Pro_2mut





GSSGGGPAP
14,852
SFV3L_P27401-Pro_2mut





GGGPAPEAAAK
14,853
WMSV_P03359_3mut





GGGEAAAK
14,854
MMTVB_P03365-Pro





GSSGSSGSSGSS
14,855
SFV3L_P27401-Pro_2mutA





PAPAPAPAPAP
14,856
FFV_O93209-Pro





GGSPAPEAAAK
14,857
FFV_O93209-Pro_2mut





GSSGSSGSSGSSGSSGSS
14,858
GALV_P21414





EAAAKEAAAKEAAAKEAA
14,859
FOAMV_P14350


AKEAAAK







GGGPAPEAAAK
14,860
MMTVB_P03365-Pro





PAPGGSGGG
14,861
MLVF5_P26810_3mutA





PAPGGSGGG
14,862
FLV_P10273_3mut





GGGEAAAKGGS
14,863
SFV3L_P27401





GSAGSAAGSGEF
14,864
MLVBM_Q7SVK7_3mut





GSSPAPGGG
14,865
MPMV_P07572_2mutB





GSSGSSGSSGSSGSSGSS
14,866
FOAMV_P14350





GGSGGGGSS
14,867
BLVJ_P03361_2mut





PAPEAAAKGSS
14,868
SFV1_P23074-Pro





GGG

FFV_O93209





EAAAKGGGGSS
14,870
SFV1_P23074_2mut





EAAAKEAAAKEAAAKEAA
14,871
SRV2_P51517


AKEAAAKEAAAK







GGGGSGGGGSGGGGSGGG
14,872
MMTVB_P03365


GSGGGGSGGGGS







GGGEAAAKGGS
14,873
MMTVB_P03365_WS





GSSGSS
14,874
SFV1_P23074





GSSGGGGGS
14,875
SFV3L_P27401





GGGGSSEAAAK
14,876
SFV1_P23074





EAAAKGSSGGS
14,877
HTL1A_P03362_2mutB





GSSEAAAKGGS
14,878
GALV_P21414_3mutA





EAAAKGSSPAP
14,879
SFV1_P23074





EAAAKPAPGSS
14,880
SFV3L_P27401_2mutA





PAPGSSGGG
14,881
SFV3L_P27401-Pro_2mut





GGGGSGGGGSGGGGSGGG
14,882
SFV3L_P27401-Pro


GSGGGGSGGGGS







EAAAKEAAAKEAAAKEAA
14,883
MMTVB_P03365_WS


AKEAAAK







GGGGSSEAAAK
14,884
MLVF5_P26810_3mutA





EAAAKGGSPAP
14,885
GALV_P21414





PAPEAAAKGSS
14,886
MMTVB_P03365_WS





GSSGGGGGS
14,887
SFVCP_Q87040_2mut





GGGGSSPAP
14,888
SFV1_P23074





EAAAKGGGGSS
14,889
XMRV6_A1Z651





PAPAPAPAP
14,890
MMTVB_P03365





GGSEAAAKGSS
14,891
SFV3L_P27401_2mutA





GSSPAPGGG
14,892
MMTVB_P03365_WS





GGGGGG
14,893
SFV3L_P27401-Pro





GGSGGSGGS
14,894
FOAMV_P14350-Pro_2mut





PAPAPAPAPAPAP
14,895
WMSV_P03359





GSSPAP
14,896
MLVBM_Q7SVK7





GGGGGSGSS
14,897
MMTVB_P03365_2mut_WS





EAAAKGSSGGS
14,898
MMTVB_P03365_2mutB_WS





EAAAK
14,899
FFV_O93209_2mutA





PAPEAAAK
14,900
SFV1_P23074-Pro





EAAAKGGSGSS
14,901
SFV3L_P27401





GGSGGSGGS
14,902
FFV_O93209-Pro





GSSGGGEAAAK
14,903
MMTVB_P03365





SGGSSGGSSGSETPGTSE
14,904
MLVFF_P26809_3mutA


SATPESSGGSSGGSS







GGSGGSGGSGGSGGSGGS
14,905
HTL1L_P0C211_2mutB





GGGEAAAK
14,906
SFV3L_P27401-Pro_2mutA





GGGGGSGSS
14,907
MMTVB_P03365





GSSPAPGGS
14,908
FOAMV_P14350_2mutA





EAAAKGSS
14,909
MLVMS_P03355





GSSGGSGGG
14,910
FFV_O93209-Pro





GGSGGGGSS
14,911
MMTVB_P03365-Pro_2mut





GGSPAPGSS
14,912
FOAMV_P14350_2mut





GGSGGSGGSGGSGGSGGS
14,913
SFVCP_Q87040-Pro_2mut





GSSEAAAKGGG
14,914
FOAMV_P14350_2mutA





GGSGGSGGS
14,915
MMTVB_P03365-Pro





GSSGSSGSSGSSGSSGSS
14,916
MMTVB_P03365_2mut_WS





GSSGSSGSSGSSGSS
14,917
MMTVB_P03365-Pro





PAPEAAAK
14,918
WDSV_O92815





GSSGSSGSSGSSGSS
14,919
FFV_O93209-Pro_2mut





EAAAKGGGGSEAAAK
14,920
MMTVB_P03365-Pro





GGSPAPEAAAK
14,921
FOAMV_P14350





GSSGSS
14,922
PERV_Q4VFZ2





GGG

MMTVB_P03365-Pro





GGGGSGGGGSGGGGS
14,924
FFV_O93209_2mut





EAAAKEAAAKEAAAKEAA
14,925
MMTVB_P03365-Pro


AKEAAAKEAAAK







GGSGSSPAP
14,926
WMSV_P03359





GGGGGGGG
14,927
SFV3L_P27401_2mut





PAPGSSEAAAK
14,928
FOAMV_P14350-Pro_2mutA





GGGGSSPAP
14,929
FOAMV_P14350_2mut





GSSGGSPAP
14,930
MLVBM_Q7SVK7_3mut





GSSGGGGGS
14,931
GALV_P21414_3mutA





EAAAKEAAAKEAAAKEAA
14,932
MMTVB_P03365


AKEAAAK







GSSGGGGGS
14,933
SFV1_P23074_2mut





GGGGSEAAAKGGGGS
14,934
SFV1_P23074





GGGEAAAKPAP
14,935
FFV_O93209





PAPGGGEAAAK
14,936
SFV1_P23074





GGSGGGEAAAK
14,937
PERV_Q4VFZ2_3mutA_WS





GSSGGG
14,938
MMTVB_P03365-Pro





EAAAKGSSGGS
14,939
FFV_O93209_2mut





GGGGG
14,940
SFV1_P23074_2mut





GGGPAP
14,941
SFV3L_P27401





GSSGGSEAAAK
14,942
FFV_O93209





SGGSSGGSSGSETPGTSE
14,943
MMTVB_P03365-Pro


SATPESSGGSSGGSS







GSSGGGEAAAK
14,944
SFV1_P23074_2mutA





GSSGSSGSSGSSGSS
14,945
SFV3L_P27401_2mut





GGSEAAAKPAP
14,946
FLV_P10273





GGGGSGGGGS
14,947
FOAMV_P14350-Pro_2mutA





GSSEAAAKPAP
14,948
SFV3L_P27401





GGGGSEAAAKGGGGS
14,949
MMTVB_P03365-Pro





PAPGSSEAAAK
14,950
MLVF5_P26810_3mut





EAAAKGGSGGG
14,951
SFV3L_P27401





GGGPAPGGS
14,952
SFV3L_P27401





GSSEAAAKGGS
14,953
FOAMV_P14350_2mutA





EAAAKGGSGGG
14,954
HTL1L_P0C211





GSSGGSPAP
14,955
SFV3L_P27401_2mutA





PAPAP
14,956
FFV_O93209





PAPGGSGSS
14,957
MMTVB_P03365_WS





EAAAKGGGGGS
14,958
FOAMV_P14350_2mut





PAPEAAAKGGS
14,959
SFV3L_P27401_2mut





GSSEAAAKPAP
14,960
MMTVB_P03365-Pro





GGSGGS
14,961
PERV_Q4VFZ2_3mut





GSSEAAAKGGG
14,962
FFV_O93209-Pro_2mutA





EAAAK
14,963
HTL1L_P0C211





GSSPAP
14,964
MLVMS_P03355





EAAAKPAPGGG
14,965
FFV_O93209-Pro_2mut





GGGGSEAAAKGGGGS
14,966
SFV1_P23074-Pro_2mut





EAAAKGSSGGS
14,967
SFV3L_P27401





GSAGSAAGSGEF
14,968
FFV_O93209_2mutA





PAPEAAAKGGS
14,969
MMTVB_P03365_2mutB_WS





EAAAKEAAAKEAAAKEAA
14,970
MMTVB_P03365


AKEAAAKEAAAK







GGS

MMTVB_P03365





GGSEAAAKPAP
14,972
SFV1_P23074





EAAAKGSSGGG
14,973
HTLV2_P03363_2mut





GGSEAAAKGGG
14,974
MMTVB_P03365_WS





GGSGGS
14,975
FFV_O93209-Pro





GSSEAAAKGGS
14,976
MMTVB_P03365-Pro





PAPAPAPAPAP
14,977
SFV1_P23074_2mutA





GGSEAAAKGGG
14,978
MMTVB_P03365_2mutB_WS





PAPAPAPAP
14,979
MMTVB_P03365_WS





GGGGSGGGGSGGGGSGGG
14,980
HTL3P_Q4U0X6_2mut


GSGGGGS







PAPGGSEAAAK
14,981
SFV1_P23074-Pro_2mut





GGSGGGPAP
14,982
MMTVB_P03365





GSSGSSGSSGSSGSSGSS
14,983
MMTVB_P03365-Pro





GGSEAAAKPAP
14,984
SFV1_P23074-Pro





GGGEAAAKGSS
14,985
SFV3L_P27401_2mutA





GGGPAPGGS
14,986
AVIRE_P03360





PAPGGG
14,987
MLVRD_P11227





GGSEAAAKGSS
14,988
SFV3L_P27401_2mut





GGGEAAAKGSS
14,989
FOAMV_P14350_2mut





GGGEAAAKGSS
14,990
SFV1_P23074-Pro





EAAAKEAAAKEAAAKEAA
14,991
MLVAV_P03356


AK







EAAAKGGGPAP
14,992
JSRV_P31623_2mutB





EAAAKGGGGSS
14,993
FOAMV_P14350_2mut





EAAAKEAAAKEAAAKEAA
14,994
SRV2_P51517


AKEAAAK







GSSGGGGGS
14,995
FFV_O93209





PAPAPAP
14,996
FOAMV_P14350_2mutA





GGSGGSGGSGGS
14,997
FOAMV_P14350





GGGEAAAK
14,998
MMTVB_P03365_WS





GGGGGS
14,999
SFV1_P23074_2mutA





GGSGGS
15,000
WMSV_P03359_3mut





EAAAKGGS
15,001
MMTVB_P03365-Pro





GGGGSS
15,002
BLVJ_P03361_2mut





PAPAP
15,003
MMTVB_P03365-Pro_2mut





PAPGGG
15,004
SMRVH_P03364





EAAAKGGGGSS
15,005
SFV3L_P27401





PAPAPAPAPAP
15,006
MMTVB_P03365





GGGPAP
15,007
MMTVB_P03365-Pro





GSSGGSGGG
15,008
MMTVB_P03365





EAAAKGGGPAP
15,009
FOAMV_P14350_2mutA





GSSGSSGSSGSS
15,010
SFV1_P23074





GGGGSGGGGS
15,011
SFV3L_P27401





GSSGGSGGG
15,012
MLVF5_P26810





GGGEAAAKPAP
15,013
MMTVB_P03365-Pro





PAPEAAAK
15,014
HTLV2_P03363_2mut





GSSGSSGSSGSS
15,015
FOAMV_P14350_2mut





GSSEAAAKPAP
15,016
MMTVB_P03365-Pro





PAPEAAAKGGG
15,017
HTL3P_Q4U0X6_2mut





GGSEAAAKGSS
15,018
MMTVB_P03365-Pro





EAAAKPAPGGS
15,019
MMTVB_P03365_2mut_WS





GSSGGSEAAAK
15,020
MLVF5_P26810_3mutA





GGGGSGGGGSGGGGSGGG
15,021
MLVF5_P26810_3mut


GSGGGGSGGGGS







EAAAKGGGGSS
15,022
MMTVB_P03365-Pro





GGGGGSGSS
15,023
HTL1A_P03362_2mutB





PAPAP
15,024
FFV_O93209-Pro_2mut





GGGGGSPAP
15,025
HTL1C_P14078_2mut





GGGPAP
15,026
HTLV2_P03363_2mut





EAAAKGGGGSEAAAK
15,027
SFVCP_Q87040





GGSEAAAKGGG
15,028
FFV_O93209-Pro_2mutA





GSSPAPGGS
15,029
FOAMV_P14350-Pro_2mut





GGGGGGG
15,030
MMTVB_P03365-Pro





EAAAKGSS
15,031
SFV3L_P27401_2mutA





EAAAKGGGGSEAAAK
15,032
MMTVB_P03365-Pro





GGGGSEAAAKGGGGS
15,033
SFV1_P23074-Pro_2mutA





EAAAKGGGGSS
15,034
MMTVB_P03365





GGGEAAAKGGS
15,035
SFV1_P23074





PAPEAAAKGGG
15,036
MLVF5_P26810





GGGGSSGGS
15,037
MMTVB_P03365





GGSGSS
15,038
MMTVB_P03365





PAPAPAPAPAPAP
15,039
KORV_Q9TTC1





EAAAKGGG
15,040
SFV1_P23074-Pro_2mut





PAPAPAPAPAPAP
15,041
SRV2_P51517





GSSGSSGSSGSSGSS
15,042
FFV_O93209-Pro_2mutA





GGGGSS
15,043
FOAMV_P14350_2mut





PAPGGGEAAAK
15,044
MMTVB_P03365_WS





GGSGGGEAAAK
15,045
FFV_O93209-Pro_2mut





PAPAPAPAPAP
15,046
MMTVB_P03365_WS





GGGEAAAKGGS
15,047
MMTVB_P03365-Pro





GGGEAAAKGSS
15,048
MMTVB_P03365_2mutB





GSSPAPEAAAK
15,049
MMTVB_P03365_WS





EAAAKEAAAKEAAAKEAA
15,050
SFV1_P23074-Pro_2mutA


AKEAAAK







PAPGGG
15,051
SFV3L_P27401





GSSEAAAKGGG
15,052
MMTVB_P03365_WS





GGGGSSEAAAK
15,053
FOAMV_P14350_2mut





PAPGSSGGS
15,054
SFV1_P23074-Pro_2mut





GSSGSSGSSGSSGSSGSS
15,055
SFV3L_P27401





EAAAKGSSGGG
15,056
MMTVB_P03365





PAPGGGGSS
15,057
WDSV_O92815_2mutA





GGSPAP
15,058
MMTVB_P03365-Pro





GGSGGSGGSGGSGGS
15,059
SFVCP_Q87040-Pro_2mut





PAPAPAPAP
15,060
MMTVB_P03365-Pro





GGGGG
15,061
HTL1A_P03362





GGSGGSGGSGGS
15,062
SFV1_P23074_2mutA





GSSGSSGSSGSSGSS
15,063
FOAMV_P14350-Pro_2mut





PAPGGSEAAAK
15,064
MMTVB_P03365_2mutB_WS





PAPAPAPAP
15,065
SFV1_P23074_2mut





PAPGGGGSS
15,066
MMTVB_P03365





GGSGSS
15,067
SFV3L_P27401_2mut





EAAAKEAAAKEAAAKEAA
15,068
MMTVB_P03365_2mut


AK







EAAAKGGSGGG
15,069
HTL3P_Q4U0X6_2mut





PAPGGGGSS
15,070
SFVCP_Q87040-Pro_2mutA





EAAAKGGGGGS
15,071
MLVAV_P03356





GGGGGS
15,072
FOAMV_P14350_2mut





GGGEAAAKGGS
15,073
FFV_O93209-Pro_2mutA





EAAAKPAPGGG
15,074
MMTVB_P03365_2mutB





GGSGGGPAP
15,075
FFV_O93209_2mut





GSSEAAAKPAP
15,076
MMTVB_P03365





PAPAPAPAPAPAP
15,077
SFV1_P23074_2mut





GGSPAPGGG
15,078
MMTVB_P03365-Pro





GGSGGGEAAAK
15,079
MMTVB_P03365





PAPAP
15,080
SFVCP_Q87040





GSSEAAAK
15,081
SFVCP_Q87040





GGGGSGGGGSGGGGS
15,082
MMTVB_P03365-Pro





GSSGSSGSS
15,083
SFV3L_P27401





EAAAKGGSGGG
15,084
MMTVB_P03365-Pro





GSSPAP
15,085
SFV1_P23074_2mut





GGGEAAAK
15,086
SFV1_P23074-Pro





AEAAAKEAAAKEAAAKEA
15,087
MMTVB_P03365-Pro


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







PAPGGS
15,088
HTL1C_P14078_2mut





PAPGSSGGS
15,089
SFV1_P23074_2mut





PAPEAAAK
15,090
MMTVB_P03365_WS





PAPAP
15,091
MMTVB_P03365-Pro





EAAAKGGS
15,092
HTL1A_P03362_2mut





GGGGSEAAAKGGGGS
15,093
HTL1C_P14078





EAAAKGSSGGS
15,094
FOAMV_P14350-Pro





PAPGGSGSS
15,095
MMTVB_P03365-Pro





PAPGGSEAAAK
15,096
SFV1_P23074_2mut





PAPGSSEAAAK
15,097
FFV_O93209-Pro_2mut





PAPGSSGGG
15,098
FOAMV_P14350-




Pro_2mutA





GSSGGGEAAAK
15,099
AVIRE_P03360





GGGGGG
15,100
SMRVH_P03364_2mut





PAPEAAAKGGG
15,101
MMTVB_P03365-Pro





GGGEAAAKGGS
15,102
SFVCP_Q87040_2mutA





PAPAPAPAPAP
15,103
SRV2_P51517





GSSGSSGSSGSSGSSGSS
15,104
MMTVB_P03365





EAAAKGGGPAP
15,105
MLVAV_P03356





PAPAPAPAPAP
15,106
FOAMV_P14350-




Pro_2mutA





PAPGGSEAAAK
15,107
FOAMV_P14350





GSSGGGPAP
15,108
HTL32_Q0R5R2_2mutB





GGGGGSPAP
15,109
HTL3P_Q4U0X6_2mutB





GSSGGSGGG
15,110
MMTVB_P03365-Pro





PAPAP
15,111
SFVCP_Q87040-Pro





GSSGGGPAP
15,112
MMTVB_P03365-Pro





GGSGSS
15,113
MMTVB_P03365-Pro_2mut





GGSPAPEAAAK
15,114
SFV1_P23074-Pro_2mut





EAAAKGGSGGG
15,115
SFV3L_P27401_2mut





GGGGSSEAAAK
15,116
MMTVB_P03365_WS





GGGGGSGSS
15,117
MMTVB_P03365_2mut





GGGGSSGGS
15,118
SFV1_P23074-Pro_2mutA





EAAAKGGGGSEAAAK
15,119
MMTVB_P03365_WS





PAPGGGEAAAK
15,120
SFV1_P23074-Pro





PAPEAAAKGGG
15,121
MMTVB_P03365





AEAAAKEAAAKEAAAKEA
15,122
MMTVB_P03365


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GSSGGSEAAAK
15,123
FOAMV_P14350-Pro_2mut





GGSPAP
15,124
MLVBM_Q7SVK7_3mut





GSSEAAAK
15,125
FOAMV_P14350





GSSEAAAK
15,126
MMTVB_P03365-Pro





EAAAKGSSGGS
15,127
HTL1A_P03362_2mut





GGGEAAAKPAP
15,128
FOAMV_P14350-Pro_2mut





EAAAKGGSPAP
15,129
FOAMV_P14350





GSSEAAAKPAP
15,130
MMTVB_P03365_WS





GSSGSSGSS
15,131
FOAMV_P14350_2mut





EAAAKEAAAKEAAAKEAA
15,132
MMTVB_P03365_WS


AK







EAAAK
15,133
MMTVB_P03365





PAPGSS
15,134
BAEVM_P10272





PAPGGS
15,135
FFV_O93209-Pro_2mut





GGSGGS
15,136
SFV1_P23074-Pro_2mutA





SGGSSGGSSGSETPGTSE
15,137
HTLV2_P03363_2mut


SATPESSGGSSGGSS







GGSGGGEAAAK
15,138
MMTVB_P03365_WS





PAPGSSGGG
15,139
HTL1A_P03362





GGSGGS
15,140
SFV3L_P27401-Pro





GSSGSS
15,141
SFV1_P23074-Pro





PAPGGSEAAAK
15,142
MMTVB_P03365





GSAGSAAGSGEF
15,143
MMTVB_P03365-Pro





PAPGGG
15,144
FOAMV_P14350_2mut





EAAAKGGSGSS
15,145
MMTVB_P03365_WS





GSSGGGEAAAK
15,146
SFV3L_P27401-Pro





GGSGGGPAP
15,147
FOAMV_P14350-Pro_2mut





PAPAPAPAPAPAP
15,148
WDSV_O92815





SGSETPGTSESATPES
15,149
SFVCP_Q87040-Pro_2mutA





GGSGGSGGS
15,150
SFV1_P23074





GGGGSS
15,151
SFVCP_Q87040_2mut





GGGGGSEAAAK
15,152
MMTVB_P03365





SGSETPGTSESATPES
15,153
MMTVB_P03365_WS





PAPAPAP
15,154
SFV3L_P27401





PAPEAAAKGSS
15,155
MMTVB_P03365_2mutB_WS





GSSGSSGSSGSSGSS
15,156
SRV2_P51517





GGGPAPGSS
15,157
HTL32_Q0R5R2_2mutB





GGSGGGGSS
15,158
MMTVB_P03365-Pro





SGSETPGTSESATPES
15,159
SRV2_P51517





EAAAKGSSGGS
15,160
MMTVB_P03365-Pro





GSSPAPEAAAK
15,161
MMTVB_P03365-Pro





GSSPAPEAAAK
15,162
SRV2_P51517





GGGGSSPAP
15,163
MMTVB_P03365-Pro





PAPGGGEAAAK
15,164
SFV1_P23074-Pro_2mutA





PAPEAAAKGGS
15,165
MMTVB_P03365





GSSGSSGSSGSSGSSGSS
15,166
FOAMV_P14350-Pro





GGSPAPGSS
15,167
SFV3L_P27401





GGGPAPGGS
15,168
SFV1_P23074-Pro_2mutA





GGGPAPGSS
15,169
MMTVB_P03365-Pro





EAAAKPAP
15,170
MLVBM_Q7SVK7





EAAAKEAAAKEAAAK
15,171
HTL1C_P14078





GSSGGSEAAAK
15,172
SRV2_P51517





PAPGGGGGS
15,173
SRV2_P51517





GGGEAAAK
15,174
FFV_O93209-Pro_2mut





EAAAKGGGPAP
15,175
HTL32_Q0R5R2





GGSGSSGGG
15,176
MMTVB_P03365





PAPEAAAKGSS
15,177
MMTVB_P03365-Pro





PAPGGGGGS
15,178
MMTVB_P03365-Pro





EAAAKGGGGGS
15,179
MMTVB_P03365_WS





GGGGGS
15,180
MMTVB_P03365-Pro





GGGGGGGGSGGGGSGGGG
15,181
HTL1C_P14078


SGGGGS







EAAAKGGSPAP
15,182
MMTVB_P03365





GGGGSSPAP
15,183
FFV_O93209-Pro_2mut





GGGGSSGGS
15,184
MMTVB_P03365-Pro





PAPGSSGGS
15,185
MMTVB_P03365-Pro





GGGGGS
15,186
SRV2_P51517





GGSGSSGGG
15,187
MMTVB_P03365





GSSGGSEAAAK
15,188
MMTVB_P03365-Pro





EAAAKEAAAKEAAAKEAA
15,189
GALV_P21414


AK







GGSEAAAKGGG
15,190
MMTVB_P03365-Pro





SGGSSGGSSGSETPGTSE
15,191
MMTVB_P03365-Pro


SATPESSGGSSGGSS







GSSEAAAKGGS
15,192
MMTVB_P03365





GGGGSGGGGSGGGGSGGG
15,193
HTL3P_Q4U0X6_2mutB


GSGGGGSGGGGS







GGGEAAAK
15,194
MMTVB_P03365-Pro





PAPAPAPAP
15,195
MMTVB_P03365-Pro





PAPGSSGGG
15,196
MMTVB_P03365





GSSGSSGSSGSSGSS
15,197
GALV_P21414





GGSPAP
15,198
MMTVB_P03365_WS





GGGGSGGGGSGGGGSGGG
15,199
MMTVB_P03365-Pro


GSGGGGSGGGGS







PAPEAAAK
15,200
MMTVB_P03365-Pro





PAPGSSGGG
15,201
SFV1_P23074-Pro_2mutA





GGGGGSEAAAK
15,202
MMTVB_P03365_2mutB_WS





PAPAPAPAPAP
15,203
MMTVB_P03365-Pro





EAAAKGGSGSS
15,204
MMTVB_P03365-Pro





EAAAKEAAAKEAAAKEAA
15,205
MLVRD_P11227_3mut


AK







PAPAPAPAP
15,206
FOAMV_P14350_2mutA





GGGPAPGSS
15,207
SFVCP_Q87040_2mut





PAPEAAAKGSS
15,208
SFVCP_Q87040_2mut





GGSPAPGGG
15,209
MMTVB_P03365-Pro





GGGGSGGGGSGGGGSGGG
15,210
MMTVB_P03365


GS







EAAAKGGS
15,211
HTL3P_Q4U0X6_2mut





PAPGSSGGS
15,212
MMTVB_P03365_WS





GGGGSGGGGS
15,213
MMTVB_P03365





GGSGGS
15,214
FOAMV_P14350





EAAAKGGGGSEAAAK
15,215
SFVCP_Q87040-Pro_2mut





EAAAKEAAAKEAAAKEAA
15,216
MMTVB_P03365-


AK

Pro_2mutB





PAPGGGEAAAK
15,217
SFVCP_Q87040-Pro





GSSGSS
15,218
JSRV_P31623_2mutB





EAAAKGGGGGS
15,219
MMTVB_P03365_2mut_WS





GSSPAPEAAAK
15,220
MMTVB_P03365-Pro





GGGEAAAK
15,221
HTL1C_P14078





PAPEAAAKGSS
15,222
HTL32_Q0R5R2_2mutB





GGGGSSEAAAK
15,223
MMTVB_P03365-Pro





PAPGSSGGS
15,224
MMTVB_P03365-Pro





EAAAKGGGGGS
15,225
MMTVB_P03365





GGGGSGGGGSGGGGSGGG
15,226
MMTVB_P03365


GS







EAAAKGGGGSS
15,227
HTL3P_Q4U0X6_2mut





GGGEAAAKGGS
15,228
SFVCP_Q87040-Pro





GGGGGSPAP
15,229
MMTVB_P03365-




Pro_2mutB





GGSGGGEAAAK
15,230
SFV3L_P27401-Pro





PAPGGGGGS
15,231
SFV3L_P27401-Pro





EAAAKGGGGSEAAAK
15,232
MMTVB_P03365





PAPEAAAKGSS
15,233
MMTVB_P03365-Pro





GGSEAAAKGGG
15,234
MMTVB_P03365-Pro





GGSGGSGGSGGSGGS
15,235
SMRVH_P03364_2mutB





GGSGGSGGSGGSGGS
15,236
HTL1L_P0C211_2mut





GGGGGG
15,237
WDSV_O92815





GGGGGSGSS
15,238
MMTVB_P03365-Pro





GGSEAAAKPAP
15,239
SFV3L_P27401-Pro_2mut





GGGPAPGSS
15,240
MMTVB_P03365_2mut_WS





GGGGGS
15,241
MMTVB_P03365_WS





GGSPAPEAAAK
15,242
MMTVB_P03365





PAPEAAAKGGS
15,243
HTL1A_P03362





EAAAKGGSGSS
15,244
MMTVB_P03365_2mut_WS





GGGPAPEAAAK
15,245
SFV3L_P27401-Pro_2mut





PAPGGGGSS
15,246
HTL32_Q0R5R2_2mut





GSSPAPGGG
15,247
HTL3P_Q4U0X6_2mut





GGGGSSGGS
15,248
BLVAU_P25059_2mut





EAAAKGGGGGS
15,249
HTL1L_P0C211





GGSEAAAKGSS
15,250
JSRV_P31623_2mutB





GSSGGG
15,251
JSRV_P31623





GGSGGSGGSGGS
15,252
MMTVB_P03365-Pro





EAAAKPAP
15,253
SFV1_P23074-Pro_2mutA





GGGGSSGGS
15,254
MMTVB_P03365_WS





GGSGGS
15,255
MMTVB_P03365_WS





EAAAKGGGGGS
15,256
MMTVB_P03365-Pro





GGGGSGGGGSGGGGSGGG
15,257
MMTVB_P03365


GSGGGGSGGGGS







GGSGGSGGS
15,258
MMTVB_P03365





GGGGGSEAAAK
15,259
MLVBM_Q7SVK7





GGSGSSPAP
15,260
MMTVB_P03365_WS





EAAAKEAAAKEAAAK
15,261
JSRV_P31623





PAPEAAAKGGS
15,262
MMTVB_P03365-Pro





GGSGSSEAAAK
15,263
FOAMV_P14350





GGGGGSGSS
15,264
MMTVB_P03365-Pro_2mut





GGGPAPGGS
15,265
MMTVB_P03365





SGSETPGTSESATPES
15,266
SFVCP_Q87040_2mut





GSSPAPGGS
15,267
SFV1_P23074-Pro_2mutA





GSSGSSGSSGSSGSS
15,268
MMTVB_P03365





EAAAKGGGPAP
15,269
MMTVB_P03365





GSSGGG
15,270
MMTVB_P03365_2mut_WS





GGGEAAAKPAP
15,271
MMTVB_P03365





PAPGGSGGG
15,272
MMTVB_P03365-Pro





GSSGGSGGG
15,273
WDSV_O92815_2mut





GGSGGG
15,274
HTL32_Q0R5R2_2mut





EAAAKGGSPAP
15,275
HTLV2_P03363_2mut





GGSPAPEAAAK
15,276
MMTVB_P03365-Pro





GSSGGSEAAAK
15,277
MMTVB_P03365_2mut





GSAGSAAGSGEF
15,278
MMTVB_P03365_WS





PAPGGSGSS
15,279
FFV_O93209





GGSEAAAKGGG
15,280
MMTVB_P03365





GGSPAPGSS
15,281
MMTVB_P03365-Pro





GSSGGSGGG
15,282
SFV3L_P27401





PAPEAAAKGGG
15,283
HTL1A_P03362_2mutB





GGGEAAAKPAP
15,284
MMTVB_P03365-Pro





GGSEAAAK
15,285
HTL32_Q0R5R2_2mutB





GGGEAAAKGSS
15,286
MPMV_P07572





GGGGGSEAAAK
15,287
MMTVB_P03365-Pro





PAPAPAPAPAP
15,288
SFVCP_Q87040-Pro_2mutA





PAPAPAPAPAP
15,289
HTL1L_P0C211_2mut





GGGGSSGGS
15,290
HTL3P_Q4U0X6





PAPGGSEAAAK
15,291
MMTVB_P03365_2mut_WS





PAPAPAPAPAP
15,292
HTL1A_P03362





EAAAKPAPGGG
15,293
MMTVB_P03365_2mut_WS





GGSEAAAK
15,294
MMTVB_P03365_2mut_WS





GGGEAAAKGSS
15,295
SFV1_P23074-Pro_2mutA





GGSPAPGSS
15,296
MMTVB_P03365-Pro





GGSEAAAKPAP
15,297
MLVBM_Q7SVK7





PAPEAAAKGGG
15,298
MMTVB_P03365_2mut_WS





GSSEAAAKPAP
15,299
MMTVB_P03365-Pro_2mutB





GGGGSEAAAKGGGGS
15,300
MMTVB_P03365-Pro_2mut





GSSEAAAKGGS
15,301
MMTVB_P03365-Pro_2mutB





GSSGSSGSSGSSGSS
15,302
SRV2_P51517_2mutB





GGGGGSPAP
15,303
HTL1L_P0C211_2mut





GGSEAAAK
15,304
MMTVB_P03365





GSSPAPEAAAK
15,305
SMRVH_P03364_2mutB





GGGPAPGGS
15,306
HTL1C_P14078_2mut





GGSPAPEAAAK
15,307
MMTVB_P03365_WS





GGSEAAAKPAP
15,308
HTL1A_P03362_2mut





PAPAPAPAP
15,309
HTLV2_P03363_2mut





GSSPAPGGG
15,310
MMTVB_P03365





GSSGSSGSSGSS
15,311
MMTVB_P03365-Pro





GGSEAAAKGSS
15,312
MMTVB_P03365_WS





GGSGSSGGG
15,313
MMTVB_P03365_2mutB





GSSGSSGSSGSSGSSGSS
15,314
JSRV_P31623_2mutB





GGSEAAAKPAP
15,315
MMTVB_P03365-Pro





GSSGGSGGG
15,316
HTLV2_P03363_2mut





AEAAAKEAAAKEAAAKEA
15,317
WDSV_O92815_2mut


AAKALEAEAAAKEAAAKE




AAAKEAAAKA







GGSPAPEAAAK
15,318
MMTVB_P03365





GGGGSSEAAAK
15,319
MMTVB_P03365





GGSGGGEAAAK
15,320
SFV1_P23074-Pro_2mutA





GGGGSEAAAKGGGGS
15,321
WDSV_O92815_2mut





GGSGSSEAAAK
15,322
MMTVB_P03365_2mutB_WS





GGSEAAAKPAP
15,323
MMTVB_P03365_WS





GSSGGGEAAAK
15,324
SFVCP_Q87040-Pro





GSSGGS
15,325
SFVCP_Q87040-Pro_2mut





GGSEAAAKPAP
15,326
SFVCP_Q87040_2mut





GSSGGSEAAAK
15,327
SFVCP_Q87040_2mut





GSSPAPEAAAK
15,328
SRV2_P51517_2mutB





GGSGGSGGSGGSGGSGGS
15,329
BLVAU_P25059





GSSGSSGSSGSSGSS
15,330
HTL1C_P14078_2mut





EAAAKGGGGSS
15,331
MMTVB_P03365_2mutB





GGGEAAAKGSS
15,332
SFVCP_Q87040-Pro









Example 3: Optimization of Lipid Nanoparticle Compositions for Delivery of Gene Modifying Systems to Correct the Pathogenic E342K Mutation Associated with Alpha-1 Antitrypsin Deficiency

In this example, lipid nanoparticle (LNP) components are formulated as described in Example 44 of WO2021/178720. Specifically, the lipid nanoparticle (LNP) components (ionizable lipid, helper lipid, sterol, PEG) are dissolved in 100% ethanol with the lipid component molar ratios of 50:10:38.5:1.5, respectively. An mRNA encoding a gene modifying polypeptide as described herein is produced by in vitro transcription and purified mRNA is dissolved in 25 mM sodium citrate, pH 4, to a final concentration of RNA cargo of 0.1 mg/mL. Similarly, a Template RNA designed to correct the E342K mutation in SERPINA1 and optionally optimized for use with the specific gene modifying polypeptide (as described herein) is dissolved in 25 mM sodium citrate, pH 4. Optionally, a second-nick gRNA as described herein is dissolved in 25 mM sodium citrate, pH 4.


Each RNA is separately formulated into distinct LNPs with a lipid amine to RNA phosphate (N:P) molar ratio of 6. The LNPs are formed by microfluidic mixing of the lipid and RNA solutions using a Precision Nanosystems NanoAssemblr™ Benchtop Instrument, using the manufacturer's recommended settings. A 3:1 ratio of aqueous to organic solvent is maintained during mixing using differential flow rates. After mixing, the LNPs are collected and dialyzed in 15 mM Tris, 5% sucrose buffer at 4° C. overnight. Formulations are concentrated by centrifugation with Amicon 10 kDa centrifugal filters (Millipore). The resulting mixture is then filtered using a 0.2 μm sterile filter. The final LNP composition is stored at −80° C. until further use.


Additional LNP formulations are generated to optimize the formulation composition and process for delivery and function of a gene modifying system. The lipid nanoparticle components are varied according to the following parameters: 30-60% ionizable lipid, e.g., an ionizable lipid in Table 19 or described elsewhere in this application, 5-15% helper phospholipid Di stearoylphosphatidylcholine (DSPC), 30-50% cholesterol, and 0.5-5% Polyethylene glycol (PEG). Beyond the lipid composition, additional formulations comprising combinations of gene modifying components are generated, e.g., an mRNA encoding the gene modifying polypeptide is co-formulated with a Template RNA for correcting the disease-causing mutation, and optionally a second-nick gRNA is either co-formulated with the mRNA and Template RNA, or formulated separately. In some embodiments, the mRNA and Template RNA, and optionally a second-nick gRNA, are co-formulated with the lipid nanoparticle components to make the total RNA cargo at a concentration approximately 0.1 mg/mL. The RNA composition for co-formulation is a mix of the mRNA and Template RNA at a 1-4:1-10 ratio by weight, respectively, or is a mix of mRNA, Template RNA, and second-nick gRNA at a ratio of 1-4:1-10:1-10, respectively.


Alternate formulations described in this example include RNAs of the system, e.g., mRNA encoding a gene modifying polypeptide, Template RNA, and optional second-nick gRNA, being separately formulated using identical or different ionizable lipids, or identical ionizable lipids formulated with different lipid component ratios as described herein. An exemplary formulation has a gene modifying polypeptide mRNA formulated using the ionizable lipid LIPIDV004, where the formulation is a ratio of 50:10:38.5:1.5 of ionizable lipid, helper lipid, sterol, and PEG, respectively. The RNA is mixed with the lipid at a lipid amine to RNA phosphate (N:P) ratio of 6. An exemplary Template RNA for use with the exemplary mRNA is formulated using the ionizable lipid LIPIDV004, where the formulation is a ratio of 50:10:38.5:1.5 of ionizable lipid, helper lipid, sterol, and PEG, respectively. The Template RNA is mixed with the lipid at an N:P ratio of 4. An exemplary optional second-nick RNA for further use in this system is formulated using the ionizable lipid LIPIDV004, where the formulation is a ratio of 50:10:38.5:1.5 of ionizable lipid, helper lipid, sterol, and PEG, respectively, with the optional second-nick gRNA being mixed with lipid at an N:P ratio of 4.


As described herein, a single-nucleotide polymorphism in the SERPINA1 gene causes the pathogenic E342K mutation that leads alpha-1 anti-trypsin deficiency (AATD). This particular amino acid change, known as the Pi*Z allele in humans, has been modeled in the transgenic mouse line B6.Cg-Tg (SERPINA1*E342K) Z11.03Slcw/ChmuJ (stock #035411, The Jackson Laboratory), which expresses the Pi*Z allele of human SERPINA1 in the liver and kidney at levels similar to human patients with AATD. To correct the amino acid substitution and ameliorate the effects caused by the non-functional AAT protein an optimized gene modifying system described herein, e.g., a gene modifying system composition described in Table 4, or a composition from Table 4 further modified to utilize an RT template region introducing a PAM disruption at the target site as in Table 5, is delivered to a transgenic mouse model of AATD by an LNP formulation described in Example 46 of WO2021/178720 or Example 4, below. To determine any efficacy-modifying effects of a second-nick gRNA, formulations including or lacking the second-nick gRNA are prepared along with the gene modifying polypeptide mRNA and disease-modifying Template RNA, and additionally prepared as separate LNPs or co-formulations. LNPs of this example are prepared as described in an example of this application and delivered intravenously to disease model mice at a total RNA amount of 1 mg/kg. Mice are monitored for correction in the liver and kidneys through various immunological, physiological, and molecular assays, including detection of wild-type human AAT, e.g., hAAT-specific ELISA, histology for detection of changes in liver and/or kidney fibrosis, immunohistochemistry to stain for intracellular hAAT, and amplicon sequencing for the genomic edit. As described herein, amplicon sequencing comprises using locus-specific primers to amplify across the target site containing the mutation, next-generation sequencing of purified amplicons, e.g., Illumina MiSeq, and computational analysis of amplicon sequencing data, e.g., analysis of editing outcome using the CRISPResso2 pipeline (Clement et al Nat Biotechnol 37(3):224-226 (2019)).


Example 4: Correction of SERPINA1 Gene Using Gene Modifying System to Treat Alpha-1 Anti-Trypsin Disease

This example describes the use of specific compositions of gene modifying systems to correct the E342K mutation in SERPINA1 that leads to alpha-1 antitrypsin deficiency in a mouse model of disease, as described in Example 3. A system for correction of the mutation in this model that employs a dual AAV delivery approach has been previously described and validated (Liu et al. bioRxiv (2020), doi.org/10.1101/2020.12.15.422970, the methods and compositions of which as related to editing the SERPINA1 gene, e.g., the methods and compositions of FIGS. 1a, 3a-d, and 5a-e, are incorporated herein by reference). Here, optimized all RNA gene modifying systems are employed to demonstrate a non-viral therapeutic approach to mutation correction. More specifically, a gene modifying system is employed that comprises (1) an mRNA that encodes a fusion protein that contains nuclear localization signals at the N- and C-terminus of the fusion of Streptococcus pyogenes Cas9 bearing a catalytic mutation, H840A, fused to the M-MLV reverse transcriptase, where the proteins are connected to one another using a 32 amino acid linker (SGGSSGGSSGSETPGTSESATPESSGGSSGGSS (SEQ ID NO: 5006)); along with (2) two guide RNAs, one functioning as a gene modifying Template RNA for targeting the genomic site in SERPINA1 for correction (UCCCCUCCAGGCCGUGCAUAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGG CUAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUCCUCUCGUCGAUGGU CAGCACAGCUUUAUGCACGGCCUGGAG (SEQ ID NO: 19529)) and another optional guide for second nicking the genome nearby to enhance correction (GGUUUGUUGAACUUGACCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGC UAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGCUUUUU (SEQ ID NO: 19530)). Here, the RNA compositions are formulated and delivered to mice using LNPs.


The mRNA and guide RNAs are formulated into LNPs using well-established mixing and purification, and concentration procedures, e.g., as described in Example 3. Briefly, the lipid nanoparticle (LNP) components (ionizable lipid (LIPIDV005 from Table 19), helper lipid (DSPC), sterol (Cholesterol), PEG (DMG-PEG-2000 (GM-20))) are dissolved in 100% ethanol with the lipid component molar ratios of 50:10:38.5:1.5, respectively. The mRNA encoding the gene modifying polypeptide (Cas-RT), Template RNA, and optionally second-nick guide RNA are dissolved in 25 mM sodium citrate, pH 4. Each RNA is separately formulated into LNPs with a lipid amine to RNA phosphate (N:P) molar ratio of 6. The LNPs are formed by microfluidic mixing of the lipid and RNA solutions using a Precision Nanosystems NanoAssemblr TM Benchtop Instrument, using the manufacturer's recommended settings. A 3:1 ratio of aqueous to organic solvent is maintained during mixing using differential flow rates. After mixing, the LNPs are collected and then dialyzed in 15 mM Tris, 5% sucrose buffer at 4° C. overnight. Formulations are concentrated by centrifugation with Amicon 10 kDa centrifugal filters (Millipore). The resulting mixture is filtered using a 0.2 μm sterile filter. The final LNPs are analyzed for particle size, polydispersity, and RNA integrity, e.g., as according to Example 44 of WO2021/178720, after aliquoting and stored at −80° C.


The LNPs are diluted from a concentrated stock to create a mixture where the molar ratio of each guide RNA (Template RNA and optionally second-nick guide RNA) is 20 times that of the mRNA. Evaluation in a mouse model is performed, as described in Example 3. The mRNA-LNP and guide RNAs-LNP mixture is injected intravenously to PiZ mice (e.g., B6.Cg-Tg(SERPINA1*E342K)Z11.03Slcw/ChmuJ, Stock No: 035411, The Jackson Laboratory) at a total RNA amount of 1 mg/kg. The mice are monitored for correction in the liver and kidneys though amplicon next-generation sequencing, production of the wild-type human alpha anti-trypsin protein, and histologic reduction in liver and kidney fibrosis.


In some embodiments, the compositions of the gene modifying system used to correct the E342K mutation in the PiZ model, as described above, are modified as follows to optimize efficiency and precision of editing.


Gene modifting polypeptide-encoding mRNA. In some embodiments, the gene modifying polypeptide comprises the bipartite SV40 NLS sequences (doi: 10.1074/jbc.M601718200) at its N-terminus and C-terminus. In some embodiments, The gene modifying system construct contains modified c-myc NLS and bipartite SV40 NLS at its N-terminus and at the C-terminus a modified bipartite SV40 NLS followed by a SV40 NLS is linked to the reverse transcriptase through a SGGS (SEQ ID NO: 25694) linker. In some embodiments, the linker between each NLS and the NLS and the fusion protein is a SGGS (SEQ ID NO: 25694) linker. In some embodiments, the 32 amino acid linker of the fusion protein encoded by the mRNA is:











(SEQ ID NO: 19531)



SGGSSGGSSGSETPGTSESATPESSGGSSGGSS






In some embodiments, the catalytic mutation of the Cas9 domain to generate the Cas9 nickase activity is H840A or N863A. In some embodiments, the mRNA has a cap, 5′ UTR containing a Kozak sequence, 3′ UTR, and a polyA tail containing at least 60 As (SEQ ID NO: 25695). In some embodiments, the mRNA has a reduced uridine content through codon selection/optimization. In some embodiments, the uridines in the mRNA are 100% substituted with 5-methoxy uridine. In some embodiments, the uridines in the mRNA are 100% substituted with N1-methyl-pseudouridine. In some embodiments, the cytosines in the mRNA are 100% substituted with 5-methylcytosine. In some embodiments, the mRNA contains a combination of 100% substitution of cytosine with 5-methylcytosine and 100% substitution of uridine with 5-methoxy uridine. In some embodiments, the mRNA contains a combination of 100% substitution of cytosine with 5-methylcytosine and 100% substitution of uridine with N1-methyl-pseudouridine. In some embodiments, combinations of modifications described above include 0-100% substitution of unmodified nucleotides, e.g., 0-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or less than 90-100% substitution. In some embodiments, the gene modifying polypeptide encoded by the mRNA of the system comprises the sequence:










c-Myc NLS-BPSV40 NLS-SpCas9H840A-linker-M-



MLV_reverse_transcriptase-SGGS linker-BPSV40 NLS-SV40





(SEQ ID NO: 19532)





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KFKVLGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEM





AKVDDSFFHRLEESFL VEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKL VDSTDKA





DLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAK





AILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDT





YDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQ





DLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELL





VKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYY





VGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPK





HSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDY





FKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDRE





MIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFA





NRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELV





KVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQ





NEKLYLYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRG





KSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVE





TRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYK VREINNYH





HAHDAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYS





NIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEV





QTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLK





SVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGE





LQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKR





VILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTST







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VEFEPKKKRKV






Template RNA and optional second-nick guide RNA. In some embodiments, the gene modifying system employs only a Template RNA in addition to the mRNA encoding the gene modifying polypeptide. In some embodiments, the gene modifying system additionally employs a second-nick guide RNA that targets the Cas9 nickase of the system to the non-edited strand of the target DNA. In some embodiments, the gene modifying Template RNA for targeting SERPINA1 is: UCCCCUCCAGGCCGUGCAUAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGG CUAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUCCUCUCGUCGAUGGU CAGCACAGCUUUAUGCACGGCCUGGAG (SEQ ID NO:19533). In some embodiments, the optional guide RNA for second nicking is: GGUUUGUUGAACUUGACCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGC UAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGCUUUUU (SEQ ID NO: 19534). In some embodiments, the Template RNA and optional second-nick guide RNA are synthesized by T7 RNA polymerase. In some embodiments, the Template RNA and optional second-nick guide RNA are chemically synthesized and contain a combination of one or multiple modifications of the following: 2′-O-methyl, 2′-Fluoro, and/or Phosphorothioate. In some embodiments, the 3 most terminal nucleotides contain 2′-O-methyl modifications with 3 phosphorothioate linkages between the nucleotides. In some embodiments, the Template RNA and optional second-nick guide RNA contain 2′-O-methyl modified nucleotides, where there are cytosines and uridines, except at nucleotides found in the seed sequence of the gRNA spacers, e.g., the seed sequences in the 3′ end of the spacer regions, where cytosines and uridines contain 2′-fluoro modifications and/or combination of 2′-fluoro and 2′ hydroxyl. In some embodiments, combinations of modifications described above include 0-100% substitution of unmodified nucleotides, e.g., 0-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or less than 90-100% substitution.


Formulations. In some embodiments, the gene modifying polypeptide mRNA and Template RNA (and optional second-nick guide RNA) are separately formulated as described above, combined prior to injection at a 1:20 RNA molar ratio, mRNA:Template RNA (and optionally mRNA:second-nick guide RNA), respectively. In some embodiments, the gene modifying polypeptide mRNA and Template RNA (and optional second-nick guide RNA) are separately formulated as described above, combined prior to injection at a 1:50 RNA molar ratio, mRNA:guide RNAs (and optionally mRNA:second-nick guide RNA), respectively. In some embodiments, the gene modifying polypeptide mRNA and Template RNA (and optional second-nick guide RNA) are separately formulated, combined prior to injection at ratio ranges from 1:10-1:250, mRNA:Template RNA (and optionally mRNA:second-nick guide RNA), respectively. In some embodiments, the mRNA and Template RNA (and optional second-nick guide NRA) are mixed together at a 1:10-1:250, mRNA:Template RNA (and optionally mRNA:second-nick guide RNA), and then formulated as described above, where the RNA concentration going into formulation is 0.1 mg/mL. In some embodiments, the mRNA and Template RNA (and optional second-nick guide RNA) are formulated separately and are injected 30-180 minutes apart, where the mRNA LNPs are delivered first followed by the Template RNA (and optional second-nick guide RNA) LNPs. In some embodiments, the ionizable lipid is LIPIDV005 from Table 19.


Dosing. In some embodiments, the gene modifying polypeptide mRNA and/or Template RNA (and optional second-nick guide RNA) are dosed at 0.01-6 mg/kg, either separately or together as a total amount of RNA-LNP. In some embodiments, the RNA-LNPs is injected as an IV bolus. In some embodiments, the RNA-LNPs is infused over a period of 30-360 minutes.


Example 5: Quantifying Activity of a Gene Editing Polypeptide and Template for Rewriting the Endogenous FAH Locus Achieved in Primary Mouse Hepatocytes

This example demonstrates the use of a gene modifying system containing a gene modifying polypeptide and a template RNA, to convert an A nucleotide to a G nucleotide in the endogenous Fah locus in mouse primary hepatocytes derived from a Fah5981SB mouse. The Fah5981SB mouse model harbors a G to A point mutation in the last nucleotide of exon 8 of the Fah gene, leading to aberrant mRNA splicing and subsequent mRNA degradation, without the production of Fah protein and, and thus serves as a mouse model of hereditary tyrosinemia type I.


In this example, the template RNA contained:

    • (1) a gRNA spacer;
    • (2) a gRNA scaffold;
    • (3) a heterologous object sequence; and
    • (4) a primer binding site (PBS) sequence.


More specifically, the template RNA (including chemical modification pattern) comprised the following sequences:









FAH1_R14_P12_Heavy RNACS048


(SEQ ID NO: 19535)


mG*mG*mA*rUrGrGrUrCrCrUrCrArUrGrArArCrGrArCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





UrUrArCrCrGrCrUrCrCrArGrUrCrGrUrUrCrArUrGrArG*mG*





mA*mC





FAH1_R15_P10_Heavy RNACS049


(SEQ ID NO: 19536)


mG*mG*mA*rUrGrGrUrCrCrUrCrArUrGrArArCrGrArCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





ArUrUrArCrCrGrCrUrCrCrArGrUrCrGrUrUrCrArUrG*mA*mG





*mG





FAH2_R19_P11_MUT_Heavy RNACS052


(SEQ ID NO: 19537)


mU*mC*mA*rGrArGrGrArArGrCrUrGrGrGrCrCrArCrCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





UrGrGrArGrCrGrGrUrArArUrGrGrCrUrGrGrUrGrGrCrCrCrA





rGrC*mU*mU*mC





FAH2_R19_P13_MUT_Heavy RNACS053


(SEQ ID NO: 19538)


mU*mC*mA*rGrArGrGrArArGrCrUrGrGrGrCrCrArCrCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





UrGrGrArGrCrGrGrUrArArUrGrGrCrUrGrGrUrGrGrCrCrCrA





rGrCrUrU*mC*mC*mU






Additional exemplary template RNAs that could be utilized in this experiment include the following:









FAH1 RNACS050


(SEQ ID NO: 19539)


mG*mG*mA*rUrGrGrUrCrCrUrCrArUrGrArArCrGrArCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





ArGrGrCrArUrUrArCrCrGrCrUrCrCrArGrUrCrGrUrUrCrArU





rGrArG*mG*mA*mC





FAH1 RNACS051


(SEQ ID NO: 19540)


mG*mG*mA*rUrGrGrUrCrCrUrCrArUrGrArArCrGrArCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





ArGrGrCrArUrUrArCrCrGrCrUrCrCrArGrUrCrGrUrUrCrArU





rG*mA*mG*mG






In the sequences above m=2′-O-methyl ribonucleotide, r=ribose and *=phosphorothioate bond.


The gene modifying polypeptides tested comprised sequence of: RNAV209 (nCas9-RT) and RNAV214 (wtCas9-RT). Specifically, the nCas9-RT and the wtCas9-RT had the following amino acid sequences:










nCas9-RT (RNAV209):



(SEQ ID NO: 19541)



MPAAKRVKLDGGDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLI






GALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVE





EDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHF





LIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQL





PGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYAD





LFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYK





EIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNG





SIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKS





EETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVK





YVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRF





NASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKV





MKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKE





DIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPENIVIEMAR





ENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMY





VDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPSEEVVKKMKN





YWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMN





TKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIK





KYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIRK





RPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKL





IARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNP





IDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYL





ASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHR





DKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRI





DLSQLGGDSGGSSGGSSGSETPGTSESATPESSGGSSGGSSTLNIEDEYRLHETSKEPDVS





LGSTWLSDFPQAWAETGGMGLAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQR





LLDQGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGL





PPSHQWYTVLDLKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTL





FNEALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNLGYRASAK





KAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQLREFLGKAGFCRLFIPG





FAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQALLTAPALGLPDLTKPFELFVDEKQG





YAKGVLTQKLGPWRRPVAYLSKKLDPVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPL





VILAPHAVEALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEG





LQHNCLDILAEAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIW





AKALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRRRGWLTS





EGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNRMADQAARKAAITETPD





TSTLLIENSSPSGGSKRTADGSEFEKRTADGSEFESPKKKAKVE





wtCas9-RT (RNAV214);


(SEQ ID NO: 19542)



MPAAKRVKLDGGDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLI






GALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVE





EDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHF





LIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQL





PGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYAD





LFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYK





EIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNG





SIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKS





EETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVK





YVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRF





NASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKV





MKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKE





DIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPENIVIEMAR





ENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMY





VDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKN





YWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMN





TKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIK





KYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIRK





RPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKL





IARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNP





IDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYL





ASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHR





DKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRI





DLSQLGGDSGGSSGGSSGSETPGTSESATPESSGGSSGGSSTLNIEDEYRLHETSKEPDVS





LGSTWLSDFPQAWAETGGMGLAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQR





LLDQGILVPCQSPWNTPLLPVKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGL





PPSHQWYTVLDLKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTL





FNEALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNLGYRASAK





KAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTPKTPRQLREFLGKAGFCRLFIPG





FAEMAAPLYPLTKPGTLFNWGPDQQKAYQEIKQALLTAPALGLPDLTKPFELFVDEKQG





YAKGVLTQKLGPWRRPVAYLSKKLDPVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPL





VILAPHAVEALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEG





LQHNCLDILAEAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIW





AKALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRRRGWLTS





EGKEIKNKDEILALLKALFLPKRLSIIHCPGHQKGHSAEARGNRMADQAARKAAITETPD





TSTLLIENSSPSGGSKRTADGSEFEKRTADGSEFESPKKKAKVE 







Underlining indicates the residue that differs between the nickase and wild-type sequences.


The gene modifying system comprising the gene modifying polypeptides listed above and the template RNA described above were transfected into primary mouse hepatocytes. The gene modifying polypeptide and the template RNA were delivered by nucleofection in the RNA format. Specifically, 4 μg of gene modifying polypeptide mRNA were combined with 10 μg of chemically synthesized template RNA in 5 μL of water. The transfection mix was added to 100,000 mouse primary hepatocytes in Buffer P3 [Lonza], and cells were nucleofected using program DG-138. After nucleofection, cells were grown at 37° C., 5% CO2 for 3 days prior to cell lysis and genomic DNA extraction. To analyze gene editing activity, primers flanking the target insertion site locus were used to amplify across the locus. Amplicons were analyzed via short read sequencing using an Illumina MiSeq. Conversion of terminal A to G sequence in exon 8 of fah gene indicates successful editing.


As shown in FIG. 2, for FAH2 templates, perfect rewrite levels (conversion of A to G with no unwanted mutations detected) of 4-8% were detected with RNAV209-013 but not with RNAV214-040. Indel levels of 4.4 to 6.6% were observed with RNAV209-013. Furthermore, the amount of WT Fah mRNA was measured using quantitative RT-PCR using primers that bind to exons 7 and 8. As shown in FIG. 3, FAH2 templates result in an increase in the abundance of Fah mRNA relative to WT by up to 12% when FAH2 template is tested with RNAV209-013 mRNA. These results demonstrate the use of a gene modifying system to reverse a mutation in the Fah gene, resulting in partial restoration of the expression of wild-type Fah mRNA.


Example 6: Quantifying Activity of a Gene Editing Polypeptide and Template In Vivo for Rewriting the Endogenous FAH Locus Achieved in Mouse Liver

This example demonstrates the use of a gene modifying system containing a gene modifying polypeptide and a template RNA, to convert an A nucleotide to a G nucleotide in the Fah5981SB mouse model into the endogenous Fah locus in mouse liver. The Fah5981SB mouse model harbors a G to A point mutation in the last nucleotide of exon 8 of the Fah gene, leading to aberrant mRNA splicing and subsequent mRNA degradation, without the production of Fah protein and serves as a mouse model of hereditary tyrosinemia type I.


In this example, the template RNA contained:

    • (1) a gRNA spacer;
    • (2) a gRNA scaffold;
    • (3) a heterologous object sequence; and
    • (4) a primer binding site (PBS) sequence.


More specifically, the template RNA comprised the following sequences:









FAH1_R14_P12_Heavy RNACS048-001


(SEQ ID NO: 19543)


mG*mG*mA*rUrGrGrUrCrCrUrCrArUrGrArArCrGrArCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





UrUrArCrCrGrCrUrCrCrArGrUrCrGrUrUrCrArUrGrArG*mG*





mA*mC





FAH1_R15_P10_Heavy RNACS049-001


(SEQ ID NO: 19544)


mG*mG*mA*rUrGrGrUrCrCrUrCrArUrGrArArCrGrArCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





ArUrUrArCrCrGrCrUrCrCrArGrUrCrGrUrUrCrArUrG*mA*mG





*mG





FAH2_R19_P11_MUT_Heavy RNACS052-001


(SEQ ID NO: 19545)


mU*mC*mA*rGrArGrGrArArGrCrUrGrGrGrCrCrArCrCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





UrGrGrArGrCrGrGrUrArArUrGrGrCrUrGrGrUrGrGrCrCrCrA





rGrC*mU*mU*mC 





FAH2_R19_P13_MUT_Heavy RNACS053-001


(SEQ ID NO: 19546)


mU*mC*mA*rGrArGrGrArArGrCrUrGrGrGrCrCrArCrCrGrUrU





rUrUrArGrAmGmCmUmAmGmAmAmAmUmAmGmCrArArGrUrUrArAr





ArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrAmAmCmUmU





mGmAmAmAmAmAmGmUmGmGmCmAmCmCmGmAmGmUmCmGmGmUmGmCr





UrGrGrArGrCrGrGrUrArArUrGrGrCrUrGrGrUrGrGrCrCrCrA





rGrCrUrU*mC*mC*mU






The gene modifying polypeptides tested comprised a sequence of: RNAV209 and RNAV214, the sequences of which are each provided in Example 3.


The gene modifying system comprising the gene modifying polypeptides and the template RNA described above was formulated in LNP and delivered to mice. Specifically, 2 mg/kg of total RNA equivalent formulated in LNPs, combined at 1:1 (w/w) of template RNA and mRNA, were dosed intravenously in 7 to 9-week-old, mixed gender Fah5981SB mice. Six hours or 6 days post-dosing, animals were sacrificed, and their liver collected for analyses. To determine the expression distribution of the gene modifying polypeptide in the liver, 6-hr liver samples were subjected to immunohistochemistry using an anti-Cas9 antibody. Upon staining, quantification of Cas9-positive hepatocytes was determined by QuPath Markup. As shown in FIG. 4, the expression of the gene modifying polypeptide was observed in 82-91% of hepatocytes.


To analyze gene editing activity, primers flanking the target insertion site locus were used to amplify across the locus in the genomic DNA of liver samples collected 6 days post-dosing. Amplicons were analyzed via short read sequencing using an Illumina MiSeq. Conversion of an A nucleotide to a G nucleotide indicates successful editing. As shown in FIG. 5, perfect rewrite levels (conversion of A to G with no unwanted mutations detected) of 0.1%-1.9% were detected across the different groups. Indel levels were in the range of 0.2%-0.4%.


To determine the phenotypic correction caused by the gene editing activity, the restoration of wild-type FAH mRNA was determined by real-time qRT-PCR, and the restoration of Fah protein expression determined by immunohistochemistry using an anti-Fah antibody. As shown in



FIG. 6, wild-type mRNA restoration of 0.1%-6%, relative to littermate heterozygous mice, was detected across the different groups. As shown in FIG. 7, Fah protein was detected in 0.1%-7% of liver cross-sectional area across the different groups. These results demonstrate the use of a gene modifying system to reverse a mutation in the Fah gene in an in vivo mouse model for hereditary tyrosinemia type I, resulting in partial restoration of expression of wild-type Fah mRNA and Fah protein.


Example 7. Gene Editing at the TTR Locus in an In Vivo Mouse Model

This Example demonstrates successful delivery of an mRNA and guide using Cas9-mediated gene editing using the protospacer sequence ACACAAAUACCAGUCCAGCG (SEQ ID NO: 25696) that targets the TTR locus using a gene modifying polypeptide and RNA in a C57Blk/6 mouse.


RNAs were prepared as follows. An mRNA encoding a gene modifying polypeptide having the sequence shown in Table 7A below was produced by in vitro transcription and the purified mRNA was dissolved in 1 mM sodium citrate, pH 6, to a final concentration of RNA of 1-2 mg/mL. Similarly, a guide RNA having a sequence shown in Table 7A below was produced by chemical synthesis and dissolved in water or aqueous buffer, to a final concentration of RNA of 1-2 mg/mL.









TABLE 7A







Sequences of Example 7











SEQ


Name
Nucleic acid sequence
ID NO





Cas9-RT
AUGCCUGCGGCUAAGCGGGUAAAAUUGGAUG
19547


gene
GUGGGGACAAGAAGUACAGCAUCGGCCUGGA



modifying
CAUCGGCACCAACUCUGUGGGCUGGGCCGUG



polypeptide
AUCACCGACGAGUACAAGGUGCCCAGCAAGA




AAUUCAAGGUGCUGGGCAACACCGACCGGCA




CAGCAUCAAGAAGAACCUGAUCGGAGCCCUG




CUGUUCGACAGCGGCGAAACAGCCGAGGCCA




CCCGGCUGAAGAGAACCGCCAGAAGAAGAUA




CACCAGACGGAAGAACCGGAUCUGCUAUCUG




CAAGAGAUCUUCAGCAACGAGAUGGCCAAGG




UGGACGACAGCUUCUUCCACAGACUGGAAGA




GUCCUUCCUGGUGGAAGAGGAUAAGAAGCAC




GAGCGGCACCCCAUCUUCGGCAACAUCGUGG




ACGAGGUGGCCUACCACGAGAAGUACCCCAC




CAUCUACCACCUGAGAAAGAAACUGGUGGAC




AGCACCGACAAGGCCGACCUGCGGCUGAUCU




AUCUGGCCCUGGCCCACAUGAUCAAGUUCCG




GGGCCACUUCCUGAUCGAGGGCGACCUGAAC




CCCGACAACAGCGACGUGGACAAGCUGUUCA




UCCAGCUGGUGCAGACCUACAACCAGCUGUU




CGAGGAAAACCCCAUCAACGCCAGCGGCGUG




GACGCCAAGGCCAUCCUGUCUGCCAGACUGA




GCAAGAGCAGACGGCUGGAAAAUCUGAUCGC




CCAGCUGCCCGGCGAGAAGAAGAAUGGCCUG




UUCGGAAACCUGAUUGCCCUGAGCCUGGGCC




UGACCCCCAACUUCAAGAGCAACUUCGACCU




GGCCGAGGAUGCCAAACUGCAGCUGAGCAAG




GACACCUACGACGACGACCUGGACAACCUGC




UGGCCCAGAUCGGCGACCAGUACGCCGACCU




GUUUCUGGCCGCCAAGAACCUGUCCGACGCC




AUCCUGCUGAGCGACAUCCUGAGAGUGAACA




CCGAGAUCACCAAGGCCCCCCUGAGCGCCUCU




AUGAUCAAGAGAUACGACGAGCACCACCAGG




ACCUGACCCUGCUGAAAGCUCUCGUGCGGCA




GCAGCUGCCUGAGAAGUACAAAGAGAUUUUC




UUCGACCAGAGCAAGAACGGCUACGCCGGCU




ACAUUGACGGCGGAGCCAGCCAGGAAGAGUU




CUACAAGUUCAUCAAGCCCAUCCUGGAAAAG




AUGGACGGCACCGAGGAACUGCUCGUGAAGC




UGAACAGAGAGGACCUGCUGCGGAAGCAGCG




GACCUUCGACAACGGCAGCAUCCCCCACCAGA




UCCACCUGGGAGAGCUGCACGCCAUUCUGCG




GCGGCAGGAAGAUUUUUACCCAUUCCUGAAG




GACAACCGGGAAAAGAUCGAGAAGAUCCUGA




CCUUCCGCAUCCCCUACUACGUGGGCCCUCUG




GCCAGGGGAAACAGCAGAUUCGCCUGGAUGA




CCAGAAAGAGCGAGGAAACCAUCACCCCCUG




GAACUUCGAGGAAGUGGUGGACAAGGGCGCU




UCCGCCCAGAGCUUCAUCGAGCGGAUGACCA




ACUUCGAUAAGAACCUGCCCAACGAGAAGGU




GCUGCCCAAGCACAGCCUGCUGUACGAGUAC




UUCACCGUGUAUAACGAGCUGACCAAAGUGA




AAUACGUGACCGAGGGAAUGAGAAAGCCCGC




CUUCCUGAGCGGCGAGCAGAAAAAGGCCAUC




GUGGACCUGCUGUUCAAGACCAACCGGAAAG




UGACCGUGAAGCAGCUGAAAGAGGACUACUU




CAAGAAAAUCGAGUGCUUCGACUCCGUGGAA




AUCUCCGGCGUGGAAGAUCGGUUCAACGCCU




CCCUGGGCACAUACCACGAUCUGCUGAAAAU




UAUCAAGGACAAGGACUUCCUGGACAAUGAG




GAAAACGAGGACAUUCUGGAAGAUAUCGUGC




UGACCCUGACACUGUUUGAGGACAGAGAGAU




GAUCGAGGAACGGCUGAAAACCUAUGCCCAC




CUGUUCGACGACAAAGUGAUGAAGCAGCUGA




AGCGGCGGAGAUACACCGGCUGGGGCAGGCU




GAGCCGGAAGCUGAUCAACGGCAUCCGGGAC




AAGCAGUCCGGCAAGACAAUCCUGGAUUUCC




UGAAGUCCGACGGCUUCGCCAACAGAAACUU




CAUGCAGCUGAUCCACGACGACAGCCUGACC




UUUAAAGAGGACAUCCAGAAAGCCCAGGUGU




CCGGCCAGGGCGAUAGCCUGCACGAGCACAU




UGCCAAUCUGGCCGGCAGCCCCGCCAUUAAG




AAGGGCAUCCUGCAGACAGUGAAGGUGGUGG




ACGAGCUCGUGAAAGUGAUGGGCCGGCACAA




GCCCGAGAACAUCGUGAUCGAAAUGGCCAGA




GAGAACCAGACCACCCAGAAGGGACAGAAGA




ACAGCCGCGAGAGAAUGAAGCGGAUCGAAGA




GGGCAUCAAAGAGCUGGGCAGCCAGAUCCUG




AAAGAACACCCCGUGGAAAACACCCAGCUGC




AGAACGAGAAGCUGUACCUGUACUACCUGCA




GAAUGGGCGGGAUAUGUACGUGGACCAGGAA




CUGGACAUCAACCGGCUGUCCGACUACGAUG




UGGACCAUAUCGUGCCUCAGAGCUUUCUGAA




GGACGACUCCAUCGACAACAAGGUGCUGACC




AGAAGCGACAAGAAUCGGGGCAAGAGCGACA




ACGUGCCCUCCGAAGAGGUCGUGAAGAAGAU




GAAGAACUACUGGCGGCAGCUGCUGAACGCC




AAGCUGAUUACCCAGAGAAAGUUCGACAAUC




UGACCAAGGCCGAGAGAGGCGGCCUGAGCGA




ACUGGAUAAGGCCGGCUUCAUCAAGAGACAG




CUGGUGGAAACCCGGCAGAUCACAAAGCACG




UGGCACAGAUCCUGGACUCCCGGAUGAACAC




UAAGUACGACGAGAAUGACAAGCUGAUCCGG




GAAGUGAAAGUGAUCACCCUGAAGUCCAAGC




UGGUGUCCGAUUUCCGGAAGGAUUUCCAGUU




UUACAAAGUGCGCGAGAUCAACAACUACCAC




CACGCCCACGACGCCUACCUGAACGCCGUCGU




GGGAACCGCCCUGAUCAAAAAGUACCCUAAG




CUGGAAAGCGAGUUCGUGUACGGCGACUACA




AGGUGUACGACGUGCGGAAGAUGAUCGCCAA




GAGCGAGCAGGAAAUCGGCAAGGCUACCGCC




AAGUACUUCUUCUACAGCAACAUCAUGAACU




UUUUCAAGACCGAGAUUACCCUGGCCAACGG




CGAGAUCCGGAAGCGGCCUCUGAUCGAGACA




AACGGCGAAACCGGGGAGAUCGUGUGGGAUA




AGGGCCGGGAUUUUGCCACCGUGCGGAAAGU




GCUGAGCAUGCCCCAAGUGAAUAUCGUGAAA




AAGACCGAGGUGCAGACAGGCGGCUUCAGCA




AAGAGUCUAUCCUGCCCAAGAGGAACAGCGA




UAAGCUGAUCGCCAGAAAGAAGGACUGGGAC




CCUAAGAAGUACGGCGGCUUCGACAGCCCCA




CCGUGGCCUAUUCUGUGCUGGUGGUGGCCAA




AGUGGAAAAGGGCAAGUCCAAGAAACUGAAG




AGUGUGAAAGAGCUGCUGGGGAUCACCAUCA




UGGAAAGAAGCAGCUUCGAGAAGAAUCCCAU




CGACUUUCUGGAAGCCAAGGGCUACAAAGAA




GUGAAAAAGGACCUGAUCAUCAAGCUGCCUA




AGUACUCCCUGUUCGAGCUGGAAAACGGCCG




GAAGAGAAUGCUGGCCUCUGCCGGCGAACUG




CAGAAGGGAAACGAACUGGCCCUGCCCUCCA




AAUAUGUGAACUUCCUGUACCUGGCCAGCCA




CUAUGAGAAGCUGAAGGGCUCCCCCGAGGAU




AAUGAGCAGAAACAGCUGUUUGUGGAACAGC




ACAAGCACUACCUGGACGAGAUCAUCGAGCA




GAUCAGCGAGUUCUCCAAGAGAGUGAUCCUG




GCCGACGCUAAUCUGGACAAAGUGCUGUCCG




CCUACAACAAGCACCGGGAUAAGCCCAUCAG




AGAGCAGGCCGAGAAUAUCAUCCACCUGUUU




ACCCUGACCAAUCUGGGAGCCCCUGCCGCCUU




CAAGUACUUUGACACCACCAUCGACCGGAAG




AGGUACACCAGCACCAAAGAGGUGCUGGACG




CCACCCUGAUCCACCAGAGCAUCACCGGCCUG




UACGAGACACGGAUCGACCUGUCUCAGCUGG




GAGGUGACUCUGGAGGAUCUAGCGGAGGAUC




CUCUGGCAGCGAGACACCAGGAACAAGCGAG




UCAGCAACACCAGAGAGCAGUGGCGGCAGCA




GCGGCGGCAGCAGCACCCUAAAUAUAGAAGA




UGAGUAUCGGCUACAUGAGACCUCAAAAGAG




CCAGAUGUUUCUCUAGGGUCCACAUGGCUGU




CUGAUUUUCCUCAGGCCUGGGCGGAAACCGG




GGGCAUGGGACUGGCAGUUCGCCAAGCUCCU




CUGAUCAUACCUCUGAAAGCAACCUCUACCC




CCGUGUCCAUAAAACAAUACCCCAUGUCACA




AGAAGCCAGACUGGGGAUCAAGCCCCACAUA




CAGAGACUGUUGGACCAGGGAAUACUGGUAC




CCUGCCAGUCCCCCUGGAACACGCCCCUGCUA




CCCGUUAAGAAACCAGGGACUAAUGAUUAUA




GGCCUGUCCAGGAUCUGAGAGAAGUCAACAA




GCGGGUGGAGGACAUCCACCCCACCGUGCCC




AACCCUUACAACCUCUUGAGCGGGCUCCCACC




GUCCCACCAGUGGUACACUGUGCUUGAUUUA




AAGGAUGCCUUUUUCUGCCUGAGACUCCACC




CCACCAGUCAGCCUCUCUUCGCCUUUGAGUG




GAGAGAUCCAGAGAUGGGAAUCUCAGGACAA




UUGACCUGGACCAGACUCCCACAGGGUUUCA




AAAACAGUCCCACCCUGUUUAAUGAGGCACU




GCACAGAGACCUAGCAGACUUCCGGAUCCAG




CACCCAGACUUGAUCCUGCUACAGUACGUGG




AUGACUUACUGCUGGCCGCCACUUCUGAGCU




AGACUGCCAACAAGGUACUCGGGCCCUGUUA




CAAACCCUAGGGAACCUCGGGUAUCGGGCCU




CGGCCAAGAAAGCCCAAAUUUGCCAGAAACA




GGUCAAGUAUCUGGGGUAUCUUCUAAAAGAG




GGUCAGAGAUGGCUGACUGAGGCCAGAAAAG




AGACUGUGAUGGGGCAGCCUACUCCGAAGAC




CCCUCGACAACUAAGGGAGUUCCUAGGGAAG




GCAGGCUUCUGUCGCCUCUUCAUCCCUGGGU




UUGCAGAAAUGGCAGCCCCCCUGUACCCUCU




CACCAAACCGGGGACUCUGUUUAAUUGGGGC




CCAGACCAACAAAAGGCCUAUCAAGAAAUCA




AGCAAGCCCUUCUAACUGCCCCAGCCCUGGG




GUUGCCAGAUUUGACUAAGCCCUUUGAACUC




UUUGUCGACGAGAAGCAGGGCUACGCCAAAG




GUGUCCUAACGCAAAAACUGGGACCUUGGCG




UCGGCCGGUGGCCUACCUGUCCAAAAAGCUA




GACCCAGUAGCAGCUGGGUGGCCCCCUUGCC




UACGGAUGGUAGCAGCCAUUGCCGUACUGAC




AAAGGAUGCAGGCAAGCUAACCAUGGGACAG




CCACUAGUCAUUCUGGCCCCCCAUGCAGUAG




AGGCACUAGUCAAACAACCCCCCGACCGCUG




GCUUUCCAACGCCCGGAUGACUCACUAUCAG




GCCUUGCUUUUGGACACGGACCGGGUCCAGU




UCGGACCGGUGGUAGCCCUGAACCCGGCUAC




GCUGCUCCCACUGCCUGAGGAAGGGCUGCAA




CACAACUGCCUUGAUAUCCUGGCCGAAGCCC




ACGGAACCCGACCCGACCUAACGGACCAGCCG




CUCCCAGACGCCGACCACACCUGGUACACGGA




UGGAAGCAGUCUCUUACAAGAGGGACAGCGU




AAGGCGGGAGCUGCGGUGACCACCGAGACCG




AGGUAAUCUGGGCUAAAGCCCUGCCAGCCGG




GACAUCCGCUCAGCGGGCUGAACUGAUAGCA




CUCACCCAGGCCCUAAAGAUGGCAGAAGGUA




AGAAGCUAAAUGUUUAUACUGAUAGCCGUUA




UGCUUUUGCUACUGCCCAUAUCCAUGGAGAA




AUAUACAGAAGGCGUGGGUGGCUCACAUCAG




AAGGCAAAGAGAUCAAAAAUAAAGACGAGAU




CUUGGCCCUACUAAAAGCCCUCUUUCUGCCC




AAAAGACUUAGCAUAAUCCAUUGUCCAGGAC




AUCAAAAGGGACACAGCGCCGAGGCUAGAGG




CAACCGGAUGGCUGACCAAGCGGCCCGAAAG




GCAGCCAUCACAGAGACUCCAGACACCUCUA




CCCUCCUCAUAGAAAAUUCAUCACCCUCUGG




CGGCUCAAAAAGAACCGCCGACGGCAGCGAA




UUCGAGAAAAGGACGGCGGAUGGUAGCGAAU




UCGAGAGCCCUAAAAAGAAGGCCAAGGUAGA




GUAA






guide RNA
mA*mC*mA*CAAAUACCAGUCCAGCGGUUUUA
19548



GAmGmCmUmAmGmAmAmAmUmAmGmCAAGU




UAAAAUAAGGCUAGUCCGUUAUCAmAmCmUm




UmGmAmAmAmAmAmGmUmGmGmCmAmCmCm




GmAmGmUmCmGmGmUmGmCmU*mU*mU*mU




m = 2'OMethyl, * = phosphorothioate linkage









Lipid nanoparticle (LNP) components (ionizable lipid, helper lipid, sterol, PEG) were dissolved in 100% ethanol with the lipid component molar ratios of 47:8:43.5:1.5, respectively. RNA (guide and mRNA) was combined in a 1:1 weight ratio and diluted to a concentration of 0.05-0.2 mg/mL in sodium acetate buffer, pH 5. RNA was formulated into distinct LNPs with a lipid amine to total RNA phosphate (N:P) molar ratio of 4.0. The LNPs were formed by microfluidic or turbulent mixing of the lipid and RNA solutions. A 3:1 ratio of aqueous to organic solvent was maintained during mixing using differential flow rates. After mixing, the LNPs were diluted, collected and buffer exchanged into 50 mM Tris, 9% sucrose buffer using tangential flow filtration. Formulations were concentrated to 1.0 mg/mL or higher then filtered through 0.2 μm sterile filter. The final LNP were stored at −80° C. until further use.


The LNP formulations were delivered intravenously by bolus tail vein injection to C57Blk/6 mice that were approximately 8 weeks old at concentrations ranging from 1-0.1 mg/kg. The expression of the Cas9-RT was measured by 6 hours after injection by euthanizing animals and collecting livers during necropsy. Animals were euthanized at 5 days after injection where liver was collected upon necropsy to which the activity of gene editing of the TTR locus was assessed. Expression of the Cas9-RT gene editing polypeptide in liver was measured by Western blot where Cas9 was detected by a mouse monoclonal antibody (7A9-3A3, Cell Signaling Technology) and GAPDH (Cell Signaling Technology) was used as a loading control. (FIG. 8). Editing of the TTR locus was quantified by Sanger sequencing followed by TIDE analysis of an amplicon of the TTR locus near the binding site of the protospacer. Editing of the TTR locus was observed, as shown in FIG. 9. TTR protein levels in serum were quantified by an ELISA using a standard curve (Aviva Biosciences). TTR protein levels in serum declined in treated animals, as shown in FIG. 10. These experiments demonstrate that the Cas9-RT polypeptide can be expressed in vivo, and can edit the TTR locus, resulting in a decrease in TTR protein levels in serum.


Example 8. Gene Editing at the TTR Locus in an In Vivo Cynomolgus Macaque Model

This Example demonstrates successful delivery of an mRNA and guide using Cas9-mediated gene editing using the protospacer sequence ACACAAAUACCAGUCCAGCG (SEQ ID NO: 25696) that targets the TTR locus using a gene modifying polypeptide and RNA in a cynomolgus model.


RNAs were prepared as follows. An mRNA encoding a gene modifying polypeptide having the sequence shown in Table 8A below was produced by in vitro transcription and the purified mRNA was dissolved in 1 mM sodium citrate, pH 6, to a final concentration of RNA of 1-2 mg/mL. Similarly, a guide RNA having a sequence shown in Table 8A below was produced by chemical synthesis and dissolved in water or aqueous buffer, to a final concentration of RNA of 1-2 mg/mL.









TABLE 8A







Sequences of Example 8











SEQ ID


Name
Nucleic acid sequence
NO





Cas9-RT gene
AUGCCUGCGGCUAAGCGGGUAAAAUUGGAUGGUGGGGACA
19549


modifying
AGAAGUACAGCAUCGGCCUGGACAUCGGCACCAACUCUGUG



polypeptide
GGCUGGGCCGUGAUCACCGACGAGUACAAGGUGCCCAGCAA




GAAAUUCAAGGUGCUGGGCAACACCGACCGGCACAGCAUCA




AGAAGAACCUGAUCGGAGCCCUGCUGUUCGACAGCGGCGAA




ACAGCCGAGGCCACCCGGCUGAAGAGAACCGCCAGAAGAAG




AUACACCAGACGGAAGAACCGGAUCUGCUAUCUGCAAGAGA




UCUUCAGCAACGAGAUGGCCAAGGUGGACGACAGCUUCUUC




CACAGACUGGAAGAGUCCUUCCUGGUGGAAGAGGAUAAGA




AGCACGAGCGGCACCCCAUCUUCGGCAACAUCGUGGACGAG




GUGGCCUACCACGAGAAGUACCCCACCAUCUACCACCUGAG




AAAGAAACUGGUGGACAGCACCGACAAGGCCGACCUGCGGC




UGAUCUAUCUGGCCCUGGCCCACAUGAUCAAGUUCCGGGGC




CACUUCCUGAUCGAGGGCGACCUGAACCCCGACAACAGCGA




CGUGGACAAGCUGUUCAUCCAGCUGGUGCAGACCUACAACC




AGCUGUUCGAGGAAAACCCCAUCAACGCCAGCGGCGUGGAC




GCCAAGGCCAUCCUGUCUGCCAGACUGAGCAAGAGCAGACG




GCUGGAAAAUCUGAUCGCCCAGCUGCCCGGCGAGAAGAAGA




AUGGCCUGUUCGGAAACCUGAUUGCCCUGAGCCUGGGCCUG




ACCCCCAACUUCAAGAGCAACUUCGACCUGGCCGAGGAUGC




CAAACUGCAGCUGAGCAAGGACACCUACGACGACGACCUGG




ACAACCUGCUGGCCCAGAUCGGCGACCAGUACGCCGACCUG




UUUCUGGCCGCCAAGAACCUGUCCGACGCCAUCCUGCUGAG




CGACAUCCUGAGAGUGAACACCGAGAUCACCAAGGCCCCCC




UGAGCGCCUCUAUGAUCAAGAGAUACGACGAGCACCACCAG




GACCUGACCCUGCUGAAAGCUCUCGUGCGGCAGCAGCUGCC




UGAGAAGUACAAAGAGAUUUUCUUCGACCAGAGCAAGAAC




GGCUACGCCGGCUACAUUGACGGCGGAGCCAGCCAGGAAGA




GUUCUACAAGUUCAUCAAGCCCAUCCUGGAAAAGAUGGACG




GCACCGAGGAACUGCUCGUGAAGCUGAACAGAGAGGACCUG




CUGCGGAAGCAGCGGACCUUCGACAACGGCAGCAUCCCCCA




CCAGAUCCACCUGGGAGAGCUGCACGCCAUUCUGCGGCGGC




AGGAAGAUUUUUACCCAUUCCUGAAGGACAACCGGGAAAA




GAUCGAGAAGAUCCUGACCUUCCGCAUCCCCUACUACGUGG




GCCCUCUGGCCAGGGGAAACAGCAGAUUCGCCUGGAUGACC




AGAAAGAGCGAGGAAACCAUCACCCCCUGGAACUUCGAGGA




AGUGGUGGACAAGGGCGCUUCCGCCCAGAGCUUCAUCGAGC




GGAUGACCAACUUCGAUAAGAACCUGCCCAACGAGAAGGUG




CUGCCCAAGCACAGCCUGCUGUACGAGUACUUCACCGUGUA




UAACGAGCUGACCAAAGUGAAAUACGUGACCGAGGGAAUG




AGAAAGCCCGCCUUCCUGAGCGGCGAGCAGAAAAAGGCCAU




CGUGGACCUGCUGUUCAAGACCAACCGGAAAGUGACCGUGA




AGCAGCUGAAAGAGGACUACUUCAAGAAAAUCGAGUGCUU




CGACUCCGUGGAAAUCUCCGGCGUGGAAGAUCGGUUCAACG




CCUCCCUGGGCACAUACCACGAUCUGCUGAAAAUUAUCAAG




GACAAGGACUUCCUGGACAAUGAGGAAAACGAGGACAUUC




UGGAAGAUAUCGUGCUGACCCUGACACUGUUUGAGGACAG




AGAGAUGAUCGAGGAACGGCUGAAAACCUAUGCCCACCUGU




UCGACGACAAAGUGAUGAAGCAGCUGAAGCGGCGGAGAUA




CACCGGCUGGGGCAGGCUGAGCCGGAAGCUGAUCAACGGCA




UCCGGGACAAGCAGUCCGGCAAGACAAUCCUGGAUUUCCUG




AAGUCCGACGGCUUCGCCAACAGAAACUUCAUGCAGCUGAU




CCACGACGACAGCCUGACCUUUAAAGAGGACAUCCAGAAAG




CCCAGGUGUCCGGCCAGGGCGAUAGCCUGCACGAGCACAUU




GCCAAUCUGGCCGGCAGCCCCGCCAUUAAGAAGGGCAUCCU




GCAGACAGUGAAGGUGGUGGACGAGCUCGUGAAAGUGAUG




GGCCGGCACAAGCCCGAGAACAUCGUGAUCGAAAUGGCCAG




AGAGAACCAGACCACCCAGAAGGGACAGAAGAACAGCCGCG




AGAGAAUGAAGCGGAUCGAAGAGGGCAUCAAAGAGCUGGG




CAGCCAGAUCCUGAAAGAACACCCCGUGGAAAACACCCAGC




UGCAGAACGAGAAGCUGUACCUGUACUACCUGCAGAAUGG




GCGGGAUAUGUACGUGGACCAGGAACUGGACAUCAACCGGC




UGUCCGACUACGAUGUGGACCAUAUCGUGCCUCAGAGCUUU




CUGAAGGACGACUCCAUCGACAACAAGGUGCUGACCAGAAG




CGACAAGAAUCGGGGCAAGAGCGACAACGUGCCCUCCGAAG




AGGUCGUGAAGAAGAUGAAGAACUACUGGCGGCAGCUGCU




GAACGCCAAGCUGAUUACCCAGAGAAAGUUCGACAAUCUGA




CCAAGGCCGAGAGAGGCGGCCUGAGCGAACUGGAUAAGGCC




GGCUUCAUCAAGAGACAGCUGGUGGAAACCCGGCAGAUCAC




AAAGCACGUGGCACAGAUCCUGGACUCCCGGAUGAACACUA




AGUACGACGAGAAUGACAAGCUGAUCCGGGAAGUGAAAGU




GAUCACCCUGAAGUCCAAGCUGGUGUCCGAUUUCCGGAAGG




AUUUCCAGUUUUACAAAGUGCGCGAGAUCAACAACUACCAC




CACGCCCACGACGCCUACCUGAACGCCGUCGUGGGAACCGC




CCUGAUCAAAAAGUACCCUAAGCUGGAAAGCGAGUUCGUG




UACGGCGACUACAAGGUGUACGACGUGCGGAAGAUGAUCG




CCAAGAGCGAGCAGGAAAUCGGCAAGGCUACCGCCAAGUAC




UUCUUCUACAGCAACAUCAUGAACUUUUUCAAGACCGAGAU




UACCCUGGCCAACGGCGAGAUCCGGAAGCGGCCUCUGAUCG




AGACAAACGGCGAAACCGGGGAGAUCGUGUGGGAUAAGGG




CCGGGAUUUUGCCACCGUGCGGAAAGUGCUGAGCAUGCCCC




AAGUGAAUAUCGUGAAAAAGACCGAGGUGCAGACAGGCGG




CUUCAGCAAAGAGUCUAUCCUGCCCAAGAGGAACAGCGAUA




AGCUGAUCGCCAGAAAGAAGGACUGGGACCCUAAGAAGUA




CGGCGGCUUCGACAGCCCCACCGUGGCCUAUUCUGUGCUGG




UGGUGGCCAAAGUGGAAAAGGGCAAGUCCAAGAAACUGAA




GAGUGUGAAAGAGCUGCUGGGGAUCACCAUCAUGGAAAGA




AGCAGCUUCGAGAAGAAUCCCAUCGACUUUCUGGAAGCCAA




GGGCUACAAAGAAGUGAAAAAGGACCUGAUCAUCAAGCUG




CCUAAGUACUCCCUGUUCGAGCUGGAAAACGGCCGGAAGAG




AAUGCUGGCCUCUGCCGGCGAACUGCAGAAGGGAAACGAAC




UGGCCCUGCCCUCCAAAUAUGUGAACUUCCUGUACCUGGCC




AGCCACUAUGAGAAGCUGAAGGGCUCCCCCGAGGAUAAUGA




GCAGAAACAGCUGUUUGUGGAACAGCACAAGCACUACCUGG




ACGAGAUCAUCGAGCAGAUCAGCGAGUUCUCCAAGAGAGU




GAUCCUGGCCGACGCUAAUCUGGACAAAGUGCUGUCCGCCU




ACAACAAGCACCGGGAUAAGCCCAUCAGAGAGCAGGCCGAG




AAUAUCAUCCACCUGUUUACCCUGACCAAUCUGGGAGCCCC




UGCCGCCUUCAAGUACUUUGACACCACCAUCGACCGGAAGA




GGUACACCAGCACCAAAGAGGUGCUGGACGCCACCCUGAUC




CACCAGAGCAUCACCGGCCUGUACGAGACACGGAUCGACCU




GUCUCAGCUGGGAGGUGACUCUGGAGGAUCUAGCGGAGGA




UCCUCUGGCAGCGAGACACCAGGAACAAGCGAGUCAGCAAC




ACCAGAGAGCAGUGGCGGCAGCAGCGGCGGCAGCAGCACCC




UAAAUAUAGAAGAUGAGUAUCGGCUACAUGAGACCUCAAA




AGAGCCAGAUGUUUCUCUAGGGUCCACAUGGCUGUCUGAU




UUUCCUCAGGCCUGGGCGGAAACCGGGGGCAUGGGACUGGC




AGUUCGCCAAGCUCCUCUGAUCAUACCUCUGAAAGCAACCU




CUACCCCCGUGUCCAUAAAACAAUACCCCAUGUCACAAGAA




GCCAGACUGGGGAUCAAGCCCCACAUACAGAGACUGUUGGA




CCAGGGAAUACUGGUACCCUGCCAGUCCCCCUGGAACACGC




CCCUGCUACCCGUUAAGAAACCAGGGACUAAUGAUUAUAGG




CCUGUCCAGGAUCUGAGAGAAGUCAACAAGCGGGUGGAGG




ACAUCCACCCCACCGUGCCCAACCCUUACAACCUCUUGAGC




GGGCUCCCACCGUCCCACCAGUGGUACACUGUGCUUGAUUU




AAAGGAUGCCUUUUUCUGCCUGAGACUCCACCCCACCAGUC




AGCCUCUCUUCGCCUUUGAGUGGAGAGAUCCAGAGAUGGG




AAUCUCAGGACAAUUGACCUGGACCAGACUCCCACAGGGUU




UCAAAAACAGUCCCACCCUGUUUAAUGAGGCACUGCACAGA




GACCUAGCAGACUUCCGGAUCCAGCACCCAGACUUGAUCCU




GCUACAGUACGUGGAUGACUUACUGCUGGCCGCCACUUCUG




AGCUAGACUGCCAACAAGGUACUCGGGCCCUGUUACAAACC




CUAGGGAACCUCGGGUAUCGGGCCUCGGCCAAGAAAGCCCA




AAUUUGCCAGAAACAGGUCAAGUAUCUGGGGUAUCUUCUA




AAAGAGGGUCAGAGAUGGCUGACUGAGGCCAGAAAAGAGA




CUGUGAUGGGGCAGCCUACUCCGAAGACCCCUCGACAACUA




AGGGAGUUCCUAGGGAAGGCAGGCUUCUGUCGCCUCUUCAU




CCCUGGGUUUGCAGAAAUGGCAGCCCCCCUGUACCCUCUCA




CCAAACCGGGGACUCUGUUUAAUUGGGGCCCAGACCAACAA




AAGGCCUAUCAAGAAAUCAAGCAAGCCCUUCUAACUGCCCC




AGCCCUGGGGUUGCCAGAUUUGACUAAGCCCUUUGAACUCU




UUGUCGACGAGAAGCAGGGCUACGCCAAAGGUGUCCUAACG




CAAAAACUGGGACCUUGGCGUCGGCCGGUGGCCUACCUGUC




CAAAAAGCUAGACCCAGUAGCAGCUGGGUGGCCCCCUUGCC




UACGGAUGGUAGCAGCCAUUGCCGUACUGACAAAGGAUGC




AGGCAAGCUAACCAUGGGACAGCCACUAGUCAUUCUGGCCC




CCCAUGCAGUAGAGGCACUAGUCAAACAACCCCCCGACCGC




UGGCUUUCCAACGCCCGGAUGACUCACUAUCAGGCCUUGCU




UUUGGACACGGACCGGGUCCAGUUCGGACCGGUGGUAGCCC




UGAACCCGGCUACGCUGCUCCCACUGCCUGAGGAAGGGCUG




CAACACAACUGCCUUGAUAUCCUGGCCGAAGCCCACGGAAC




CCGACCCGACCUAACGGACCAGCCGCUCCCAGACGCCGACC




ACACCUGGUACACGGAUGGAAGCAGUCUCUUACAAGAGGG




ACAGCGUAAGGCGGGAGCUGCGGUGACCACCGAGACCGAGG




UAAUCUGGGCUAAAGCCCUGCCAGCCGGGACAUCCGCUCAG




CGGGCUGAACUGAUAGCACUCACCCAGGCCCUAAAGAUGGC




AGAAGGUAAGAAGCUAAAUGUUUAUACUGAUAGCCGUUAU




GCUUUUGCUACUGCCCAUAUCCAUGGAGAAAUAUACAGAA




GGCGUGGGUGGCUCACAUCAGAAGGCAAAGAGAUCAAAAA




UAAAGACGAGAUCUUGGCCCUACUAAAAGCCCUCUUUCUGC




CCAAAAGACUUAGCAUAAUCCAUUGUCCAGGACAUCAAAAG




GGACACAGCGCCGAGGCUAGAGGCAACCGGAUGGCUGACCA




AGCGGCCCGAAAGGCAGCCAUCACAGAGACUCCAGACACCU




CUACCCUCCUCAUAGAAAAUUCAUCACCCUCUGGCGGCUCA




AAAAGAACCGCCGACGGCAGCGAAUUCGAGAAAAGGACGGC




GGAUGGUAGCGAAUUCGAGAGCCCUAAAAAGAAGGCCAAG




GUAGAGUAA






guide RNA
mA*mC*mA*CAAAUACCAGUCCAGCGGUUUUAGAmGmCmUm
19550



AmGmAmAmAmUmAmGmCAAGUUAAAAUAAGGCUAGUCCG




UUAUCAmAmCmUmUmGmAmAmAmAmAmGmUmGmGmCmAm




CmCmGmAmGmUmCmGmGmUmGmCmU*mU*mU*mU




m = 2'OMethyl, * = phosphorothioate linkage









Lipid nanoparticle (LNP) components (ionizable lipid, helper lipid, sterol, PEG) were dissolved in 100% ethanol with the lipid component molar ratios of 47:8:43.5:1.5, respectively. RNA (guide and mRNA) was combined in a 1:1 weight ratio and diluted to a concentration of 0.05-0.2 mg/mL in sodium acetate buffer, pH 5. RNA was formulated into distinct LNPs with a lipid amine to total RNA phosphate (N:P) molar ratio of 4.0. The LNPs were formed by microfluidic or turbulent mixing of the lipid and RNA solutions. A 3:1 ratio of aqueous to organic solvent was maintained during mixing using differential flow rates. After mixing, the LNPs were diluted, collected and buffer exchanged into 50 mM Tris, 9% sucrose buffer using tangential flow filtration. Formulations were concentrated to 1.0 mg/mL or higher then filtered through 0.2 μm sterile filter. The final LNP were stored at −80° C. until further use. The LNP formulations were delivered intravenously by infusion over the course of 1 hour at 2 mg/kg where the volume of the infusion was 5 ml/kg. Cynomolgus macaques from mainland Asia were given dexamethasone 2 mg/kg bolus via intramuscular injection 1.5-2 h prior to intravenous infusion using a syringe pump. Animals were monitored after infusion and the expression of the Cas9-RT was measured by laparoscopic biopsies taken from the liver 8-12 h, 24 h, and 48 h after infusion. Animals were euthanized 14 days after infusion and liver was harvested by dividing the organ up into 8 different segments to which the activity of gene editing of the TTR locus was assessed. Expression of the Cas9-RT gene editing polypeptide in liver was quantified by capillary electrophoresis western blot using the ProteinSimple Jess system (bio-techne) where Cas9 was detected by a mouse monoclonal antibody (7A9-3A3, Cell Signaling Technology). Relative expression of the Cas9-RT gene editing polypeptide was measured by an area under curve analysis, as shown in FIG. 11. Editing of the TTR locus was quantified by amplicon-sequencing of the TTR locus near the binding site of the protospacer. Editing of the TTR locus was observed, as shown in FIG. 12. These experiments demonstrate that the Cas9-RT polypeptide can be expressed in vivo in a non-human primate model and can edit the TTR locus.


Example 9. Screening of Cas9 Variant and Spacer Combinations for High indel Activity

This Example characterizes screening experiments conducted to identify Cas9 variants and spacer combinations capable of producing high indel activity. In this example, an sgRNA contains:

    • (1) A gRNA spacer
    • (2) A gRNA scaffold


An initial screen was performed in HEK293T cells using wild type SpCas9 polypeptide variants in combination with template RNAs comprising several spacer sequences, selected for close proximity to the mutation to be corrected. This initial screen evaluated indel activity as an indicator of a spacer's utility for editing the target PiZ mutation. Following this analysis, variant Cas9 domains were used to generate exemplary gene modifying polypeptides comprising the selected Cas9 domains, a linker, and an exemplary RT domain, and the exemplary gene modifying polypeptides were used to screen compatible template RNAs using indel activity.


A gene modifying system comprising either:

    • a) a compatible gene modifying polypeptide described herein (e.g., having: an NLS, a linker, an RT sequence, and a second NLS as recited below in this Example), and a wild type Cas9 having a sequence of Table X1, or
    • (i-b) a compatible wild type Cas9 polypeptide;
    • and (ii) a single guide RNA (sgRNA) (e.g., A1AT-Sp-sgRNA-1)
    • was transfected into the HEK293T landing pad cell line (described in Example 1). The gene modifying polypeptide and the sgRNA or wild type Cas9 polypeptide and the sgRNA were delivered by transfection in DNA format. Specifically, 50 ng of gene modifying polypeptide/Cas9 polypeptide plasmid was combined with 50 of sgRNA. This combination of plasmids was mixed with 0.5 uL of TransIT 293, in 10 uL of OptiMeM solution, and added to 20,000 cells. After transfection, cells were grown at 37° C., 5% CO2 for 3 days prior to cell lysis and genomic DNA extraction. To analyze gene editing activity, primers flanking the A1 AT PiZ mutation site were used to amplify across the locus. Amplicons were analyzed via short read sequencing using an Illumina MiSeq.









SpCas9 spacer sequences used in FIG. 13:


A1AT-Sp-sgRNA-1:


(SEQ ID NO: 23786)


gggtatggcctctaaaaaca (PLV3676) [cut site is 30 bp


from PiZ mutation]





A1AT-Sp-sgRNA-2:


(SEQ ID NO: 23787)


tcccctccaggccgtgcata (PLV3712) [cut site is 23 bp


from PiZ mutation]





A1AT-Sp-sgRNA-3:


(SEQ ID NO: 23788)


tctctgcttctctcccctcc (PLV3735) [cut site is 35 bp


from PiZ mutation]





A1AT-Sp-sgRNA-4:


(SEQ ID NO: 23789)


gtcccctccaggccgtgcata (PLV3690) [cut site is 23


bp from PiZ mutation]





A1AT-Sp-sgRNA-5:


(SEQ ID NO: 23790)


gtctctgcttctctcccctcc (PLV3668) [cut site is 35


bp from PiZ mutation]







Bar 6 in FIG. 13 was a no sgRNA control.


Exemplary gene modifying polypeptides comprising Cas9 variants comprised:


an N-terminal NLS having an amino acid sequence of:











(SEQ ID NO: 23791)



MPAAKRVKLDGGKRTADGSEFESPKKKRKV;







a C-terminal NLS having an amino acid sequence of











(SEQ ID NO: 23792)



KRTADSQHSTPPKTKRKVEFEPKKKRKV;







an RT domain having an amino acid sequence of









(SEQ ID NO: 23793)


TLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAETGGMGLAVRQAPLI





IPLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPWNTPL





LPVKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYT





VLDLKDAFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSP





TLFNEALHRDLADFRIQHPDLILLQYVDDLLLAATSELDCQQGTRALLQ





TLGNLGYRASAKKAQICQKQVKYLGYLLKEGQRWLTEARKETVMGQPTP





KTPRQLREFLGKAGFCRLFIPGFAEMAAPLYPLTKPGTLFNWGPDQQKA





YQEIKQALLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWRRPV





AYLSKKLDPVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVE





ALVKQPPDRWLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEG





LQHNCLDILAEAHGTRPDLTDQPLPDADHTWYTDGSSLLQEGQRKAGAA





VTTETEVIWAKALPAGTSAQRAELIALTQALKMAEGKKLNVYTDSRYAF





ATAHIHGEIYRRRGWLTSEGKEIKNKDEILALLKALFLPKRLSIIHCPG





HQKGHSAEARGNRMADQAARKAAITETPDTSTLLIENSSPSGGSKRTAD





GSEF;







and a linker between the RT domain and Cas domain having an amino acid sequence of











(SEQ ID NO: 25689)



SGGSSGGSSGSETPGTSE







(SEQ ID NO: 23794)



SATPESSGGSSGGSS













TABLE X1







Exemplary Gene Modifying Polypeptide Cas9 Variant Sequences and Identifiers















SEQ


Plasmid
Cas9
Mu-

ID


Number
Variant
tation
Cas9 domain
NO





PLV5191
SpyCas9
WT
DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATR
23795





LKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEV






AYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQL






VQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPN






FKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEIT






KAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKF






IKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNRE






KIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKN






LPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVK






QLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLF






EDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDG






FANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVK






VMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKL






YLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS






EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQ






ILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVG






TALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI






RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLI






ARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFL






EAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYE






KLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQA






ENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD



PLV919

N863A
DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATR
23796





LKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEV






AYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQL






VQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPN






FKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEIT






KAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKF






IKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNRE






KIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKN






LPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVK






QLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLF






EDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDG






FANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVK






VMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKL






YLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS






EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQ






ILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVG






TALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI






RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLI






ARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFL






EAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYE






KLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQA






ENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD






PLV7476
BlatCas9
WT
AYTMGIDVGIASCGWAIVDLERQRIIDIGVRTFEKAENPKNGEALAVPRREARSSRRRLRRK
23797





KHRIERLKHMFVRNGLAVDIQHLEQTLRSQNEIDVWQLRVDGLDRMLTQKEWLRVLIHLA






QRRGFQSNRKTDGSSEDGQVLVNVTENDRLMEEKDYRTVAEMMVKDEKFSDHKRNKNGN






YHGVVSRSSLLVEIHTLFETQRQHHNSLASKDFELEYVNIWSAQRPVATKDQIEKMIGTCTFL






PKEKRAPKASWHFQYFMLLQTINHIRITNVQGTRSLNKEEIEQVVNMALTKSKVSYHDTRKI






LDLSEEYQFVGLDYGKEDEKKKVESKETIIKLDDYHKLNKIFNEVELAKGETWEADDYDTV






AYALTFFKDDEDIRDYLQNKYKDSKNRLVKNLANKEYTNELIGKVSTLSFRKVGHLSLKAL






RKIIPFLEQGMTYDKACQAAGFDFQGISKKKRSVVLPVIDQISNPVVNRALTQTRKVINALIK






KYGSPETIHIETARELSKTFDERKNITKDYKENRDKNEHAKKHLSELGIINPTGLDIVKYKLW






CEQQGRCMYSNQPISFERLKESGYTEVDHIIPYSRSMNDSYNNRVLVMTRENREKGNQTPFE






YMGNDTQRWYEFEQRVTTNPQIKKEKRQNLLLKGFTNRRELEMLERNLNDTRYITKYLSHF






ISTNLEFSPSDKKKKVVNTSGRITSHLRSRWGLEKNRGQNDLHHAMDAIVIAVTSDSFIQQVT






NYYKRKERRELNGDDKFPLPWKFFREEVIARLSPNPKEQIEALPNHFYSEDELADLQPIFVSR






MPKRSITGEAHQAQFRRVVGKTKEGKNITAKKTALVDISYDKNGDFNMYGRETDPATYEAI






KERYLEFGGNVKKAFSTDLHKPKKDGTKGPLIKSVRIMENKTLVHPVNKGKGVVYNSSIVR






TDVFQRKEKYYLLPVYVTDVTKGKLPNKVIVAKKGYHDWIEVDDSFTFLFSLYPNDLIFIRQ






NPKKKISLKKRIESHSISDSKEVQEIHAYYKGVDSSTAAIEFIIHDGSYYAKGVGVQNLDCFEK






YQVDILGNYFKVKGEKRLELETSDSNHKGKDVNSIKSTSR



PLV7475

N607A
AYTMGIDVGIASCGWAIVDLERQRIIDIGVRTFEKAENPKNGEALAVPRREARSSRRRLRRK
23798





KHRIERLKHMFVRNGLAVDIQHLEQTLRSQNEIDVWQLRVDGLDRMLTQKEWLRVLIHLA






QRRGFQSNRKTDGSSEDGQVLVNVTENDRLMEEKDYRTVAEMMVKDEKFSDHKRNKNGN






YHGVVSRSSLLVEIHTLFETQRQHHNSLASKDFELEYVNIWSAQRPVATKDQIEKMIGTCTFL






PKEKRAPKASWHFQYFMLLQTINHIRITNVQGTRSLNKEEIEQVVNMALTKSKVSYHDTRKI






LDLSEEYQFVGLDYGKEDEKKKVESKETIIKLDDYHKLNKIFNEVELAKGETWEADDYDTV






AYALTFFKDDEDIRDYLQNKYKDSKNRLVKNLANKEYTNELIGKVSTLSFRKVGHLSLKAL






RKIIPFLEQGMTYDKACQAAGFDFQGISKKKRSVVLPVIDQISNPVVNRALTQTRKVINALIK






KYGSPETIHIETARELSKTFDERKNITKDYKENRDKNEHAKKHLSELGIINPTGLDIVKYKLW






CEQQGRCMYSNQPISFERLKESGYTEVDHIIPYSRSMNDSYNNRVLVMTREAREKGNQTPFE






YMGNDTQRWYEFEQRVTTNPQIKKEKRQNLLLKGFTNRRELEMLERNLNDTRYITKYLSHF






ISTNLEFSPSDKKKKVVNTSGRITSHLRSRWGLEKNRGQNDLHHAMDAIVIAVTSDSFIQQVT






NYYKRKERRELNGDDKFPLPWKFFREEVIARLSPNPKEQIEALPNHFYSEDELADLQPIFVSR






MPKRSITGEAHQAQFRRVVGKTKEGKNITAKKTALVDISYDKNGDFNMYGRETDPATYEAI






KERYLEFGGNVKKAFSTDLHKPKKDGTKGPLIKSVRIMENKTLVHPVNKGKGVVYNSSIVR






TDVFQRKEKYYLLPVYVTDVTKGKLPNKVIVAKKGYHDWIEVDDSFTFLFSLYPNDLIFIRQ






NPKKKISLKKRIESHSISDSKEVQEIHAYYKGVDSSTAAIEFIIHDGSYYAKGVGVQNLDCFEK






YQVDILGNYFKVKGEKRLELETSDSNHKGKDVNSIKSTSR






PLV4929
Nme2Cas9
WT
AAFKPNPINYILGLDIGIASVGWAMVEIDEEENPIRLIDLGVRVFERAEVPKTGDSLAMARRL
23799





ARSVRRLTRRRAHRLLRARRLLKREGVLQAADFDENGLIKSLPNTPWQLRAAALDRKLTPL






EWSAVLLHLIKHRGYLSQRKNEGETADKELGALLKGVANNAHALQTGDFRTPAELALNKF






EKESGHIRNQRGDYSHTFSRKDLQAELILLFEKQKEFGNPHVSGGLKEGIETLLMTQRPALSG






DAVQKMLGHCTFEPAEPKAAKNTYTAERFIWLTKLNNLRILEQGSERPLTDTERATLMDEP






YRKSKLTYAQARKLLGLEDTAFFKGLRYGKDNAEASTLMEMKAYHAISRALEKEGLKDKK






SPLNLSSELQDEIGTAFSLFKTDEDITGRLKDRVQPEILEALLKHISFDKFVQISLKALRRIVPL






MEQGKRYDEACAEIYGDHYGKKNTEEKIYLPPIPADEIRNPVVLRALSQARKVINGVVRRYG






SPARIHIETAREVGKSFKDRKEIEKRQEENRKDREKAAAKFREYFPNFVGEPKSKDILKLRLY






EQQHGKCLYSGKEINLVRLNEKGYVEIDHALPFSRTWDDSFNNKVLVLGSENQNKGNQTPY






EYFNGKDNSREWQEFKARVETSRFPRSKKQRILLQKFDEDGFKECNLNDTRYVNRFLCQFV






ADHILLTGKGKRRVFASNGQITNLLRGFWGLRKVRAENDRHHALDAVVVACSTVAMQQKI






TRFVRYKEMNAFDGKTIDKETGKVLHQKTHFPQPWEFFAQEVMIRVFGKPDGKPEFEEADT






PEKLRTLLAEKLSSRPEAVHEYVTPLFVSRAPNRKMSGAHKDTLRSAKRFVKHNEKISVKRV






WLTEIKLADLENMVNYKNGREIELYEALKARLEAYGGNAKQAFDPKDNPFYKKGGQLVKA






VRVEKTQESGVLLNKKNAYTIADNGDMVRVDVFCKVDKKGKNQYFIVPIYAWQVAENILP






DIDCKGYRIDDSYTFCFSLHKYDLIAFQKDEKSKVEFAYYINCDSSNGRFYLAWHDKGSKEQ






QFRISTQNLVLIQKYQVNELGKEIRPCRLKKRPPVR



PLV4947

N611A
AAFKPNPINYILGLDIGIASVGWAMVEIDEEENPIRLIDLGVRVFERAEVPKTGDSLAMARRL
23800





ARSVRRLTRRRAHRLLRARRLLKREGVLQAADFDENGLIKSLPNTPWQLRAAALDRKLTPL






EWSAVLLHLIKHRGYLSQRKNEGETADKELGALLKGVANNAHALQTGDFRTPAELALNKF






EKESGHIRNQRGDYSHTFSRKDLQAELILLFEKQKEFGNPHVSGGLKEGIETLLMTQRPALSG






DAVQKMLGHCTFEPAEPKAAKNTYTAERFIWLTKLNNLRILEQGSERPLTDTERATLMDEP






YRKSKLTYAQARKLLGLEDTAFFKGLRYGKDNAEASTLMEMKAYHAISRALEKEGLKDKK






SPLNLSSELQDEIGTAFSLFKTDEDITGRLKDRVQPEILEALLKHISFDKFVQISLKALRRIVPL






MEQGKRYDEACAEIYGDHYGKKNTEEKIYLPPIPADEIRNPVVLRALSQARKVINGVVRRYG






SPARIHIETAREVGKSFKDRKEIEKRQEENRKDREKAAAKFREYFPNFVGEPKSKDILKLRLY






EQQHGKCLYSGKEINLVRLNEKGYVEIDHALPFSRTWDDSFNNKVLVLGSEAQNKGNQTPY






EYFNGKDNSREWQEFKARVETSRFPRSKKQRILLQKFDEDGFKECNLNDTRYVNRFLCQFV






ADHILLTGKGKRRVFASNGQITNLLRGFWGLRKVRAENDRHHALDAVVVACSTVAMQQKI






TRFVRYKEMNAFDGKTIDKETGKVLHQKTHFPQPWEFFAQEVMIRVFGKPDGKPEFEEADT






PEKLRTLLAEKLSSRPEAVHEYVTPLFVSRAPNRKMSGAHKDTLRSAKRFVKHNEKISVKRV






WLTEIKLADLENMVNYKNGREIELYEALKARLEAYGGNAKQAFDPKDNPFYKKGGQLVKA






VRVEKTQESGVLLNKKNAYTIADNGDMVRVDVFCKVDKKGKNQYFIVPIYAWQVAENILP






DIDCKGYRIDDSYTFCFSLHKYDLIAFQKDEKSKVEFAYYINCDSSNGRFYLAWHDKGSKEQ






QFRISTQNLVLIQKYQVNELGKEIRPCRLKKRPPVR






PLV4928
PpnCas9
WT
QNNPLNYILGLDLGIASIGWAVVEIDEESSPIRLIDVGVRTFERAEVAKTGESLALSRRLARSS
23801





RRLIKRRAERLKKAKRLLKAEKILHSIDEKLPINVWQLRVKGLKEKLERQEWAAVLLHLSKH






RGYLSQRKNEGKSDNKELGALLSGIASNHQMLQSSEYRTPAEIAVKKFQVEEGHIRNQRGSY






THTFSRLDLLAEMELLFQRQAELGNSYTSTTLLENLTALLMWQKPALAGDAILKMLGKCTF






EPSEYKAAKNSYSAERFVWLTKLNNLRILENGTERALNDNERFALLEQPYEKSKLTYAQVR






AMLALSDNAIFKGVRYLGEDKKTVESKTTLIEMKFYHQIRKTLGSAELKKEWNELKGNSDL






LDEIGTAFSLYKTDDDICRYLEGKLPERVLNALLENLNFDKFIQLSLKALHQILPLMLQGQRY






DEAVSAIYGDHYGKKSTETTRLLPTIPADEIRNPVVLRTLTQARKVINAVVRLYGSPARIHIET






AREVGKSYQDRKKLEKQQEDNRKQRESAVKKFKEMFPHFVGEPKGKDILKMRLYELQQAK






CLYSGKSLELHRLLEKGYVEVDHALPFSRTWDDSFNNKVLVLANENQNKGNLTPYEWLDG






KNNSERWQHFVVRVQTSGFSYAKKQRILNHKLDEKGFIERNLNDTRYVARFLCNFIADNML






LVGKGKRNVFASNGQITALLRHRWGLQKVREQNDRHHALDAVVVACSTVAMQQKITRFV






RYNEGNVFSGERIDRETGEIIPLHFPSPWAFFKENVEIRIFSENPKLELENRLPDYPQYNHEWV






QPLFVSRMPTRKMTGQGHMETVKSAKRLNEGLSVLKVPLTQLKLSDLERMVNRDREIALYE






SLKARLEQFGNDPAKAFAEPFYKKGGALVKAVRLEQTQKSGVLVRDGNGVADNASMVRV






DVFTKGGKYFLVPIYTWQVAKGILPNRAATQGKDENDWDIMDEMATFQFSLCQNDLIKLVT






KKKTIFGYFNGLNRATSNINIKEHDLDKSKGKLGIYLEVGVKLAISLEKYQVDELGKNIRPCR






PTKRQHVR



PLV4946

N605A
QNNPLNYILGLDLGIASIGWAVVEIDEESSPIRLIDVGVRTFERAEVAKTGESLALSRRLARSS
23802





RRLIKRRAERLKKAKRLLKAEKILHSIDEKLPINVWQLRVKGLKEKLERQEWAAVLLHLSKH






RGYLSQRKNEGKSDNKELGALLSGIASNHQMLQSSEYRTPAEIAVKKFQVEEGHIRNQRGSY






THTFSRLDLLAEMELLFQRQAELGNSYTSTTLLENLTALLMWQKPALAGDAILKMLGKCTF






EPSEYKAAKNSYSAERFVWLTKLNNLRILENGTERALNDNERFALLEQPYEKSKLTYAQVR






AMLALSDNAIFKGVRYLGEDKKTVESKTTLIEMKFYHQIRKTLGSAELKKEWNELKGNSDL






LDEIGTAFSLYKTDDDICRYLEGKLPERVLNALLENLNFDKFIQLSLKALHQILPLMLQGQRY






DEAVSAIYGDHYGKKSTETTRLLPTIPADEIRNPVVLRTLTQARKVINAVVRLYGSPARIHIET






AREVGKSYQDRKKLEKQQEDNRKQRESAVKKFKEMFPHFVGEPKGKDILKMRLYELQQAK






CLYSGKSLELHRLLEKGYVEVDHALPFSRTWDDSFNNKVLVLANEAQNKGNLTPYEWLDG






KNNSERWQHFVVRVQTSGFSYAKKQRILNHKLDEKGFIERNLNDTRYVARFLCNFIADNML






LVGKGKRNVFASNGQITALLRHRWGLQKVREQNDRHHALDAVVVACSTVAMQQKITRFV






RYNEGNVFSGERIDRETGEIIPLHFPSPWAFFKENVEIRIFSENPKLELENRLPDYPQYNHEWV






QPLFVSRMPTRKMTGQGHMETVKSAKRLNEGLSVLKVPLTQLKLSDLERMVNRDREIALYE






SLKARLEQFGNDPAKAFAEPFYKKGGALVKAVRLEQTQKSGVLVRDGNGVADNASMVRV






DVFTKGGKYFLVPIYTWQVAKGILPNRAATQGKDENDWDIMDEMATFQFSLCQNDLIKLVT






KKKTIFGYFNGLNRATSNINIKEHDLDKSKGKLGIYLEVGVKLAISLEKYQVDELGKNIRPCR






PTKRQHVR






PLV4924
SauCas9
WT
KRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGARRLKRRRRHRI
23803





QRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSAALLHLAKRRGVHNVNEVEE






DTGNELSTKEQISRNSKALEEKYVAELQLERLKKDGEVRGSINRFKTSDYVKEAKQLLKVQ






KAYHQLDQSFIDTYIDLLETRRTYYEGPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVK






YAYNADLYNALNDLNNLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIK






GYRVTSTGKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSELT






QEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKVDLSQQKEIPTTL






VDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNSKDAQKMINEMQKRNRQTNERI






EEIIRTTGKENAKYLIEKIKLHDMQEGKCLYSLEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFN






NKVLVKQEENSKKGNRTPFQYLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDIN






RFSVQKDFINRNLVDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKER






NKGYKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQEYKEIFIT






PHQIKHIKDFKDYKYSHRVDKKPNRELINDTLYSTRKDDKGNTLIVNNLNGLYDKDNDKLK






KLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNPLYKYYEETGNYLTKYSKKDNGPVIK






KIKYYGNKLNAHLDITDDYPNSRNKVVKLSLKPYRFDVYLDNGVYKFVTVKNLDVIKKEN






YYEVNSKCYEEAKKLKKISNQAEFIASFYNNDLIKINGELYRVIGVNNDLLNRIEVNMIDITY






REYLENMNDKRPPRIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG



PLV4949

N580A
KRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGARRLKRRRRHRI
23804





QRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSAALLHLAKRRGVHNVNEVEE






DTGNELSTKEQISRNSKALEEKYVAELQLERLKKDGEVRGSINRFKTSDYVKEAKQLLKVQ






KAYHQLDQSFIDTYIDLLETRRTYYEGPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVK






YAYNADLYNALNDLNNLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIK






GYRVTSTGKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSELT






QEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKVDLSQQKEIPTTL






VDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNSKDAQKMINEMQKRNRQTNERI






EEIIRTTGKENAKYLIEKIKLHDMQEGKCLYSLEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFN






NKVLVKQEEASKKGNRTPFQYLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDIN






RFSVQKDFINRNLVDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKER






NKGYKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQEYKEIFIT






PHQIKHIKDFKDYKYSHRVDKKPNRELINDTLYSTRKDDKGNTLIVNNLNGLYDKDNDKLK






KLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNPLYKYYEETGNYLTKYSKKDNGPVIK






KIKYYGNKLNAHLDITDDYPNSRNKVVKLSLKPYRFDVYLDNGVYKFVTVKNLDVIKKEN






YYEVNSKCYEEAKKLKKISNQAEFIASFYNNDLIKINGELYRVIGVNNDLLNRIEVNMIDITY






REYLENMNDKRPPRIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG






PLV4932
SauCas9-
WT
KRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGARRLKRRRRHRI
23805



KKH

QRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSAALLHLAKRRGVHNVNEVEE






DTGNELSTKEQISRNSKALEEKYVAELQLERLKKDGEVRGSINRFKTSDYVKEAKQLLKVQ






KAYHQLDQSFIDTYIDLLETRRTYYEGPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVK






YAYNADLYNALNDLNNLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIK






GYRVTSTGKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSELT






QEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKVDLSQQKEIPTTL






VDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNSKDAQKMINEMQKRNRQTNERI






EEIIRTTGKENAKYLIEKIKLHDMQEGKCLYSLEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFN






NKVLVKQEENSKKGNRTPFQYLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDIN






RFSVQKDFINRNLVDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKER






NKGYKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQEYKEIFIT






PHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIVNNLNGLYDKDNDKLK






KLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNPLYKYYEETGNYLTKYSKKDNGPVIK






KIKYYGNKLNAHLDITDDYPNSRNKVVKLSLKPYRFDVYLDNGVYKFVTVKNLDVIKKEN






YYEVNSKCYEEAKKLKKISNQAEFIASFYKNDLIKINGELYRVIGVNNDLLNRIEVNMIDITY






REYLENMNDKRPPHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG



PLV4945

N580A
KRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEGRRSKRGARRLKRRRRHRI
23806





QRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEFSAALLHLAKRRGVHNVNEVEE






DTGNELSTKEQISRNSKALEEKYVAELQLERLKKDGEVRGSINRFKTSDYVKEAKQLLKVQ






KAYHQLDQSFIDTYIDLLETRRTYYEGPGEGSPFGWKDIKEWYEMLMGHCTYFPEELRSVK






YAYNADLYNALNDLNNLVITRDENEKLEYYEKFQIIENVFKQKKKPTLKQIAKEILVNEEDIK






GYRVTSTGKPEFTNLKVYHDIKDITARKEIIENAELLDQIAKILTIYQSSEDIQEELTNLNSELT






QEEIEQISNLKGYTGTHNLSLKAINLILDELWHTNDNQIAIFNRLKLVPKKVDLSQQKEIPTTL






VDDFILSPVVKRSFIQSIKVINAIIKKYGLPNDIIIELAREKNSKDAQKMINEMQKRNRQTNERI






EEIIRTTGKENAKYLIEKIKLHDMQEGKCLYSLEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFN






NKVLVKQEEASKKGNRTPFQYLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDIN






RFSVQKDFINRNLVDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKER






NKGYKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQEYKEIFIT






PHQIKHIKDFKDYKYSHRVDKKPNRKLINDTLYSTRKDDKGNTLIVNNLNGLYDKDNDKLK






KLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNPLYKYYEETGNYLTKYSKKDNGPVIK






KIKYYGNKLNAHLDITDDYPNSRNKVVKLSLKPYRFDVYLDNGVYKFVTVKNLDVIKKEN






YYEVNSKCYEEAKKLKKISNQAEFIASFYKNDLIKINGELYRVIGVNNDLLNRIEVNMIDITY






REYLENMNDKRPPHIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQIIKKG






PLV4930
Sauri-
WT
QENQQKQNYILGLDIGITSVGYGLIDSKTREVIDAGVRLFPEADSENNSNRRSKRGARRLKRR
23807



Cas9

RIHRLNRVKDLLADYQMIDLNNVPKSTDPYTIRVKGLREPLTKEEFAIALLHIAKRRGLHNIS






VSMGDEEQDNELSTKQQLQKNAQQLQDKYVCELQLERLTNINKVRGEKNRFKTEDFVKEV






KQLCETQRQYHNIDDQFIQQYIDLVSTRREYFEGPGNGSPYGWDGDLLKWYEKLMGRCTYF






PEELRSVKYAYSADLFNALNDLNNLVVTRDDNPKLEYYEKYHIIENVFKQKKNPTLKQIAKE






IGVQDYDIRGYRITKSGKPQFTSFKLYHDLKNIFEQAKYLEDVEMLDEIAKILTIYQDEISIKK






ALDQLPELLTESEKSQIAQLTGYTGTHRLSLKCIHIVIDELWESPENQMEIFTRLNLKPKKVE






MSEIDSIPTTLVDEFILSPVVKRAFIQSIKVINAVINRFGLPEDIIIELAREKNSKDRRKFINK






LQKQNEATRKKIEQLLAKYGNTNAKYMIEKIKLHDMQEGKCLYSLEAIPLEDLLSNPTHYEVDH






IIPRSVSFDNSLNNKVLVKQSENSKKGNRTPYQYLSSNESKISYNQFKQHILNLSKAKDRISKK






KRDMLLEERDINKFEVQKEFINRNLVDTRYATRELSNLLKTYFSTHDYAVKVKTINGGFTNH






LRKVWDFKKHRNHGYKHHAEDALVIANADFLFKTHKALRRTDKILEQPGLEVNDTTVKVD






TEEKYQELFETPKQVKNIKQFRDFKYSHRVDKKPNRQLINDTLYSTREIDGETYVVQTLKDL






YAKDNEKVKKLFTERPQKILMYQHDPKTFEKLMTILNQYAEAKNPLAAYYEDKGEYVTKY






AKKGNGPAIHKIKYIDKKLGSYLDVSNKYPETQNKLVKLSLKSFRFDIYKCEQGYKMVSIGY






LDVLKKDNYYYIPKDKYEAEKQKKKIKESDLFVGSFYYNDLIMYEDELFRVIGVNSDINNLV






ELNMVDITYKDFCEVNNVTGEKRIKKTIGKRVVLIEKYTTDILGNLYKTPLPKKPQLIFKRGE






L



PLV4950

N588A
QENQQKQNYILGLDIGITSVGYGLIDSKTREVIDAGVRLFPEADSENNSNRRSKRGARRLKRR
23808





RIHRLNRVKDLLADYQMIDLNNVPKSTDPYTIRVKGLREPLTKEEFAIALLHIAKRRGLHNIS






VSMGDEEQDNELSTKQQLQKNAQQLQDKYVCELQLERLTNINKVRGEKNRFKTEDFVKEV






KQLCETQRQYHNIDDQFIQQYIDLVSTRREYFEGPGNGSPYGWDGDLLKWYEKLMGRCTYF






PEELRSVKYAYSADLFNALNDLNNLVVTRDDNPKLEYYEKYHIIENVFKQKKNPTLKQIAKE






IGVQDYDIRGYRITKSGKPQFTSFKLYHDLKNIFEQAKYLEDVEMLDEIAKILTIYQDEISIKK






ALDQLPELLTESEKSQIAQLTGYTGTHRLSLKCIHIVIDELWESPENQMEIFTRLNLKPKKVE






MSEIDSIPTTLVDEFILSPVVKRAFIQSIKVINAVINRFGLPEDIIIELAREKNSKDRRKFINK






LQKQNEATRKKIEQLLAKYGNTNAKYMIEKIKLHDMQEGKCLYSLEAIPLEDLLSNPTHYEVDH






IIPRSVSFDNSLNNKVLVKQSEASKKGNRTPYQYLSSNESKISYNQFKQHILNLSKAKDRISKK






KRDMLLEERDINKFEVQKEFINRNLVDTRYATRELSNLLKTYFSTHDYAVKVKTINGGFTNH






LRKVWDFKKHRNHGYKHHAEDALVIANADFLFKTHKALRRTDKILEQPGLEVNDTTVKVD






TEEKYQELFETPKQVKNIKQFRDFKYSHRVDKKPNRQLINDTLYSTREIDGETYVVQTLKDL






YAKDNEKVKKLFTERPQKILMYQHDPKTFEKLMTILNQYAEAKNPLAAYYEDKGEYVTKY






AKKGNGPAIHKIKYIDKKLGSYLDVSNKYPETQNKLVKLSLKSFRFDIYKCEQGYKMVSIGY






LDVLKKDNYYYIPKDKYEAEKQKKKIKESDLFVGSFYYNDLIMYEDELFRVIGVNSDINNLV






ELNMVDITYKDFCEVNNVTGEKRIKKTIGKRVVLIEKYTTDILGNLYKTPLPKKPQLIFKRGE






L






PLV4926
Sauri
WT
QENQQKQNYILGLDIGITSVGYGLIDSKTREVIDAGVRLFPEADSENNSNRRSKRGARRLKRR
23809



Cas9-

RIHRLNRVKDLLADYQMIDLNNVPKSTDPYTIRVKGLREPLTKEEFAIALLHIAKRRGLHNIS




KKH

VSMGDEEQDNELSTKQQLQKNAQQLQDKYVCELQLERLTNINKVRGEKNRFKTEDFVKEV






KQLCETQRQYHNIDDQFIQQYIDLVSTRREYFEGPGNGSPYGWDGDLLKWYEKLMGRCTYF






PEELRSVKYAYSADLFNALNDLNNLVVTRDDNPKLEYYEKYHIIENVFKQKKNPTLKQIAKE






IGVQDYDIRGYRITKSGKPQFTSFKLYHDLKNIFEQAKYLEDVEMLDEIAKILTIYQDEISIKK






ALDQLPELLTESEKSQIAQLTGYTGTHRLSLKCIHIVIDELWESPENQMEIFTRLNLKPKKVE






MSEIDSIPTTLVDEFILSPVVKRAFIQSIKVINAVINRFGLPEDIIIELAREKNSKDRRKFINK






LQKQNEATRKKIEQLLAKYGNTNAKYMIEKIKLHDMQEGKCLYSLEAIPLEDLLSNPTHYEVDH






IIPRSVSFDNSLNNKVLVKQSENSKKGNRTPYQYLSSNESKISYNQFKQHILNLSKAKDRISKK






KRDMLLEERDINKFEVQKEFINRNLVDTRYATRELSNLLKTYFSTHDYAVKVKTINGGFTNH






LRKVWDFKKHRNHGYKHHAEDALVIANADFLFKTHKALRRTDKILEQPGLEVNDTTVKVD






TEEKYQELFETPKQVKNIKQFRDFKYSHRVDKKPNRKLINDTLYSTREIDGETYVVQTLKDL






YAKDNEKVKKLFTERPQKILMYQHDPKTFEKLMTILNQYAEAKNPLAAYYEDKGEYVTKY






AKKGNGPAIHKIKYIDKKLGSYLDVSNKYPETQNKLVKLSLKSFRFDIYKCEQGYKMVSIGY






LDVLKKDNYYYIPKDKYEAEKQKKKIKESDLFVGSFYKNDLIMYEDELFRVIGVNSDINNLV






ELNMVDITYKDFCEVNNVTGEKHIKKTIGKRVVLIEKYTTDILGNLYKTPLPKKPQLIFKRGE






L



PLV4952

N588A
QENQQKQNYILGLDIGITSVGYGLIDSKTREVIDAGVRLFPEADSENNSNRRSKRGARRLKRR
23810





RIHRLNRVKDLLADYQMIDLNNVPKSTDPYTIRVKGLREPLTKEEFAIALLHIAKRRGLHNIS






VSMGDEEQDNELSTKQQLQKNAQQLQDKYVCELQLERLTNINKVRGEKNRFKTEDFVKEV






KQLCETQRQYHNIDDQFIQQYIDLVSTRREYFEGPGNGSPYGWDGDLLKWYEKLMGRCTYF






PEELRSVKYAYSADLFNALNDLNNLVVTRDDNPKLEYYEKYHIIENVFKQKKNPTLKQIAKE






IGVQDYDIRGYRITKSGKPQFTSFKLYHDLKNIFEQAKYLEDVEMLDEIAKILTIYQDEISIKK






ALDQLPELLTESEKSQIAQLTGYTGTHRLSLKCIHIVIDELWESPENQMEIFTRLNLKPKKVE






MSEIDSIPTTLVDEFILSPVVKRAFIQSIKVINAVINRFGLPEDIIIELAREKNSKDRRKFINK






LQKQNEATRKKIEQLLAKYGNTNAKYMIEKIKLHDMQEGKCLYSLEAIPLEDLLSNPTHYEVDH






IIPRSVSFDNSLNNKVLVKQSEASKKGNRTPYQYLSSNESKISYNQFKQHILNLSKAKDRISKK






KRDMLLEERDINKFEVQKEFINRNLVDTRYATRELSNLLKTYFSTHDYAVKVKTINGGFTNH






LRKVWDFKKHRNHGYKHHAEDALVIANADFLFKTHKALRRTDKILEQPGLEVNDTTVKVD






TEEKYQELFETPKQVKNIKQFRDFKYSHRVDKKPNRKLINDTLYSTREIDGETYVVQTLKDL






YAKDNEKVKKLFTERPQKILMYQHDPKTFEKLMTILNQYAEAKNPLAAYYEDKGEYVTKY






AKKGNGPAIHKIKYIDKKLGSYLDVSNKYPETQNKLVKLSLKSFRFDIYKCEQGYKMVSIGY






LDVLKKDNYYYIPKDKYEAEKQKKKIKESDLFVGSFYKNDLIMYEDELFRVIGVNSDINNLV






ELNMVDITYKDFCEVNNVTGEKHIKKTIGKRVVLIEKYTTDILGNLYKTPLPKKPQLIFKRGE






L






PLV4925
Sca-
WT
EKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTNRKSIKKNLMGALLFDSGETAEATR
23811



Cas9++

LKRTARRRYTRRKNRIRYLQEIFANEMAKLDDSFFQRLEESFLVEEDKKNERHPIFGNLADE






VAYHRNYPTIYHLRKKLADSPEKADLRLIYLALAHIIKFRGHFLIEGKLNAENSDVAKLFYQL






IQTYNQLFEESPLDEIEVDAKGILSARLSKSKRLEKLIAVFPNEKKNGLFGNIIALALGLTPNFK






SNFDLTEDAKLQLSKDTYDDDLDELLGQIGDQYADLFSAAKNLSDAILLSDILRSNSEVTKAP






LSASMVKRYDEHHQDLALLKTLVRQQFPEKYAEIFKDDTKNGYAGYVGADKKLRKRSGKL






ATEEEFYKFIKPILEKMDGAEELLAKLNRDDLLRKQRTFDNGSIPHQIHLKELHAILRRQEEF






YPFLKENREKIEKILTFRIPYYVGPLARGNSRFAWLTRKSEEAITPWNFEEVVDKGASAQSFIE






RMTNFDEQLPNKKVLPKHSLLYEYFTVYNELTKVKYVTERMRKPEFLSGEQKKAIVDLLFK






TNRKVTVKQLKEDYFKKIECFDSVEIIGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILE






DIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQLKRRHYTGWGRLSRKMINGIRDKQSGKTI






LDFLKSDGFSNRNFMQLIHDDSLTFKEEIEKAQVSGQGDSLHEQIADLAGSPAIKKGILQTVKI






VDELVKVMGHKPENIVIEMARENQTTTKGLQQSRERKKRIEEGIKELESQILKENPVENTQL






QNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFIKDDSIDNKVLTRSVENRGKS






DNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSEADKAGFIKRQLVETRQIT






KHVARILDSRMNTKRDKNDKPIREVKVITLKSKLVSDFRKDFQLYKVRDINNYHHAHDAYL






NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKRFFYSNIMNFFKTEV






KLANGEIRKRPLIETNGETGEVVWNKEKDFATVRKVLAMPQVNIVKKTEVQTGGFSKESILS






KRESAKLIPRKKGWDTRKYGGFGSPTVAYSILVVAKVEKGKAKKLKSVKVLVGITIMEKGS






YEKDPIGFLEAKGYKDIKKELIFKLPKYSLFELENGRRRMLASAKELQKANELVLPQHLVRL






LYYTQNISATTGSNNLGYIEQHREEFKEIFEKIIDFSEKYILKNKVNSNLKSSFDEQFAVSDSI






LLSNSFVSLLKYTSFGASGGFTFLDLDVKQGRLRYQTVTEVLDATLIYQSITGLYETRTDLSQL






GGD



PLV4951

N872A
EKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTNRKSIKKNLMGALLFDSGETAEATR
23812





LKRTARRRYTRRKNRIRYLQEIFANEMAKLDDSFFQRLEESFLVEEDKKNERHPIFGNLADE






VAYHRNYPTIYHLRKKLADSPEKADLRLIYLALAHIIKFRGHFLIEGKLNAENSDVAKLFYQL






IQTYNQLFEESPLDEIEVDAKGILSARLSKSKRLEKLIAVFPNEKKNGLFGNIIALALGLTPNFK






SNFDLTEDAKLQLSKDTYDDDLDELLGQIGDQYADLFSAAKNLSDAILLSDILRSNSEVTKAP






LSASMVKRYDEHHQDLALLKTLVRQQFPEKYAEIFKDDTKNGYAGYVGADKKLRKRSGKL






ATEEEFYKFIKPILEKMDGAEELLAKLNRDDLLRKQRTFDNGSIPHQIHLKELHAILRRQEEF






YPFLKENREKIEKILTFRIPYYVGPLARGNSRFAWLTRKSEEAITPWNFEEVVDKGASAQSFIE






RMTNFDEQLPNKKVLPKHSLLYEYFTVYNELTKVKYVTERMRKPEFLSGEQKKAIVDLLFK






TNRKVTVKQLKEDYFKKIECFDSVEIIGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILE






DIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQLKRRHYTGWGRLSRKMINGIRDKQSGKTI






LDFLKSDGFSNRNFMQLIHDDSLTFKEEIEKAQVSGQGDSLHEQIADLAGSPAIKKGILQTVKI






VDELVKVMGHKPENIVIEMARENQTTTKGLQQSRERKKRIEEGIKELESQILKENPVENTQL






QNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFIKDDSIDNKVLTRSVEARGKS






DNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSEADKAGFIKRQLVETRQIT






KHVARILDSRMNTKRDKNDKPIREVKVITLKSKLVSDFRKDFQLYKVRDINNYHHAHDAYL






NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKRFFYSNIMNFFKTEV






KLANGEIRKRPLIETNGETGEVVWNKEKDFATVRKVLAMPQVNIVKKTEVQTGGFSKESILS






KRESAKLIPRKKGWDTRKYGGFGSPTVAYSILVVAKVEKGKAKKLKSVKVLVGITIMEKGS






YEKDPIGFLEAKGYKDIKKELIFKLPKYSLFELENGRRRMLASAKELQKANELVLPQHLVRL






LYYTQNISATTGSNNLGYIEQHREEFKEIFEKIIDFSEKYILKNKVNSNLKSSFDEQFAVSDSI






LLSNSFVSLLKYTSFGASGGFTFLDLDVKQGRLRYQTVTEVLDATLIYQSITGLYETRTDLSQL






GGD






PLV4931
SpyCas9-
WT
DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAERTRL
23813



SpRY

KRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVA






YHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLV






QTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNF






KSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITK






APLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFI






KPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREK






IEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNL






PNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQ






LKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFE






DREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGF






ANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKV






MGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY






LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSE






EVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQI






LDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGT






ALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIR






KRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESIRPKRNSDKLIA






RKKDWDPKKYGGFLWPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFL






EAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAKQLQKGNELALPSKYVNFLYLASHYE






KLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQA






ENIIHLFTLTRLGAPRAFKYFDTTIDPKQYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD



PLV4948

N863A
DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAERTRL
23814





KRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVA






YHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLV






QTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNF






KSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITK






APLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFI






KPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREK






IEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNL






PNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQ






LKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFE






DREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGF






ANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKV






MGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY






LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPSE






EVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQI






LDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGT






ALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIR






KRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESIRPKRNSDKLIA






RKKDWDPKKYGGFLWPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFL






EAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAKQLQKGNELALPSKYVNFLYLASHYE






KLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQA






ENIIHLFTLTRLGAPRAFKYFDTTIDPKQYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD






PLV4927
SpyCas9-
WT
DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATR
23815



NG

LKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEV






AYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQL






VQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPN






FKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEIT






KAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKF






IKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNRE






KIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKN






LPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVK






QLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLF






EDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDG






FANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVK






VMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKL






YLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS






EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQ






ILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVG






TALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI






RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESIRPKRNSDKLI






ARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFL






EAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASARFLQKGNELALPSKYVNFLYLASHYEK






LKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAE






NIIHLFTLTNLGAPRAFKYFDTTIDRKVYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD



PLV4943

N863A
DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATR
23816





LKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEV






AYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQL






VQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPN






FKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEIT






KAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKF






IKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNRE






KIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKN






LPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVK






QLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLF






EDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDG






FANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVK






VMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKL






YLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS






EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQ






ILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVG






TALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEI






RKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESIRPKRNSDKLI






ARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFL






EAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASARFLQKGNELALPSKYVNFLYLASHYEK






LKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAE






NIIHLFTLTNLGAPRAFKYFDTTIDRKVYRSTKEVLDATLIHQSITGLYETRIDLSQLGGD






PLV4933
St1Cas9
WT
SDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQGRRLARRKKHRRV
23817





RLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFIALKNMVKHRGISYLDDASDDG






NSSVGDYAQIVKENSKQLETKTPGQIQLERYQTYGQLRGDFTVEKDGKKHRLINVFPTSAYR






SEALRILQTQQEFNPQITDEFINRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGI






LIGKCTFYPDEFRAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGP






AKLFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETLDKLAYV






LTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGWHNFSVKLMMELIPELY






ETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIYNPVVAKSVRQAIKIVNAAIKEYGDF






DNIVIEMARETNEDDEKKAIQKIQKANKDEKDAAMLKAANQYNGKAELPHSVFHGHKQLA






TKIRLWHQQGERCLYTGKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKG






QRTPYQALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLVDTR






YASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYHHHAVDALIIAASS






QLNLWKKQKNTLVSYSEDQLLDIETGELISDDEYKESVFKAPYQHFVDTLKSKEFEDSILFSY






QVDSKFNRKISDATIYATRQAKVGKDKADETYVLGKIKDIYTQDGYDAFMKIYKKDKSKFL






MYRHDPQTFEKVIEPILENYPNKQINEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLK






YYDSKLGNHIDITPKDSNNKVVLQSVSPWRADVYFNKTTGKYEILGLKYADLQFEKGTGTY






KISQEKYNDIKKKEGVDSDSEFKFTLYKNDLLLVKDTETKEQQLFRFLSRTMPKQKHYVELK






PYDKQKFEGGEALIKVLGNVANSGQCKKGLGKSNISIYKVRTDVLGNQHIIKNEGDKPKLDF



PLV4944

N622A
SDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVRRTNRQGRRLARRKKHRRV
23818





RLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEELFIALKNMVKHRGISYLDDASDDG






NSSVGDYAQIVKENSKQLETKTPGQIQLERYQTYGQLRGDFTVEKDGKKHRLINVFPTSAYR






SEALRILQTQQEFNPQITDEFINRYLEILTGKRKYYHGPGNEKSRTDYGRYRTSGETLDNIFGI






LIGKCTFYPDEFRAAKASYTAQEFNLLNDLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGP






AKLFKYIAKLLSCDVADIKGYRIDKSGKAEIHTFEAYRKMKTLETLDIEQMDRETLDKLAYV






LTLNTEREGIQEALEHEFADGSFSQKQVDELVQFRKANSSIFGKGWHNFSVKLMMELIPELY






ETSEEQMTILTRLGKQKTTSSSNKTKYIDEKLLTEEIYNPVVAKSVRQAIKIVNAAIKEYGDF






DNIVIEMARETNEDDEKKAIQKIQKANKDEKDAAMLKAANQYNGKAELPHSVFHGHKQLA






TKIRLWHQQGERCLYTGKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATAAQEKG






QRTPYQALDSMDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLVDTR






YASRVVLNALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYHHHAVDALIIAASS






QLNLWKKQKNTLVSYSEDQLLDIETGELISDDEYKESVFKAPYQHFVDTLKSKEFEDSILFSY






QVDSKFNRKISDATIYATRQAKVGKDKADETYVLGKIKDIYTQDGYDAFMKIYKKDKSKFL






MYRHDPQTFEKVIEPILENYPNKQINEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPEIKSLK






YYDSKLGNHIDITPKDSNNKVVLQSVSPWRADVYFNKTTGKYEILGLKYADLQFEKGTGTY






KISQEKYNDIKKKEGVDSDSEFKFTLYKNDLLLVKDTETKEQQLFRFLSRTMPKQKHYVELK






PYDKQKFEGGEALIKVLGNVANSGQCKKGLGKSNISIYKVRTDVLGNQHIIKNEGDKPKLDF









First the indel activity of gene modifying systems comprising template RNAs comprising 5 SpCas9 spacers, in combination with wildtype SpCas9 polypeptide was evaluated in HEK293T cells.


As shown in FIG. 13, out of the 5 spacers tested, spacers 1, 3, and 4 showed indel activity of >15%. Template RNAs were designed using these 3 spacers and tested for rewrite activity using exemplary gene modifying polypeptides comprising SpCas9. However, these combinations did not result in rewrite activity above a target threshold (data not shown). Without wishing to be bound by theory, SpCas9 spacers did not yield rewrite activity above the target threshold likely due to the distance from the PiZ mutation. Based on this result, Cas9 variants and compatible spacers were evaluated.


Twelve different Cas9 variants (Table X1) with several different spacers for each variant were screened. Exemplary gene modifying polypeptides comprising the different Cas9 domains were generated (Table X1) and tested with compatible template RNAs (Table X2). FIG. 14 shows the indel % at the PiZ mutation site in HEK293T landing pad cells after treatment with the gene modifying systems. The results showed that many combinations of Cas9 variant domains and compatible template RNAs showed promising indel activity. Of the combinations examined, five Cas9 variants were identified that showed similar or higher indel activity relative to SpCas9 (FIG. 14). The active spacers were ranked by indel activity and distance from the PiZ mutation, and selected spacer:Cas9 combinations that showed high indel activity and were located within 20 bp of the PiZ mutation were selected for further screening (FIG. 15 and Table XX).









TABLE X2







sgRNAs used for spacer screen















SEQ

SEQ


Plasmid

Spacer
ID

ID


Number
Name
sequence
NO
Scaffold Sequence
NO





PLV5610
pU6-
CTGTG
23819
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23938



Spy-
CTGAC

AAAGTGGCACCGAGTCGGTGC




A1AT-
CATCG






sgRNA-
ACAAG






1









PLV5611
pU6-
GCTGT
23820
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23939



Spy-
GCTGA

AAAGTGGCACCGAGTCGGTGC




A1AT-
CCATC






sgRNA-
GACAA






1G
G








PLV5612
pU6-
CAGCT
23821
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23940



Spy-
TCAGT

AAAGTGGCACCGAGTCGGTGC




A1AT-
CCCTTT






sgRNA-
CTTG






2









PLV5613
pU6-
GCAGC
23822
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23941



Spy-
TTCAG

AAAGTGGCACCGAGTCGGTGC




A1AT-
TCCCTT






sgRNA-
TCTTG






2G









PLV5614
pU6-
GGCTG
23823
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23942



Spy-
TGCTG

AAAGTGGCACCGAGTCGGTGC




A1AT-
ACCAT






sgRNA-
CGACA






3









PLV5615
pU6-
AGGCT
23824
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23943



Spy-
GTGCT

AAAGTGGCACCGAGTCGGTGC




A1AT-
GACCA






sgRNA-
TCGAC






4









PLV5616
pU6-
GAGGC
23825
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23944



Spy-
TGTGC

AAAGTGGCACCGAGTCGGTGC




A1AT-
TGACC






sgRNA-
ATCGA






4G
C








PLV5617
pU6-
AGCAG
23826
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23945



Spy-
CTTCA

AAAGTGGCACCGAGTCGGTGC




A1AT-
GTCCC






sgRNA-
TTTCT






5









PLV5618
pU6-
GAGCA
23827
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23946



Spy-
GCTTC

AAAGTGGCACCGAGTCGGTGC




A1AT-
AGTCC






sgRNA-
CTTTCT






5G









PLV5619
pU6-
GGCCG
23828
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23947



Spy-
TGCAT

AAAGTGGCACCGAGTCGGTGC




A1AT-
AAGGC






sgRNA-
TGTGC






6









PLV5620
pU6-
CCAGG
23829
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23948



Spy-
CCGTG

AAAGTGGCACCGAGTCGGTGC




A1AT-
CATAA






sgRNA-
GGCTG






7









PLV5621
pU6-
GCCAG
23830
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23949



Spy-
GCCGT

AAAGTGGCACCGAGTCGGTGC




A1AT-
GCATA






sgRNA-
AGGCT






7G
G








PLV5622
pU6-
CAGCA
23831
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23950



Spy-
GCTTC

AAAGTGGCACCGAGTCGGTGC




A1AT-
AGTCC






sgRNA-
CTTTC






8









PLV5623
pU6-
GCAGC
23832
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23951



Spy-
AGCTT

AAAGTGGCACCGAGTCGGTGC




A1AT-
CAGTC






sgRNA-
CCTTTC






8G









PLV5624
pU6-
AGGCC
23833
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23952



Spy-
GTGCA

AAAGTGGCACCGAGTCGGTGC




A1AT-
TAAGG






sgRNA-
CTGTG






9









PLV5625
pU6-
GAGGC
23834
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23953



Spy-
CGTGC

AAAGTGGCACCGAGTCGGTGC




A1AT-
ATAAG






sgRNA-
GCTGT






9G
G








PLV5626
pU6-
TCCAG
23835
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23954



Spy-
GCCGT

AAAGTGGCACCGAGTCGGTGC




A1AT-
GCATA






sgRNA-
AGGCT






10









PLV5627
pU6-
GTCCA
23836
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23955



Spy-
GGCCG

AAAGTGGCACCGAGTCGGTGC




A1AT-
TGCAT






sgRNA-
AAGGC






10G
T








PLV5628
pU6-
ACCTC
23837
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23956



Spy-
GGGGG

AAAGTGGCACCGAGTCGGTGC




A1AT-
GGATA






sgRNA-
GACAT






11









PLV5629
pU6-
GACCT
23838
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23957



Spy-
CGGGG

AAAGTGGCACCGAGTCGGTGC




A1AT-
GGGAT






sgRNA-
AGACA






11G
T








PLV5630
pU6-
TGTTG
23839
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23958



Spy-
AACTT

AAAGTGGCACCGAGTCGGTGC




A1AT-
GACCT






sgRNA-
CGGGG






12









PLV5631
pU6-
GTGTT
23840
GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAA
23959



Spy-
GAACT

AAAGTGGCACCGAGTCGGTGC




A1AT-
TGACC






sgRNA-
TCGGG






12G
G








PLV5564
pU6-
AAGGC
23841
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23960



Sau-
TGTGC

CTCGTCAACTTGTTGGCGAGA




A1AT-
TGACC






sgRNA-
ATCGA






1
C








PLV5565
pU6-
GAAGG
23842
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23961



Sau-
CTGTG

CTCGTCAACTTGTTGGCGAGA




A1AT-
CTGAC






sgRNA-
CATCG






1G
AC








PLV5566
pU6-
AGCAG
23843
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23962



Sau-
CTTCA

CTCGTCAACTTGTTGGCGAGA




A1AT-
GTCCC






sgRNA-
TTTCTT






2









PLV5567
pU6-
GAGCA
23844
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23963



Sau-
GCTTC

CTCGTCAACTTGTTGGCGAGA




A1AT-
AGTCC






sgRNA-
CTTTCT






2G
T








PLV5568
pU6-
CCAGG
23845
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23964



Sau-
CCGTG

CTCGTCAACTTGTTGGCGAGA




A1AT-
CATAA






sgRNA-
GGCTG






3
T








PLV5569
pU6-
GCCAG
23846
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23965



Sau-
GCCGT

CTCGTCAACTTGTTGGCGAGA




A1AT-
GCATA






sgRNA-
AGGCT






3G
GT








PLV5570
pU6-
TAAAA
23847
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23966



Sau-
ACATG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GCCCC






sgRNA-
AGCAG






4
C








PLV5571
pU6-
GTAAA
23848
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23967



Sau-
AACAT

CTCGTCAACTTGTTGGCGAGA




A1AT-
GGCCC






sgRNA-
CAGCA






4G
GC








PLV5572
pU6-
GGCCT
23849
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23968



Sau-
CTAAA

CTCGTCAACTTGTTGGCGAGA




A1AT-
AACAT






sgRNA-
GGCCC






5
C








PLV5573
pU6-
TATGG
23850
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23969



Sau-
CCTCT

CTCGTCAACTTGTTGGCGAGA




A1AT-
AAAAA






sgRNA-
CATGG






6
C








PLV5574
pU6-
GTATG
23851
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23970



Sau-
GCCTC

CTCGTCAACTTGTTGGCGAGA




A1AT-
TAAAA






sgRNA-
ACATG






G
GC








PLV5575
pU6-
TTGAC
23852
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23971



Sau-
CTCGG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GGGGG






sgRNA-
ATAGA






7
C








PLV5576
pU6-
GTTGA
23853
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23972



Sau-
CCTCG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GGGGG






sgRNA-
GATAG






7G
AC








PLV5577
pU6-
TTTGTT
23854
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23973



Sau-
GAACT

CTCGTCAACTTGTTGGCGAGA




A1AT-
TGACC






sgRNA-
TCGGG






8









PLV5578
pU6-
GTTTG
23855
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23974



Sau-
TTGAA

CTCGTCAACTTGTTGGCGAGA




A1AT-
CTTGA






sgRNA-
CCTCG






8G
GG








PLV5579
pU6-
ACGTG
23856
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23975



Sau-
AGCCT

CTCGTCAACTTGTTGGCGAGA




A1AT-
TGCTC






sgRNA-
GAGGC






9
C








PLV5580
pU6-
GACGT
23857
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23976



Sau-
GAGCC

CTCGTCAACTTGTTGGCGAGA




A1AT-
TTGCT






sgRNA-
CGAGG






9G
CC








PLV5581
pU6-
ATTAA
23858
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23977



Sau-
GAAGA

CTCGTCAACTTGTTGGCGAGA




A1AT-
CAAAG






sgRNA-
GGTTT






10
G








PLV5582
pU6-
GATTA
23859
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23978



Sau-
AGAAG

CTCGTCAACTTGTTGGCGAGA




A1AT-
ACAAA






sgRNA-
GGGTT






10G
TG








PLV5583
pU6-
AGGTG
23860
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23979



Sau-
TCCAC

CTCGTCAACTTGTTGGCGAGA




A1AT-
GTGAG






sgRNA-
CCTTG






11
C








PLV5584
pU6-
GAGGT
23861
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23980



Sau-
GTCCA

CTCGTCAACTTGTTGGCGAGA




A1AT-
CGTGA






sgRNA-
GCCTT






11G
GC








PLV5585
pU6-
TGTTC
23862
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23981



Sau-
AATCA

CTCGTCAACTTGTTGGCGAGA




A1AT-
TTAAG






sgRNA-
AAGAC






12
A








PLV5586
pU6-
GTGTT
23863
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23982



Sau-
CAATC

CTCGTCAACTTGTTGGCGAGA




A1AT-
ATTAA






sgRNA-
GAAGA






12G
CA








PLV5587
pU6-
CGCTT
23864
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23983



Sau-
CCTGG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GAGGT






sgRNA-
GTCCA






13
C








PLV5588
pU6-
GCGCT
23865
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23984



Sau-
TCCTG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GGAGG






sgRNA-
TGTCC






13G
AC








PLV5589
pU6-
TCTCC
23866
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23985



Sauri-
CCTCC

CTCGTCAACTTGTTGGCGAGA




A1AT-
AGGCC






sgRNA-
GTGCA






1
T








PLV5590
pU6-
GTCTC
23867
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23986



Sauri-
CCCTC

CTCGTCAACTTGTTGGCGAGA




A1AT-
CAGGC






sgRNA-
CGTGC






1G
AT








PLV5591
pU6-
ATGGG
23868
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23987



Sauri-
TATGG

CTCGTCAACTTGTTGGCGAGA




A1AT-
CCTCT






sgRNA-
AAAAA






2
C








PLV5592
pU6-
GATGG
23869
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23988



Sauri-
GTATG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GCCTC






sgRNA-
TAAAA






2G
AC








PLV5593
pU6-
TAAGG
23870
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23989



Sauri-
CTGTG

CTCGTCAACTTGTTGGCGAGA




A1AT-
CTGAC






sgRNA-
CATCG






3
A








PLV5594
pU6-
GTAAG
23871
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23990



Sauri-
GCTGT

CTCGTCAACTTGTTGGCGAGA




A1AT-
GCTGA






sgRNA-
CCATC






3G
GA








PLV5595
pU6-
AAAAA
23872
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23991



Sauri-
CATGG

CTCGTCAACTTGTTGGCGAGA




A1AT-
CCCCA






sgRNA-
GCAGC






4
T








PLV5596
pU6-
GAAAA
23873
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23992



Sauri-
ACATG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GCCCC






sgRNA-
AGCAG






4G
CT








PLV5597
pU6-
GCCTC
23874
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23993



Sauri-
TAAAA

CTCGTCAACTTGTTGGCGAGA




A1AT-
ACATG






sgRNA-
GCCCC






5
A








PLV5598
pU6-
ATGGC
23875
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23994



Sauri-
CTCTA

CTCGTCAACTTGTTGGCGAGA




A1AT-
AAAAC






sgRNA-
ATGGC






6
C








PLV5599
pU6-
GATGG
23876
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23995



Sauri-
CCTCT

CTCGTCAACTTGTTGGCGAGA




A1AT-
AAAAA






sgRNA-
CATGG






6G
CC








PLV5600
pU6-
CTCTC
23877
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23996



Sauri-
CCCTC

CTCGTCAACTTGTTGGCGAGA




A1AT-
CAGGC






sgRNA-
CGTGC






7
A








PLV5601
pU6-
GCTCT
23878
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23997



Sauri-
CCCCT

CTCGTCAACTTGTTGGCGAGA




A1AT-
CCAGG






sgRNA-
CCGTG






7G
CA








PLV5602
pU6-
TGTCT
23879
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23998



Sauri-
CTGCT

CTCGTCAACTTGTTGGCGAGA




A1AT-
TCTCTC






sgRNA-
CCCTC






8









PLV5603
pU6-
GTGTC
23880
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
23999



Sauri-
TCTGC

CTCGTCAACTTGTTGGCGAGA




A1AT-
TTCTCT






sgRNA-
CCCCT






8G
C








PLV5604
pU6-
TGACC
23881
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
24000



Sauri-
TCGGG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GGGGA






sgRNA-
TAGAC






9
A








PLV5605
pU6-
GTGAC
23882
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
24001



Sauri-
CTCGG

CTCGTCAACTTGTTGGCGAGA




A1AT-
GGGGG






sgRNA-
ATAGA






9G
CA








PLV5606
pU6-
AAGGG
23883
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
24002



Sauri-
TTTGTT

CTCGTCAACTTGTTGGCGAGA




A1AT-
GAACT






sgRNA-
TGACC






10









PLV5607
pU6-
GAAGG
23884
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
24003



Sauri-
GTTTG

CTCGTCAACTTGTTGGCGAGA




A1AT-
TTGAA






sgRNA-
CTTGA






10G
CC








PLV5608
pU6-
GTGTC
23885
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
24004



Sauri-
CACGT

CTCGTCAACTTGTTGGCGAGA




A1AT-
GAGCC






sgRNA-
TTGCT






11
C








PLV5609
pU6-
GTTCA
23886
GTTTTAGTACTCTGGAAACAGAATCTACTAAAACAAGGCAAAATGCCGTGTTTAT
24005



Sauri-
ATCAT

CTCGTCAACTTGTTGGCGAGA




A1AT-
TAAGA






sgRNA-
AGACA






12
A








PLV5537
pU6-
GTAAA
23887
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24006



Nme2-
AACAT

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
GGCCC

GGCATCGTTTA




sgRNA-
CAGCA






1
GCTT








PLV5538
pU6-
GTCCA
23888
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24007



Nme2-
GGCCG

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
TGCAT

GGCATCGTTTA




sgRNA-
AAGGC






2
TGTG








PLV5539
pU6-
GCATG
23889
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24008



Nme2-
GGTAT

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
GGCCT

GGCATCGTTTA




sgRNA-
CTAAA






3
AACA








PLV5540
pU6-
GACAT
23890
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24009



Nme2-
GGGTA

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
TGGCC

GGCATCGTTTA




sgRNA-
TCTAA






4
AAAC








PLV5541
pU6-
GCGTG
23891
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24010



Nme2-
TCTCT

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
GCTTC

GGCATCGTTTA




sgRNA-
TCTCC






5
CCTC








PLV5542
pU6-
GCCTT
23892
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24011



Nme2-
ACAAC

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
GTGTC

GGCATCGTTTA




sgRNA-
TCTGC






6
TTCT








PLV5543
pU6-
GCCTC
23893
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24012



Nme2-
GGGGG

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
GGATA

GGCATCGTTTA




sgRNA-
GACAT






7
GGGT








PLV5544
pU6-
GTGAG
23894
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24013



Nme2-
CCTTG

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
CTCGA

GGCATCGTTTA




sgRNA-
GGCCT






8
GGGA








PLV5545
pU6-
GAGAA
23895
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24014



Nme2-
GACAA

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
AGGGT

GGCATCGTTTA




sgRNA-
TTGTT






9
GAAC








PLV5546
pU6-
GAGGT
23896
GTTGTAGCTCCCTTTCTCATTTCGGAAACGAAATGAGAACCGTTGCTACAATAAG
24015



Nme2-
GTCCA

GCCGTCTGAAAAGATGTGCCGCAACGCTCTGCCCCTTAAAGCTTCTGCTTTAAGG




A1AT-
CGTGA

GGCATCGTTTA




sgRNA-
GCCTT






10
GCTC








PLV5519
pU6-
TGCAT
23897
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24016



Blat-
AAGGC

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
TGTGC

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
TGACC






1
A








PLV5520
pU6-
GTGCA
23898
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24017



Blat-
TAAGG

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
CTGTG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
CTGAC






1G
CA








PLV5521
pU6-
TGGCC
23899
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24018



Blat-
CCAGC

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
AGCTT

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
CAGTC






2
C








PLV5522
pU6-
GTGGC
23900
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24019



Blat-
CCCAG

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




AIAT-
CAGCT

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
TCAGT






2G
CC








PLV5523
pU6-
CCGTG
23901
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24020



Blat-
CATAA

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
GGCTG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
TGCTG






3
A








PLV5524
pU6-
GCCGT
23902
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24021



Blat-
GCATA

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
AGGCT

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GTGCT






3G
GA








PLV5525
pU6-
AGGCC
23903
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24022



Blat-
GTGCA

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
TAAGG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
CTGTG






4
C








PLV5526
pU6-
GAGGC
23904
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24023



Blat-
CGTGC

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
ATAAG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GCTGT






4G
GC








PLV5527
pU6-
AAAAC
23905
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24024



Blat-
ATGGC

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
CCCAG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
CAGCT






5
T








PLV5528
pU6-
GAAAA
23906
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24025



Blat-
CATGG

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
CCCCA

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GCAGC






5G
TT








PLV5529
pU6-
GGGGG
23907
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24026



Blat-
GATAG

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
ACATG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GGTAT






6
G








PLV5530
pU6-
TGAAC
23908
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24027



Blat-
TTGAC

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
CTCGG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GGGGG






7
A








PLV5531
pU6-
GTGAA
23909
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24028



Blat-
CTTGA

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
CCTCG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GGGGG






7G
GA








PLV5532
pU6-
TCGAG
23910
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24029



Blat-
GCCTG

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
GGATC

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
AGCCT






8
T








PLV5533
pU6-
GTCGA
23911
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24030



Blat-
GGCCT

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
GGGAT

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
CAGCC






8G
TT








PLV5534
pU6-
ACAAA
23912
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24031



Blat-
GGGTT

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
TGTTG

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
AACTT






9
G








PLV5535
pU6-
GACAA
23913
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24032



Blat-
AGGGT

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
TTGTT

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
GAACT






9G
TG








PLV5536
pU6-
GCCTT
23914
GCTATAGTTCCTTACTGAAAGGTAAGTTGCTATAGTAAGGGCAACAGACCCGAGG
24033



Blat-
GCTCG

CGTTGGGGATCGCCTAGCCCGTGTTTACGGGCTCTCCCCATATTCAAAATAATGA




A1AT-
AGGCC

CAGACGAGCACCTTGGAGCATTTATCTCCGAGGTGCT




sgRNA-
TGGGA






10
T








PLV5547
pU6-
AAAGG
23915
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24034



Ppn-
GACTG

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
AAGCT

C




sgRNA-
GCTGG






1
GG








PLV5548
pU6-
GAAAG
23916
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24035



Ppn-
GGACT

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
GAAGC

C




sgRNA-
TGCTG






1G
GGG








PLV5549
pU6-
CCTGG
23917
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24036



Ppn-
AGGGG

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
AGAGA

C




sgRNA-
AGCAG






2
AG








PLV5550
pU6-
GCCTG
23918
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24037



Ppn-
GAGGG

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
GAGAG

C




sgRNA-
AAGCA






2G
GAG








PLV5551
pU6-
CCCAT
23919
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24038



Ppn-
GTCTA

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
TCCCC

C




sgRNA-
CCCGA






3
GG








PLV5552
pU6-
GCCCA
23920
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24039



Ppn-
TGTCT

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
ATCCC

C




sgRNA-
CCCCG






3G
AGG








PLV5553
pU6-
TCAAT
23921
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24040



Ppn-
CATTA

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
AGAAG

C




sgRNA-
ACAAA






4
GG








PLV5554
pU6-
GTCAA
23922
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24041



Ppn-
TCATT

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
AAGAA

C




sgRNA-
GACAA






4G
AGG








PLV5555
pU6-
TTGTTC
23923
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24042



Ppn-
AATCA

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
TTAAG

C




sgRNA-
AAGAC






5
A








PLV5556
pU6-
GTTGT
23924
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24043



Ppn-
TCAAT

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
CATTA

C




sgRNA-
AGAAG






5G
ACA








PLV5557
pU6-
TCAAC
23925
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24044



Ppn-
AAACC

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
CTTTGT

C




sgRNA-
CTTCTT






6









PLV5558
pU6-
GTCAA
23926
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24045



Ppn-
CAAAC

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
CCTTT

C




sgRNA-
GTCTT






6G
CTT








PLV5559
pU6-
GGGGA
23927
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24046



Ppn-
GACTT

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
GGTAT

C




sgRNA-
TTTGTT






7
C








PLV5560
pU6-
CATGA
23928
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24047



Ppn-
AGAGG

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
GGAGA

C




sgRNA-
CTTGG






8
TA








PLV5561
pU6-
GCATG
23929
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24048



Ppn-
AAGAG

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
GGGAG

C




sgRNA-
ACTTG






8G
GTA








PLV5562
pU6-
TTTCCC
23930
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24049



Ppn-
ATGAA

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
GAGGG

C




sgRNA-
GAGAC






9
T








PLV5563
pU6-
GTTTC
23931
GTTGTAGCTCCCTTTTTCATTTCGCGAAAGCGAAATGAAAAACGTTGTTACAATA
24050



Ppn-
CCATG

AGAGATGAATTTCTCGCAAAGCTCTGCCTCTTGAAATTTCGGTTTCAAGAGGCAT




A1AT-
AAGAG

C




sgRNA-
GGGAG






9G
ACT








PLV5632
pU6-
AAGGC
23932
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATAAGGCTTCATGCCGAAATCAA
24051



St1-
TGTGC

CACCCTGTCATTTTATGGCAGGGTGTTTT




A1AT-
TGACC






sgRNA-
ATCGA






1









PLV5633
pU6-
GAAGG
23933
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATAAGGCTTCATGCCGAAATCAA
24052



St1-
CTGTG

CACCCTGTCATTTTATGGCAGGGTGTTTT




A1AT-
CTGAC






sgRNA-
CATCG






1G
A








PLV5634
pU6-
AAGGC
23934
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATAAGGCTTCATGCCGAAATCAA
24053



St1-
TCACG

CACCCTGTCATTTTATGGCAGGGTGTTTT




A1AT-
TGGAC






sgRNA-
ACCTC






2









PLV5635
pU6-
GAAGG
23935
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATAAGGCTTCATGCCGAAATCAA
24054



St1-
CTCAC

CACCCTGTCATTTTATGGCAGGGTGTTTT




A1AT-
GTGGA






sgRNA-
CACCT






2G
C








PLV5636
pU6-
TACCA
23936
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATAAGGCTTCATGCCGAAATCAA
24055



St1-
AGTCT

CACCCTGTCATTTTATGGCAGGGTGTTTT




A1AT-
CCCCT






sgRNA-
CTTCA






3









PLV5637
pU6-
GTACC
23937
GTCTTTGTACTCTGGTACCAGAAGCTACAAAGATAAGGCTTCATGCCGAAATCAA
24056



St1-
AAGTC

CACCCTGTCATTTTATGGCAGGGTGTTTT




A1AT-
TCCCC






sgRNA-
TCTTC






3G
A
















TABLE XX







Selected Cas Spacer Combination











Cas

SEQ ID



Variant
Spacer
NO







SpRY
CTGTGCTGACCATCGACAAG
24057







SpRY
GCTGTGCTGACCATCGACAAG
24058







SpRY
GGCTGTGCTGACCATCGACA
24059







SpRY
AGGCTGTGCTGACCATCGAC
24060







ScaCas9
AGGCTGTGCTGACCATCGAC
24061







SpCas9
CCAGGCCGTGCATAAGGCTG
24062







SpRY
AGGCCGTGCATAAGGCTGTG
24063







SpCas9
AGGCCGTGCATAAGGCTGTG
24064







Nme2
GTAAAAACATGGCCCCAGCAGCTT
24065







Nme2
GTCCAGGCCGTGCATAAGGCTGTG
24066







St1
AAGGCTGTGCTGACCATCGA
24067







St1
GAAGGCTGTGCTGACCATCGA
24068










Example 10. Screening of Cas9 Variant and Spacer Combinations for High Rewrite Activity

This example describes the use of an exemplary gene modifying system containing a gene modifying polypeptide and exemplary template RNAs comprising varied lengths of heterologous object sequences and PBS sequences to quantify the activity of template RNAs for correction of the PiZ mutation. In this example, a template RNA contains:

    • (1) a gRNA spacer;
    • (2) a gRNA scaffold;
    • (3) a heterologous object sequence; and
    • (4) a primer binding site (PBS) sequence.


The exemplary gene modifying polypeptides used are described in Example 9 above. The 11 Cas9/spacer combinations that were identified in Example 9 to have high spacer activity (indels) were then used to screen for template RNAs having rewriting activity in combination with exemplary gene modifying polypeptides comprising matched Cas9 domains. The heterologous object sequences and PBS sequences were designed to correct the AATD PiZ mutation in a landing pad by replacing a “T” nucleotide with a “C” nucleotide at the mutation site via gene editing, to reverse a PiZ mutation in the corresponding protein. Sequences of the exemplary template RNAs tested are shown in Table X3.


A gene modifying system comprising a (i) compatible gene modifying polypeptide described comprising: an NLS as described in Example 9, a compatible nickase Cas9 having a sequence of Table X1 (e.g., SpyCas9-SpRY), a linker as described in Example 9, an RT sequence as described in Example 9 (e.g., PLV4931), and a second NLS as described in Example 9 and (ii) a template RNA from Table X3 was transfected into the HEK293T landing pad cell line. The gene modifying polypeptide and the sgRNAs were delivered by transfection in RNA format. Specifically, 75 ng of gene modifying polypeptide mRNA was combined with 1 pmol of template RNA. This combination of RNAs was then mixed with 0.5 uL of Lipofectamine MessengerMax, in 10 uL of OptiMEM solution, and added to 20,000 cells. After transfection, cells were grown at 37° C., 5% CO2 for 3 days prior to cell lysis and genomic DNA extraction. To analyze gene editing activity, primers flanking the A1 AT PiZ mutation site were used to amplify across the locus. Amplicons were analyzed via short read sequencing using an Illumina MiSeq.


The gene modifying systems comprising combinations of gene modifying polypeptides and template RNAs, wherein the gene modifying polypeptides comprised Cas9 domains matched to the template RNA spacers from Example 9, were tested for rewriting activity. The various template RNAs tested comprised different PBS and RT template lengths (FIG. 16). The results showed that, of the combinations examined, 5 combinations of Cas9 variants and spacers, demonstrated rewriting activity above a target threshold of 2% (FIG. 16 and Table X5).









TABLE X5







Cas9 Variant and Spacer Pairs








Cas9 variant
Spacer











pU6_A1AT_SpRY_ED0-_ (having a
CTGTGCTGACCATCGACAAG


Cas9 variant amino acid
(SEQ ID NO: 17087)


sequence according to SEQ ID 



NO: 23813)






pU6_A1AT_SpRY_ED2- (having a
GGCTGTGCTGACCATCGACA


Cas9 variant amino acid
(SEQ ID NO: 17108)


sequence according to SEQ ID



NO: 23813)






pU6_A1AT_SpRY_ED3- (having a
AGGCTGTGCTGACCATCGAC


Cas9 variant amino acid
(SEQ ID NO: 17120)


sequence according to SEQ ID 



NO: 23813)






pU6_A1AT_SpRY_ED15- (having
AGGCCGTGCATAAGGCTGTG


a Cas9 variant amino acid
(SEQ ID NO: 17220)


sequence according to SEQ ID 



NO: 23813)






pU6_A1AT_St1_ED4- (having a
GAAGGCTGTGCTGACCATCG


Cas9 variant amino acid
A


sequence according to SEQ ID 
(SEQ ID NO: 20065)


NO: 23817)










FIG. 17A-17B show heat maps graphing the % rewriting of gene modifying systems comprising various SpRY EDO template RNAs (varying PBS and RT lengths) and an exemplary SpRY Cas9-containing gene modifying polypeptide (FIG. 17A) and gene modifying systems comprising various St1_ED4 template RNAs (varying PBS and RT lengths) and an exemplary St1Cas9-containing gene modifying polypeptide (FIG. 17B). The results identify several optimal RT lengths (e.g., 6, 8, 10, 12, and 15 nucleotides) and PBS lengths (8 10, 11, and 12 nucleotides) and combinations thereof for SpRY EDO template RNAs. The results identified several optimal RT lengths (e.g., 8, 14, and 16 nucleotides) and PBS lengths (8, 9, 10, 11, and 12 nucleotides) and combinations thereof for St1_ED4 template RNAs. For SpRY Cas9 and SpRY EDO spacer, the best performing template RNA showed 10.3% perfect rewriting and comprised an RT length of 6 nucleotides and a PBS length of 8 nucleotides. For St1Cas9 and St1_ED4 spacer, the best performing RT length was 8 nucleotides and PBS length 9 nucleotides. The rewriting activities of the top-performing 17 combinations of template RNAs and gene modifying polypeptides comprising Cas9 variants (as ranked by rewriting activity) were graphed in FIG. 18. The results further demonstrate that the tested gene modifying systems show rewriting activity at the target PiZ mutation site, and that rewriting levels are generally higher than indel levels. Exemplary gene modifying systems comprising a SpRYCas9 gene modifying polypeptide with SpRY_ED0_PBS8_RT6 template RNA performed best with 10.3% rewriting and less than 4% indels.









TABLE X3







Exemplary Template RNAs for Correcting PiZ Mutation











tgRNA

SEQ


ID
Name
tgRNA_seq_IDT_formatted
ID NO













0
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24069



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*




PB17
mA*mG






1
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24070



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*




PB16
mC*mA






2
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24071



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*




PB15
mC






3
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24072



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*




PB14
mA






4
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24073



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







5
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24074



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







6
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24075



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







7
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24076



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







8
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24077



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







9
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24078



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







10
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24079



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







11
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24080



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







12
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24081



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







13
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24082



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







14
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24083



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







15
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24084



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







16
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24085



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







17
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24086



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







18
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24087



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







19
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24088



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







20
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24089



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







21
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24090



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







22
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24091



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







23
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24092



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







24
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24093



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







25
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24094



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







26
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24095



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







27
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24096



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







28
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24097



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







29
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24098



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







30
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24099



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







31
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24100



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







32
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24101



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







33
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24102



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







34
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24103



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







35
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24104



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







36
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24105



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







37
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24106



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







38
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24107



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







39
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24108



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







40
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24109



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







41
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24110



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







42
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24111



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







43
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24112



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







44
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24113



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







45
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24114



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







46
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24115



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







47
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24116



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







48
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24117



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







49
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24118



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







50
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24119



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







51
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24120



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







52
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24121



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







53
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24122



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







54
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24123



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







55
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24124



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







56
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24125



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







57
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24126



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







58
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24127



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







59
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24128



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







60
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24129



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







61
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24130



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







62
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24131



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







63
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24132



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







64
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24133



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







65
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24134



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







66
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24135



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







67
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24136



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







68
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24137



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







69
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24138



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







70
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24139



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







71
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24140



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







72
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24141



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







73
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24142



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







74
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24143



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







75
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24144



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







76
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24145



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







77
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24146



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







78
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24147



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







79
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24148



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







80
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24149



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB17







81
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24150



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB16







82
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24151



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB15







83
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24152



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB14







84
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24153



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB13







85
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24154



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB12







86
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24155



ED0-_
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




PB11







87
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24156



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC




PB10







88
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24157



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArU*mG*mG*mU




PB9







89
A1AT_SpRY_
mC*mU*mG*rUrGrCrUrGrArCrCrArUrCrGrArCrArArGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24158



ED0-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrA*mU*mG*mG




PB8







90
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24159



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*




PB17
mG*mC*mC






91
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24160



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*




PB16
mA*mG*mC






92
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24161



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*




PB15
mA*mG






93
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24162



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*




PB14
mC*mA






94
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24163



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*




PB13
mC






95
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24164



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*




PB12
mA






96
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24165



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB11







97
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24166



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB10







98
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24167



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_PB9
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






99
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24168



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




30FE_PB8
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






100
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24169



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*




PB17
mC






101
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24170



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB16







102
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24171



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB15







103
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24172



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB14







104
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24173



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB13







105
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24174



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB12







106
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24175



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB11







107
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24176



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB10







108
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24177



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_PB9
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






109
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24178



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




25FE_PB8
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






110
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24179



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB17







111
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24180



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB16







112
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24181



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB15







113
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24182



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB14







114
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24183



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB13







115
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24184



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB12







116
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24185



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB11







117
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24186



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB10







118
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24187



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_PB9
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






119
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24188



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




20FE_PB8
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






120
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24189



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB17







121
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24190



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB16







122
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24191



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB15







123
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24192



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB14







124
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24193



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB13







125
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24194



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB12







126
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24195



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB11







127
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24196



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB10







128
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24197



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_PB9
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






129
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24198



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




14FE_PB8
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






130
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24199



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB17







131
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24200



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB16







132
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24201



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB15







133
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24202



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB14







134
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24203



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB13







135
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24204



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB12







136
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24205



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB11







137
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24206



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB10







138
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24207



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_PB9
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






139
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24208



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




11FE_PB8
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






140
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24209



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB17
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC






141
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24210



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB16
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC






142
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24211



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB15
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG






143
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24212



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB14
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA






144
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24213



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB13
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC






145
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24214



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB12
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA






146
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24215



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB11
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC






147
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24216



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB10
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG






148
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24217



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB9
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






149
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24218



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




9FE_PB8
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






150
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24219



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB17
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC






151
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24220



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB16
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC






152
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24221



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB15
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG






153
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24222



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB14
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA






154
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24223



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB13
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC






155
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24224



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB12
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA






156
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24225



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB11
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC






157
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24226



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB10
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG






158
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24227



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB9
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






159
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24228



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




7FE_PB8
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






160
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24229



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB17
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC






161
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24230



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB16
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC






162
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24231



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB15
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG






163
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24232



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB14
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA






164
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24233



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB13
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC






165
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24234



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB12
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA






166
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24235



ED2-_
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB11
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC






167
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24236



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB10
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG






168
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24237



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB9
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






169
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24238



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




5FE_PB8
GrCrUrUrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






170
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24239



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB17
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC






171
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24240



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB16
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC






172
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24241



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB15
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG






173
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24242



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB14
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA






174
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24243



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB13
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC






175
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24244



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB12
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA






176
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24245



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB11
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC






177
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24246



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB10
GrCrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG






178
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24247



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB9
GrCrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA






179
A1AT_SpRY_
mG*mG*mC*rUrGrUrGrCrUrGrArCrCrArUrCrGrArCrArGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24248



ED2-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




3FE_PB8
GrCrUrCrUrCrGrUrCrGrArUrG*mG*mU*mC






180
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24249



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
G*mC*mC*mU




PB17







181
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24250



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*




30FE_
mG*mC*mC




PB16







182
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24251



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*




30FE_
mA*mG*mC




PB15







183
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24252



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*




30FE_
mA*mG




PB14







184
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24253



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*




30FE_
mC*mA




PB13







185
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24254



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*




30FE_
mC




PB12







186
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24255



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*




30FE_
mA




PB11







187
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24256



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




30FE_





PB10







188
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24257



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




30FE_PB9







189
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24258



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




30FE_PB8







190
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24259



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*




25FE_
mU




PB17







191
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24260



ScaCas9++_
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*m




25FE_
C




PB16







192
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24261



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




25FE_





PB15







193
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24262



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




25FE_





PB14







194
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24263



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




25FE_





PB13







195
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24264



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




25FE_





PB12







196
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24265



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




25FE_





PB11







197
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24266



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




25FE_





PB10







198
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24267



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




25FE_PB9







199
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24268



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




25FE_PB8







200
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24269



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




20FE_





PB17







201
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24270



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




20FE_





PB16







202
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24271



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




20FE_





PB15







203
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24272



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




20FE_





PB14







204
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24273



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




20FE_





PB13







205
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24274



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




20FE_





PB12







206
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24275



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




20FE_





PB11







207
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24276



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




20FE_





PB10







208
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24277



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




20FE_PB9







209
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24278



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




20FE_PB8







210
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24279



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




14FE_





PB17







211
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24280



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




14FE_





PB16







212
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24281



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




14FE_





PB15







213
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24282



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




14FE_





PB14







214
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24283



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




14FE_





PB13







215
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24284



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




14FE_





PB12







216
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24285



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




14FE_





PB11







217
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24286



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




14FE_





PB10







218
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24287



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




14FE_PB9







219
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24288



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




14FE_PB8







220
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24289



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




11FE_





PB17







221
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24290



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




11FE_





PB16







222
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24291



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




11FE_





PB15







223
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24292



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




11FE_





PB14







224
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24293



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




11FE_





PB13







225
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24294



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




11FE_





PB12







226
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24295



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




11FE_





PB11







227
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24296



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




11FE_





PB10







228
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24297



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




11FE_PB9







229
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24298



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




11FE_PB8







230
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24299



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




9FE_PB17







231
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24300



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




9FE_PB16







232
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24301



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




9FE_PB15







233
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24302



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




9FE_PB14







234
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24303



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




9FE_PB13







235
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24304



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




9FE_PB12







236
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24305



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




9FE_PB11







237
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24306



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




9FE_PB10







238
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24307



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




9FE_PB9







239
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24308



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




9FE_PB8







240
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24309



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




7FE_PB17







241
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24310



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




7FE_PB16







242
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24311



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




7FE_PB15







243
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24312



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




7FE_PB14







244
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24313



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




7FE_PB13







245
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24314



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




7FE_PB12







246
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24315



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




7FE_PB11







247
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24316



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




7FE_PB10







248
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24317



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




7FE_PB9







249
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24318



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




7FE_PB8







250
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24319



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




5FE_PB17







251
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24320



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




5FE_PB16







252
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24321



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




5FE_PB15







253
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24322



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




5FE_PB14







254
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24323



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




5FE_PB13







255
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24324



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




5FE_PB12







256
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24325



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




5FE_PB11







257
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24326



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




5FE_PB10







258
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24327



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




5FE_PB9







259
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24328



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




5FE_PB8







260
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24329



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




3FE_PB17







261
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24330



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




3FE_PB16







262
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24331



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




3FE_PB15







263
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24332



ScaCas9++_
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




3FE_PB14







264
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24333



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




3FE_PB13







265
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24334



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




3FE_PB12







266
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24335



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




3FE_PB11







267
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24336



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




3FE_PB10







268
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24337



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




3FE_PB9







269
A1AT_
mA*mG*mG*rCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24338



ScaCas9++_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




SpRY_ED3-_
GrCrUrCrUrCrGrUrCrGrArUrGrG*mU*mC*mA




3FE_PB8







270
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24339



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB17
UrCrArGrCrArCrArGrC*mC*mU*mU






271
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24340



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB16
UrCrArGrCrArCrArG*mC*mC*mU






272
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24341



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB15
UrCrArGrCrArCrA*mG*mC*mC






273
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24342



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB14
UrCrArGrCrArC*mA*mG*mC






274
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24343



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB13
UrCrArGrCrA*mC*mA*mG






275
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24344



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB12
UrCrArGrC*mA*mC*mA






276
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24345



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB11
UrCrArG*mC*mA*mC






277
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24346



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB10
UrCrA*mG*mC*mA






278
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24347



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB9
UrC*mA*mG*mC






279
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24348



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_30FE_
ArGrGrGrUrGrUrUrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGr




PB8
U*mC*mA*mG






280
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24349



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB17
ArCrArGrC*mC*mU*mU






281
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24350



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB16
ArCrArG*mC*mC*mU






282
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24351



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB15
ArCrA*mG*mC*mC






283
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24352



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB14
ArC*mA*mG*mC






284
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24353



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB13
A*mC*mA*mG






285
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24354



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC




PB12
*mA*mC*mA






286
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24355



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*m




PB11
C*mA*mC






287
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24356



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG




PB10
*mC*mA






288
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24357



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*




PB9
mG*mC






289
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24358



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_25FE_
ArGrGrGrUrGrUrUrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*




PB8
mG






290
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24359



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB17
*mC*mU*mU






291
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24360



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*




PB16
mC*mC*mU






292
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24361



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG




PB15
*mC*mC






293
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24362



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*




PB14
mG*mC






294
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24363



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA




PB13
*mG






295
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24364



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*




PB12
mA






296
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24365



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







297
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24366



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







298
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24367



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







299
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24368



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_20FE_
ArGrGrGrUrGrUrUrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







300
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24369



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC*mC*mU*mU




PB17







301
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24370



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




PB16







302
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24371



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB15







303
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24372



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB14







304
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24373



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB13







305
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24374



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB12







306
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24375



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







307
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24376



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







308
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24377



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







309
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24378



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_14FE_
ArGrGrGrUrGrUrUrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







310
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24379



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC*mC*mU*mU




PB17







311
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24380



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




PB16







312
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24381



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB15







313
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24382



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB14







314
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24383



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB13







315
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24384



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB12







316
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24385



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







317
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24386



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







318
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24387



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







319
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24388



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_11FE_
ArGrGrGrUrGrUrUrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







320
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24389



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC*mC*mU*mU




PB17







321
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24390



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




PB16







322
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24391



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB15







323
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24392



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_PB
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




14







324
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24393



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB13







325
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24394



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB12







326
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24395



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







327
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24396



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







328
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24397



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







329
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24398



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_9FE_
ArGrGrGrUrGrUrUrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







330
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24399



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC*mC*mU*mU




1PB7







331
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24400



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




PB16







332
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24401



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB15







333
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24402



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB14







334
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24403



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB13







335
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24404



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB12







336
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24405



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







337
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24406



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







338
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24407



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







339
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24408



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_7FE_
ArGrGrGrUrGrUrUrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







340
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24409



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC*mC*mU*mU




PB17







341
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24410



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




PB16







342
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24411



ED4-_
ArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB15







343
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24412



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB14







344
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24413



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB13







345
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24414



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB12







346
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24415



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







347
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24416



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







348
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24417



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







349
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24418



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_5FE_
ArGrGrGrUrGrUrUrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







350
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24419



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC*mC*mU*mU




PB17







351
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24420



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArG*mC*mC*mU




PB16







352
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24421



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA*mG*mC*mC




PB15







353
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24422



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArC*mA*mG*mC




PB14







354
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24423



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrA*mC*mA*mG




PB13







355
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24424



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrC*mA*mC*mA




PB12







356
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24425



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArG*mC*mA*mC




PB11







357
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24426



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrA*mG*mC*mA




PB10







358
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24427



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrC*mA*mG*mC




PB9







359
A1AT_St1_
mA*mA*mG*rGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArGrUrCrUrUrUrGrUrArCrUrCrUrGrGrUrArCrCrArGrArArGrCrU
24428



ED4-_
rArCrArArArGrArUrArArGrGrCrUrUrCrArUrGrCrCrGrArArArUrCrArArCrArCrCrCrUrGrUrCrArUrUrUrUrArUrGrGrCr




G_3FE_
ArGrGrGrUrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrU*mC*mA*mG




PB8







360
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24429



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB17
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArUrG*mU*mU*mU






361
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24430



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB16
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArU*mG*mU*mU






362
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24431



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB15
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrA*mU*mG*mU






363
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24432



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB14
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrC*mA*mU*mG






364
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24433



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB13
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrC*mC*mA*mU






365
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24434



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB12
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrG*mC*mC*mA






366
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24435



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB11
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrG*mG*mC*mC






367
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24436



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr




PB10
UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrG*mG*mG*mC






368
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24437



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr





UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrG*mG*mG*mG






369
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24438



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




30FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrAr





UrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrU*mG*mG*mG






370
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24439



ED15+_G_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB17
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArUrG*mU*mU*mU






371
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24440



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB16
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArU*mG*mU*mU






372
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24441



ED15+_G_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB15
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrA*mU*mG*mU






373
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24442



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB14
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrC*mA*mU*mG






374
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24443



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB13
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrC*mC*mA*mU






375
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24444



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB12
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrG*mC*mC*mA






376
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24445



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB11
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrG*mG*mC*mC






377
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24446



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr




PB10
GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrG*mG*mG*mC






378
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24447



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr





GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrG*mG*mG*mG






379
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24448



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




25FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrArUrArArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCr





GrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrU*mG*mG*mG






380
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24449



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB17
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArUrG*mU*mU*mU






381
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24450



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB16
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArU*mG*mU*mU






382
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24451



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB15
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrCrA*mU*mG*mU






383
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24452



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB14
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrCrC*mA*mU*mG






384
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24453



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




3PB1
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrGrC*mC*mA*mU






385
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24454



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB12
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrGrG*mC*mC*mA






386
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24455



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB11
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrGrG*mG*mC*mC






387
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24456



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr




PB10
ArGrGrGrArCrUrGrArArGrCrUrGrCrUrGrG*mG*mG*mC






388
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24457



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr





ArGrGrGrArCrUrGrArArGrCrUrGrCrUrG*mG*mG*mG






389
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24458



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




20FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrGrCrUrGrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArAr





ArGrGrGrArCrUrGrArArGrCrUrGrCrU*mG*mG*mG






390
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24459



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB17
UrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArUrG*mU*mU*mU






391
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24460



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB16
UrGrArArGrCrUrGrCrUrGrGrGrGrCrCrArU*mG*mU*mU






392
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24461



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB15
UrGrArArGrCrUrGrCrUrGrGrGrGrCrCrA*mU*mG*mU






393
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24462



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB14
UrGrArArGrCrUrGrCrUrGrGrGrGrCrC*mA*mU*mG






394
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24463



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB13
UrGrArArGrCrUrGrCrUrGrGrGrGrC*mC*mA*mU






395
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24464



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB12
UrGrArArGrCrUrGrCrUrGrGrGrG*mC*mC*mA






396
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24465



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB11
UrGrArArGrCrUrGrCrUrGrGrG*mG*mC*mC






397
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24466



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr




PB10
UrGrArArGrCrUrGrCrUrGrG*mG*mG*mC






398
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24467



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr





UrGrArArGrCrUrGrCrUrG*mG*mG*mG






399
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24468



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




14FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrCrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCr





UrGrArArGrCrUrGrCrU*mG*mG*mG






400
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24469



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB17
ArGrCrUrGrCrUrGrGrGrGrCrCrArUrG*mU*mU*mU






401
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24470



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB16
ArGrCrUrGrCrUrGrGrGrGrCrCrArU*mG*mU*mU






402
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24471



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB15
ArGrCrUrGrCrUrGrGrGrGrCrCrA*mU*mG*mU






403
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24472



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB14
ArGrCrUrGrCrUrGrGrGrGrCrC*mA*mU*mG






404
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24473



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB13
ArGrCrUrGrCrUrGrGrGrGrC*mC*mA*mU






405
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24474



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB12
ArGrCrUrGrCrUrGrGrGrG*mC*mC*mA






406
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24475



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB11
ArGrCrUrGrCrUrGrGrG*mG*mC*mC






407
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24476



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr




PB10
ArGrCrUrGrCrUrGrG*mG*mG*mC






408
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24477



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr





ArGrCrUrGrCrUrG*mG*mG*mG






409
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24478



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




11FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrGrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrAr





ArGrCrUrGrCrU*mG*mG*mG






410
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24479



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB17
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrGrGrGrCrCrArUrG*mU*mU*mU






411
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24480



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB16
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrGrGrGrCrCrArU*mG*mU*mU






412
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24481



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB15
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrGrGrGrCrCrA*mU*mG*mU






413
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24482



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB14
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrGrGrGrCrC*mA*mU*mG






414
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU




ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB13
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr
24483




CrUrGrCrUrGrGrGrGrC*mC*mA*mU






415
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24484



ED15+_G_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB12
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrGrGrG*mC*mC*mA






416
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24485



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB11
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrGrG*mG*mC*mC






417
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24486



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB10
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrGrG*mG*mG*mC






418
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24487



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrUrG*mG*mG*mG






419
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24488



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




9FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrCrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGr





CrUrGrCrU*mG*mG*mG






420
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24489



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB17
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrGrGrCrCrArUrG*mU*mU*mU






421
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24490



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB16
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrGrGrCrCrArU*mG*mU*mU






422
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24491



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB15
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrGrGrCrCrA*mU*mG*mU






423
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24492



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB14
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrGrGrCrC*mA*mU*mG






424
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24493



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB13
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrGrGrC*mC*mA*mU






425
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24494



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB12
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrGrG*mC*mC*mA






426
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24495



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB11
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrGrG*mG*mC*mC






427
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24496



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB10
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrGrG*mG*mG*mC






428
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24497



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrUrG*mG*mG*mG






429
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24498



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




6FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGr





CrU*mG*mG*mG






430
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24499



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB17
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrGrGrCrCrArUrG*mU*mU*mU






431
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24500



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB16
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrGrGrCrCrArU*mG*mU*mU






432
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24501



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB15
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrGrGrCrCrA*mU*mG*mU






433
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24502



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB14
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrGrGrCrC*mA*mU*mG






434
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24503



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB13
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrGrGrC*mC*mA*mU






435
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24504



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB12
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrGrG*mC*mC*mA






436
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24505



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB11
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrGrG*mG*mC*mC






437
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24506



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB10
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrGrG*mG*mG*mC






438
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24507



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





UrG*mG*mG*mG






439
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24508



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




5FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCr





U*mG*mG*mG






440
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24509



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB17
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrGrGrCrCrArUrG*mU*mU*mU






441
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24510



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB16
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrGrGrCrCrArU*mG*mU*mU






442
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24511



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB15
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrGrGrCrCrA*mU*mG*mU






443
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24512



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB14
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrGrGrCrC*mA*mU*mG






444
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24513



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB13
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrGrGrC*mC*mA*mU






445
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24514



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB12
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrGrG*mC*mC*mA






446
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24515



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB11
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





GrG*mG*mC*mC






447
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24516



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB10
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrGr





G*mG*mG*mC






448
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24517



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB9
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrUrG*





mG*mG*mG






449
A1AT_Nme2_
mU*mA*mA*rArArArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24518



ED15+_G_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




3FE_PB8
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrArCrGrArGrArArArGrGrGrArCrUrGrArArGrCrUrGrCrU*m





G*mG*mG






450
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24519



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




30FE_





PB17







451
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24520



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




30FE_





PB16







452
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24521



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArUrGrCrArC*mG*mG*mC




30FE_





PB15







453
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24522



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArUrGrCrA*mC*mG*mG




30FE_





PB14







454
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24523



9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArUrGrC*mA*mC*mG




30FE_





PB13







455
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24524



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArUrG*mC*mA*mC




30FE_





PB12







456
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24525



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrArU*mG*mC*mA




30FE_





PB11







457
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24526



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrUrA*mU*mG*mC




30FE_





PB10







458
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24527



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrUrU*mA*mU*mG




30FE_PB9







459
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24528



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




ED15-_
rGrCrCrU*mU*mA*mU




30FE_PB8







460
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24529



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED15-_
rArUrGrCrArCrGrG*mC*mC*mU




25FE_





PB17







461
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24530



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED15-_
rArUrGrCrArCrG*mG*mC*mC




25FE_





PB16







462
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24531



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED15-_
rArUrGrCrArC*mG*mG*mC




25FE_





PB15







463
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24532



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED 15-
rArUrGrCrA*mC*mG*mG




25FE_





PB14







464
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24533



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED 15-
rArUrGrC*mA*mC*mG




25FE_





PB13







465
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24534



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED15-_
rArUrG*mC*mA*mC




25FE_





PB12







466
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr




SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU
24535



ED15-_
rArU*mG*mC*mA




25FE_





PB11







467
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24536



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED15-_
rA*mU*mG*mC




25FE_





PB10







468
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24537



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU




ED15-_
*mA*mU*mG




25FE_PB9







469
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24538



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*m




ED15-_
U*mA*mU




25FE_PB8







470
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24539



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA




ED 15-
rCrGrG*mC*mC*mU




20FE_





PB17







471
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24540



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA




ED15-_
rCrG*mG*mC*mC




20FE_





PB16







472
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24541



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA




ED15-_
rC*mG*mG*mC




20FE_





PB15







473
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24542



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA




ED15-_
*mC*mG*mG




20FE_





PB14







474
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24543



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*




ED15-_
mA*mC*mG




20FE_





PB13







475
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24544



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC




ED15-_
*mA*mC




20FE_





PB12







476
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24545



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*




ED15-_
mC*mA




20FE_





PB11







477
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24546



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG




ED15-_
*mC




20FE_





PB10







478
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24547



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*




ED15-_
mG




20FE_PB9







479
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24548



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





20FE_PB8







480
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24549



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*




ED15-_
mC*mU




14FE_





PB17







481
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24550



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC




ED15-_
*mC




14FE_





PB16







482
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24551



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*




ED15-_
mC




14FE_





PB15







483
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24552



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




ED15-_





14FE_





PB14







484
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24553



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




ED15-_





14FE_





PB13







485
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24554



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




ED15-_





14FE_





PB12







486
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24555



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




ED15-_





14FE_





PB11







487
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24556



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




ED15-_





14FE_





PB10







488
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24557



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG




ED15-_





14FE_PB9







489
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24558



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





14FE_PB8







490
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24559



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




ED15-_





11FE_





PB17







491
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24560



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




ED15-_





11FE_





PB16







492
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24561



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




ED15-_





11FE_





PB15







493
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24562



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




ED15-_





11FE_





PB14







494
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24563



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




ED15-_





11FE_





PB13







495
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24564



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




ED15-_





11FE_





PB12







496
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24565



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




ED15-_





11FE_





PB11







497
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24566



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




ED15-_





11FE_





PB10







498
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24567



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG




ED15-_





11FE_PB9







499
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24568



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





11FE_PB8







500
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24569



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




ED15-_





9FE_PB17







501
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24570



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




ED15-_





9FE_PB16







502
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24571



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




ED15-_





9FE_PB15







503
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24572



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




ED15-_





9FE_PB14







504
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24573



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




ED15-_





9FE_PB13







505
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24574



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




ED15-_





9FE_PB12







506
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24575



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




ED15-_





9FE_PB11







507
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24576



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




ED15-_





9FE_PB10







508
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24577



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG




ED15-_





9FE_PB9







509
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24578



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





9FE_PB8







510
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24579



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




ED15-_





7FE_PB17







511
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24580



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




ED15-_





7FE_PB16







512
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24581



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




ED15-_





7FE_PB15







513
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24582



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




ED15-_





7FE_PB14







514
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24583



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




ED15-_





7FE_PB13







515
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24584



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




ED15-_





7FE_PB12







516
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24585



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




ED15-_





7FE_PB11







517
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24586



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




ED15-_





7FE_PB10







518
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24587



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG




ED15-_





7FE_PB9







519
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24588



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





7FE_PB8







520
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24589



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




ED15-_





5FE_PB17







521
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24590



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




ED15-_





5FE_PB16







522
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24591



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




ED15-_





5FE_PB15







523
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24592



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




ED15-_





5FE_PB14







524
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24593



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




ED15-_





5FE_PB13







525
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24594



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




ED15-_





5FE_PB12







526
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24595



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




ED15-_





5FE_PB11







527
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24596



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




ED15-_





5FE_PB10







528
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24597



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG




ED15-_





5FE_PB9







529
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24598



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





5FE_PB8







530
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24599



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




ED15-_





3FE_PB17







531
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24600



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




ED15-_





3FE_PB16







532
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24601



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




ED15-_





3FE_PB15







533
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24602



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




ED15-_





3FE_PB14







534
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24603



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




ED15-_





3FE_PB13







535
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24604



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




ED15-_





3FE_PB12







536
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24605



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




ED15-_





3FE_PB11







537
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24606



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




ED15-_





3FE_PB10







538
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24607



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG




ED15-_





3FE_PB9







539
A1AT_
mA*mG*mG*rCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUr
24608



SpCas9-
ArArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrU




NG_SpRY_
rGrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU




ED15-_





3FE_PB8







540
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24609



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB17
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






541
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24610



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB16
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






542
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24611



ED15-_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB15
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC






543
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24612



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB14
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG






544
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24613



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB13
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG






545
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24614



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB12
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC






546
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24615



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB11
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA






547
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24616



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB10
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC






548
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24617



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB9
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG






549
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24618



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_30FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrU




PB8
rUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU






550
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24619



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB17
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






551
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24620



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB16
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






552
A1AT_Nme2_
mU*mC*mC*ArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24621



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB15
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC






553
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24622



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB14
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG






554
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24623



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB13
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG






555
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24624



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB12
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC






556
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24625



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB11
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA






557
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24626



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB10
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC






558
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24627



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB9
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG






559
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24628



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_25FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrC




PB8
rGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU






560
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24629



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB17
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






561
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24630



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB16
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






562
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24631



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB15
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC






563
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24632



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB14
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG






564
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24633



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB13
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG






565
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24634



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB12
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC






566
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24635



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB11
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA






567
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24636



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB10
rUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC






568
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24637



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB9
rUrGrGrUrCrArGrCrArCrArGrCrCrUrU*mA*mU*mG






569
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24638



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_20FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrA




PB8
rUrGrGrUrCrArGrCrArCrArGrCrCrU*mU*mA*mU






570
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24639



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB17
ArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






571
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24640



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB16
ArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






572
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24641



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB15
ArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC






573
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24642



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB14
ArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG






574
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24643



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB13
ArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG






575
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24644



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB12
ArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC






576
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24645



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB11
ArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA






577
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24646



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB10
ArGrCrArCrArGrCrCrUrUrA*mU*mG*mC






578
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24647



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB9
ArGrCrArCrArGrCrCrUrU*mA*mU*mG






579
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24648



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_14FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCr




PB8
ArGrCrArCrArGrCrCrU*mU*mA*mU






580
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24649



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB17
ArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






581
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24650



ED15-_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB16
ArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






582
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24651



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB15
ArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC






583
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24652



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB14
ArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG






584
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24653



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB13
ArCrArGrCrCrUrUrArUrGrC*mA*mC*mG






585
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24654



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB12
ArCrArGrCrCrUrUrArUrG*mC*mA*mC






586
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24655



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB11
ArCrArGrCrCrUrUrArU*mG*mC*mA






587
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24656



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB10
ArCrArGrCrCrUrUrA*mU*mG*mC






588
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24657



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB9
ArCrArGrCrCrUrU*mA*mU*mG






589
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24658



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_11FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCr




PB8
ArCrArGrCrCrU*mU*mA*mU






590
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24659



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB17
rGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






591
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24660



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB16
rGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






592
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24661



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB15
rGrCrCrUrUrArUrGrCrArC*mG*mG*mC






593
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24662



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB14
rGrCrCrUrUrArUrGrCrA*mC*mG*mG






594
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24663



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB13
rGrCrCrUrUrArUrGrC*mA*mC*mG






595
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24664



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB12
rGrCrCrUrUrArUrG*mC*mA*mC






596
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24665



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB11
rGrCrCrUrUrArU*mG*mC*mA






597
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24666



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB10
rGrCrCrUrUrA*mU*mG*mC






598
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24667



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB9
rGrCrCrUrU*mA*mU*mG






599
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24668



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_9FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrA




PB8
rGrCrCrU*mU*mA*mU






600
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24669



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB17
rCrUrUrArUrGrCrArCrGrG*mC*mC*mU






601
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24670



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB16
rCrUrUrArUrGrCrArCrG*mG*mC*mC






602
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24671



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB15
rCrUrUrArUrGrCrArC*mG*mG*mC






603
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24672



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB14
rCrUrUrArUrGrCrA*mC*mG*mG






604
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24673



ED15-_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB13
rCrUrUrArUrGrC*mA*mC*mG






605
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24674



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB12
rCrUrUrArUrG*mC*mA*mC






606
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24675



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB11
rCrUrUrArU*mG*mC*mA






607
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24676



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB10
rCrUrUrA*mU*mG*mC






608
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24677



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB9
rCrUrU*mA*mU*mG






609
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24678



ED15-_
ArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_7FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrC




PB8
rCrU*mU*mA*mU






610
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24679



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB17
UrUrArUrGrCrArCrGrG*mC*mC*mU






611
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24680



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB16
UrUrArUrGrCrArCrG*mG*mC*mC






612
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24681



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB15
UrUrArUrGrCrArC*mG*mG*mC






613
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24682



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB14
UrUrArUrGrCrA*mC*mG*mG






614
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24683



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB13
UrUrArUrGrC*mA*mC*mG






615
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24684



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB12
UrUrArUrG*mC*mA*mC






616
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24685



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB11
UrUrArU*mG*mC*mA






617
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24686



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB10
UrUrA*mU*mG*mC






618
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24687



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB9
UrU*mA*mU*mG






619
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24688



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_5FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCr




PB8
U*mU*mA*mU






620
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24689



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB17
ArUrGrCrArCrGrG*mC*mC*mU






621
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24690



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB16
ArUrGrCrArCrG*mG*mC*mC






622
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24691



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB15
ArUrGrCrArC*mG*mG*mC






623
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24692



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB14
ArUrGrCrA*mC*mG*mG






624
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24693



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB13
ArUrGrC*mA*mC*mG






625
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24694



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB12
ArUrG*mC*mA*mC






626
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24695



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB11
ArU*mG*mC*mA






627
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24696



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB10
A*mU*mG*mC






628
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24697



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrU*




PB9
mA*mU*mG






629
A1AT_Nme2_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrGrGrUrUrGrUrArGrCrUrCrCrCrGrArArArCrGrUrUrGrCrU
24698



ED15-_
rArCrArArUrArArGrGrCrCrGrUrCrUrGrArArArArGrArUrGrUrGrCrCrGrCrArArCrGrCrUrCrUrGrCrCrCrCrUrUrArArArG




G_3FE_
rCrUrUrCrUrGrCrUrUrUrArArGrGrGrGrCrArUrCrGrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrU*mU




PB8
*mA*mU






630
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24699



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




PB17







631
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24700



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




PB16







632
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24701



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




PB15







633
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24702



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




PB14







634
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24703



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrCrArC*mG*mG*mC




PB13







635
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24704



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrCrA*mC*mG*mG




PB12







636
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24705



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrGrC*mA*mC*mG




PB11







637
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24706



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_
GrCrCrUrUrArUrG*mC*mA*mC




PB10







638
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24707



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_PB9
GrCrCrUrUrArU*mG*mC*mA






639
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24708



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




30FE_PB8
GrCrCrUrUrA*mU*mG*mC






640
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24709



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrCrArCrGrGrCrC*mU*mG*mG




PB17







641
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24710



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrCrArCrGrGrC*mC*mU*mG




PB16







642
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24711



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrCrArCrGrG*mC*mC*mU




PB15







643
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24712



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrCrArCrG*mG*mC*mC




PB14







644
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24713



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrCrArC*mG*mG*mC




PB13







645
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24714



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrCrA*mC*mG*mG




PB12







646
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24715



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrGrC*mA*mC*mG




PB11







647
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24716



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_
ArUrG*mC*mA*mC




PB10







648
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24717



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_PB9
ArU*mG*mC*mA






649
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24718



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




25FE_PB8
A*mU*mG*mC






650
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24719



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




20FE_
CrGrGrCrC*mU*mG*mG




PB17







651
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24720



SpCas9-
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




20FE_
CrGrGrC*mC*mU*mG




PB16







652
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24721



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




20FE_
CrGrG*mC*mC*mU




PB15







653
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24722



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




20FE_
CrG*mG*mC*mC




PB14







654
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24723



SpCas9-
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




20FE_
C*mG*mG*mC




PB13







655
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24724



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA




20FE_
*mC*mG*mG




PB12







656
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24725



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*




20FE_
mA*mC*mG




PB11







657
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24726



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*




20FE_
mA*mC




PB10







658
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24727



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*




20FE_PB9
mC*mA






659
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24728



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*




20FE_PB8
mC






660
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24729



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*




14FE_
mU*mG*mG




PB17







661
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24730



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*




14FE_
mU*mG




PB16







662
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24731



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*




14FE_
mC*mU




PB15







663
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24732



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*




14FE_
mC




PB14







664
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24733



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*




14FE_
mC




PB13







665
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24734



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




14FE_





PB12







666
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24735



SpCas9-
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




14FE_





PB11







667
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24736



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




14FE_





PB10







668
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24737



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




14FE_PB9







669
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24738



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




14FE_PB8







670
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24739



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*




11FE_
mG




PB17







671
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24740



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*




11FE_
mG




PB16







672
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24741



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




11FE_





PB15







673
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24742



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




11FE_





PB14







674
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24743



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




11FE_





PB13







675
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24744



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




11FE_





PB12







676
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24745



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




11FE_





PB11







677
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24746



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




11FE_





PB10







678
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24747



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




11FE_PB9







679
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24748



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




11FE_PB8







680
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24749



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




9FE_PB17







681
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24750



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




9FE_PB16







682
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24751



SpCas9-
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




9FE_PB15







683
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24752



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




9FE_PB14







684
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24753



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




9FE_PB13







685
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24754



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




9FE_PB12







686
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24755



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




9FE_PB11







687
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24756



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




9FE_PB10







688
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24757



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




9FE_PB9







689
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24758



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




9FE_PB8







690
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24759



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




7FE_PB17







691
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24760



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




7FE_PB16







692
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24761



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




7FE_PB15







693
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24762



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




7FE_PB14







694
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24763



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




7FE_PB13







695
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24764



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




7FE_PB12







696
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24765



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




7FE_PB11







697
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24766



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




7FE_PB10







698
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24767



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




7FE_PB9







699
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24768



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




7FE_PB8







700
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24769



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




5FE_PB17







701
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24770



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




5FE_PB16







702
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24771



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




5FE_PB15







703
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24772



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




5FE_PB14







704
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24773



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




5FE_PB13







705
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24774



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




5FE_PB12







706
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24775



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




5FE_PB11







707
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24776



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




5FE_PB10







708
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24777



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




5FE_PB9







709
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24778



SpCas9-
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




5FE_PB8







710
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24779



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




3FE_PB17







711
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24780



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




3FE_PB16







712
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24781



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




3FE_PB15







713
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24782



SpCas9-
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




3FE_PB14







714
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24783



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




3FE_PB13







715
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24784



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




3FE_PB12







716
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24785



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




3FE_PB11







717
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24786



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




3FE_PB10







718
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24787



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




3FE_PB9







719
A1AT_
mC*mC*mA*rGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24788



SpCas9-
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




NG_ED17-_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrA*mU*mG*mC




3FE_PB8







720
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24789



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB17
GrCrCrUrUrArUrGrCrArCrGrGrCrCrU*mG*mG*mA






721
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24790



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB16
GrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG






722
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24791



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB15
GrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG






723
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24792



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB14
GrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU






724
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24793



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB13
GrCrCrUrUrArUrGrCrArCrG*mG*mC*mC






725
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24794



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB12
GrCrCrUrUrArUrGrCrArC*mG*mG*mC






726
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24795



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB11
GrCrCrUrUrArUrGrCrA*mC*mG*mG






727
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24796



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB10
GrCrCrUrUrArUrGrC*mA*mC*mG






728
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24797



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB9
GrCrCrUrUrArUrG*mC*mA*mC






729
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24798



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_30FE_
GrCrArCrArUrGrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrAr




PB8
GrCrCrUrUrArU*mG*mC*mA






730
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24799



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB17
ArUrGrCrArCrGrGrCrCrU*mG*mG*mA






731
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24800



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB16
ArUrGrCrArCrGrGrCrC*mU*mG*mG






732
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24801



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB15
ArUrGrCrArCrGrGrC*mC*mU*mG






733
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24802



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB14
ArUrGrCrArCrGrG*mC*mC*mU






734
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24803



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB13
ArUrGrCrArCrG*mG*mC*mC






735
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24804



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB12
ArUrGrCrArC*mG*mG*mC






736
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24805



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB11
ArUrGrCrA*mC*mG*mG






737
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24806



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB10
ArUrGrC*mA*mC*mG






738
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24807



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB9
ArUrG*mC*mA*mC






739
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24808



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_25FE_
GrCrGrCrCrCrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUr




PB8
ArU*mG*mC*mA






740
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24809



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




PB17
CrGrGrCrCrU*mG*mG*mA






741
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24810



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




PB16
CrGrGrCrC*mU*mG*mG






742
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24811



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




PB15
CrGrGrC*mC*mU*mG






743
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24812



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




PB14
CrGrG*mC*mC*mU






744
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24813



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




PB13
CrG*mG*mC*mC






745
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24814



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrAr




PB12
C*mG*mG*mC






746
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24815



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA




PB11
*mC*mG*mG






747
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24816



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*




PB10
mA*mC*mG






748
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24817



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*




PB9
mA*mC






749
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24818



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_20FE_
GrCrArGrCrArGrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*




PB8
mC*mA






750
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24819



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrCrU*




PB17
mG*mG*mA






751
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24820



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*




PB16
mU*mG*mG






752
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24821



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*




PB15
mU*mG






753
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24822



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*




PB14
mC*mU






754
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24823



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*




PB13
mC






755
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24824



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*




PB12
mC






756
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24825



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




PB11







757
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24826



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




PB10







758
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24827



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




PB9







759
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24828



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_14FE_
GrCrUrUrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




PB8







760
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24829



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrCrU*mG*




PB17
mG*mA






761
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24830



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*




PB16
mG






762
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24831



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*




PB15
mG






763
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24832



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




PB14







764
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24833



ED18-_
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




PB13







765
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24834



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




PB12







766
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24835



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




PB11







767
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24836



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




PB10







768
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24837



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




PB9







769
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24838



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_11FE_
GrCrArGrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




PB8







770
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24839



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrCrU*mG*mG*




PB17
mA






771
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24840



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




PB16







772
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24841



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




PB15







773
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24842



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




PB14







774
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24843



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




PB13







775
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24844



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




PB12







776
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24845



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




PB11







777
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24846



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




PB10







778
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24847



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




PB9







779
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24848



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_9FE_
GrCrUrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




PB8







780
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24849



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrCrU*mG*mG*mA




PB17







781
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24850



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




PB16







782
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24851



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




PB15







783
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24852



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




PB14







784
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24853



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




PB13







785
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24854



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




PB12







786
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24855



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




PB11







787
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24856



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




PB10







788
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24857



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




PB9







789
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24858



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_7FE_
GrCrCrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUmG*mC*mA




PB8







790
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24859



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrCrU*mG*mG*mA




PB17







791
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24860



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




PB16







792
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24861



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




PB15







793
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24862



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




PB14







794
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24863



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




PB13







795
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24864



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




PB12







796
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24865



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




PB11







797
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24866



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




PB10







798
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24867



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




PB9







799
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24868



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_5FE_
GrCrUrUrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




PB8







800
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24869



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrCrU*mG*mG*mA




PB17







801
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24870



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrCrC*mU*mG*mG




PB16







802
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24871



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrGrC*mC*mU*mG




PB15







803
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24872



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrGrG*mC*mC*mU




PB14







804
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24873



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArCrG*mG*mC*mC




PB13







805
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24874



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrArC*mG*mG*mC




PB12







806
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24875



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrCrA*mC*mG*mG




PB11







807
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24876



ED18-_
ArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrGrC*mA*mC*mG




PB10







808
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24877



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArUrG*mC*mA*mC




PB9







809
A1AT_SpRY_
mU*mC*mC*rArGrGrCrCrGrUrGrCrArUrArArGrGrCrUrGrUrUrUrUrArGrArGrCrUrArGrArArArUrArGrCrArArGrUrUrA
24878



ED18-_
rArArArUrArArGrGrCrUrArGrUrCrCrGrUrUrArUrCrArArCrUrUrGrArArArArArGrUrGrGrCrArCrCrGrArGrUrCrGrGrUr




G_3FE_
GrCrUrCrUrCrGrUrCrGrArUrGrGrUrCrArGrCrArCrArGrCrCrUrUrArU*mG*mC*mA




PB8









Table X3A shows the sequences of X3 without modifications. In some embodiments, the sequences used in this table can be used without chemical modifications.









TABLE X3A







Table X3 Sequences without Modifications













SEQ





ID


ID
tgRNA Name
tgNA_seq_IDT_formatted
NO













0
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24879



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAG




PB17







1
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24880



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACA




PB16







2
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24881



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CAC




PB15







3
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24882



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CA




PB14







4
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24883



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
C




PB13







5
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24884



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_





PB12







6
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24885



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




30FE_





PB11







7
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24886



SpRY_ED0-_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




G_





30FE_





PB10







8
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24887



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGU




30FE_





PB9







9
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24888



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGG




30FE_





PB8







10
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24889



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_





PB17







11
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24890



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




25FE_





PB16







12
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24891



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




25FE_





PB15







13
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24892



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




25FE_





PB14







14
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24893



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




25FE_





PB13







15
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24894



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




25FE_





PB12







16
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24895



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




25FE_





PB11







17
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24896



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




25FE_





PB10







18
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24897



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGU




25FE_





PB9







19
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24898



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGG




ED0-_G_





25FE_





PB8







20
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24899



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




20FE_





PB17







21
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24900



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




ED0-_G_





20FE_





PB16







22
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24901



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




20FE_





PB15







23
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24902



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




ED0-_G_





20FE_





PB14







24
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24903



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




ED0-_G_





20FE_





PB13







25
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24904



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




20FE_





PB12







26
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24905



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




20FE_





PB11







27
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24906



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




20FE_





PB10







28
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24907



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGU




20FE_





PB9







29
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24908



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGG




20FE_





PB8







30
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24909



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




14FE_





PB17







31
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24910



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




14FE_





PB16







32
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24911



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




14FE_





PB11_





5







33
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24912



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




14FE_





PB14







34
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24913



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




14FE_





PB13







35
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24914



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




14FE_





PB12







36
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24915



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCA




14FE_





PB11







37
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24916



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUC




14FE_





PB10







38
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24917



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGU




14FE_





PB9







39
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24918



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGG




14FE_





PB8







40
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24919



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




11FE_





PB17







41
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24920



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACA




11FE_





PB16







42
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24921



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCAC




11FE_





PB15







43
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24922



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCA




11FE_





PB14







44
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24923



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGC




ED0-_G_





11FE_





PB13







45
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24924



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAG




11FE_





PB12







46
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24925



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCA




11FE_





PB11







47
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24926



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUC




ED0-_G_





11FE_





PB10







48
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24927



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGU




11FE_





PB9







49
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24928



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGG




11FE_





PB8







50
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24929



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAG




9FE_





PB17







51
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24930



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACA




9FE_





PB16







52
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24931



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCAC




9FE_





PB15







53
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24932



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCA




9FE_





PB14







54
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24933



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGC




9FE_





PB13







55
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24934



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAG




9FE_





PB12







56
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24935



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCA




9FE_





PB11







57
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24936



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUC




9FE_





PB10







58
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24937



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGU




9FE_





PB9







59
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24938



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGG




9FE_





PB8







60
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24939



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAG




7FE_





PB17







61
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24940



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACA




7FE_





PB16







62
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24941



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCAC




7FE_





PB15







63
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24942



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCA




7FE_





PB14







64
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24943



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGC




7FE_





PB13







65
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24944



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAG




7FE_





PB12







66
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24945



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCA




7FE_





PB11







67
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24946



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUC




7FE_





PB10







68
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24947



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGU




7FE_





PB9







69
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24948



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGG




7FE_





PB8







70
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24949



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAG




5FE_





PB17







71
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24950



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACA




5FE_





PB16







72
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24951



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCAC




5FE_





PB15







73
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24952



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCA




5FE_





PB14







74
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24953



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGC




5FE_





PB13







75
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24954



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAG




5FE_





PB12







76
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24955



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCA




5FE_





PB11







77
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24956



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUC




5FE_





PB10







78
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24957



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGU




5FE_





PB9







79
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24958



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGG




5FE_





PB8







80
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24959



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAG




3FE_





PB17







81
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24960



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACA




3FE_





PB16







82
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24961



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCAC




3FE_





PB15







83
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24962



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCA




3FE_





PB14







84
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24963



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGC




3FE_





PB13







85
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24964



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAG




3FE_





PB12







86
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24965



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCA




3FE_





PB11







87
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24966



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUC




3FE_





PB10







88
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24967



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGU




3FE_





PB9







89
A1AT_
CUGUGCUGACCAUCGACAAGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24968



SpRY_ED0-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGG




3FE_





PB8







90
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24969



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB17
CACAGCC






91
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24970



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB16
CACAGC






92
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24971



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB15
CACAG






93
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24972



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB14
CACA






94
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24973



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB13
CAC






95
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24974



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB12
CA






96
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24975



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB11
C






97
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24976



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB10







98
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24977



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




PB9







99
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24978



ED2-_30FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




PB8







100
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24979



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




PB17
CC






101
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24980



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




PB16
C






102
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24981



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




PB15







103
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24982



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




PB14







104
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24983



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




PB13







105
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24984



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




PB12







106
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24985



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




PB11







107
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24986



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB10







108
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24987



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




PB9







109
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24988



ED2-_25FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




PB8







110
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24989



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




PB17







111
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24990



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




PB16







112
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24991



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




PB15







113
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24992



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




PB14







114
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24993



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




PB13







115
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24994



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




PB12







116
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24995



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




PB11







117
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24996



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB10







118
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24997



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




PB9







119
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24998



ED2-_20FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




PB8







120
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
24999



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




PB17







121
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25000



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




PB16







122
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25001



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




PB15







123
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25002



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




PB14







124
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25003



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




PB13







125
A1AT_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25004



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




ED2-_14FE_





PB12







126
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25005



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




PB11







127
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25006



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




PB10







128
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25007



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCA




PB9







129
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25008



ED2-_14FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUC




PB8







130
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25009



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




PB17







131
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25010



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




PB16







132
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25011



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




PB15







133
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25012



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACA




PB14







134
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25013



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCAC




PB13







135
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25014



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCA




PB12







136
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25015



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGC




PB11







137
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25016



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAG




PB10







138
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25017



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCA




PB9







139
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25018



ED2-_11FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUC




PB8







140
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25019



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCC




PB17







141
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25020



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGC




PB16







142
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25021



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAG




PB15







143
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25022



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACA




PB14







144
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25023



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCAC




PB13







145
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25024



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCA




PB12







146
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25025



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGC




PB11







147
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25026



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAG




PB10







148
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25027



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCA




PB9







149
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25028



ED2-_9FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUC




PB8







150
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25029



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCC




PB17







151
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25030



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGC




PB16







152
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25031



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAG




PB15







153
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25032



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACA




PB14







154
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25033



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCAC




PB13







155
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25034



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCA




PB12







156
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25035



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGC




PB11







157
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25036



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAG




PB10







158
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25037



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCA




PB9







159
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25038



ED2-_7FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUC




PB8







160
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25039



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCC




PB17







161
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25040



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGC




PB16







162
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25041



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAG




PB15







163
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25042



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACA




PB14







164
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25043



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCAC




PB13







165
A1AT_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25044



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCA




ED2-_5FE_





PB12







166
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25045



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGC




PB11







167
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25046



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAG




PB10







168
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25047



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCA




PB9







169
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25048



ED2-_5FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUC




PB8







170
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25049



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCC




PB17







171
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25050



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGC




PB16







172
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25051



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAG




PB15







173
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25052



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACA




PB14







174
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25053



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCAC




PB13







175
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25054



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCA




PB12







176
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25055



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGC




PB11







177
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25056



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAG




PB10







178
A1AT_SpRY_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25057



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCA




PB9







179
AIAT_
GGCUGUGCUGACCAUCGACAGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25058



ED2-_3FE_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUC




PB8







180
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25059



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCU




ED3-_30FE_





PB17







181
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25060



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCC




ED3-_30FE_





PB16







182
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25061



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGC




ED3-_30FE_





PB15







183
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25062



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAG




ED3-_30FE_





PB14







184
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25063



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACA




ED3-_30FE_





PB13







185
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25064



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CAC




ED3-_30FE_





PB12







186
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25065



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CA




ED3-_30FE_





PB11







187
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25066



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
C




ED3-_30FE_





PB10







188
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25067



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_





ED3-_30FE_





PB9







189
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25068



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_30FE_





PB8







190
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25069



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCU




ED3-_25FE_





PB17







191
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25070



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CC




ED3-_25FE_





PB16







192
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25071



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
C




ED3-_25FE_





PB15







193
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25072



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_25FE_





PB14







194
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25073



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_25FE_





PB13







195
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25074



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_25FE_





PB12







196
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25075



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_25FE_





PB11







197
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25076



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_25FE_





PB10







198
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25077



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_





ED3-_25FE_





PB9







199
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25078



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_25FE_





PB8







200
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25079



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_20FE_





PB17







201
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25080



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_20FE_





PB16







202
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25081



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_20FE_





PB15







203
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25082



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_20FE_





PB14







204
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25083



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_20FE_





PB13







205
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25084



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_20FE_





PB12







206
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25085



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_20FE_





PB11







207
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25086



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_20FE_





PB10







208
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25087



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_





E





D3-_20FE_





PB9







209
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25088



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_20FE_





PB8







210
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25089



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_14FE_





PB17







211
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25090



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_14FE_





PB16







212
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25091



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_14FE_





PB15







213
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25092



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_14FE_





PB14







214
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25093



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_14FE_





PB13







215
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25094



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_14FE_





PB12







216
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25095



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_14FE_





PB11







217
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25096



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_14FE_





PB10







218
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25097



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_





E





D3-_14FE_





PB9







219
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25098



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_14FE_





PB8







220
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25099



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_11FE_





PB17







221
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25100



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_11FE_





PB16







222
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25101



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_11FE_





PB15







223
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25102



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_11FE_





PB14







224
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25103



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_11FE_





PB13







225
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25104



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_11FE_





PB12







226
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25105



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_11FE_





PB11







227
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25106



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_11FE_





PB10







228
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25107



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_





ED3-_11FE_





PB9







229
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25108



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_11FE_





PB8







230
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25109



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_9FE_





PB17







231
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25110



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_9FE_





PB16







232
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25111



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_9FE_





PB15







233
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25112



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_9FE_





PB14







234
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25113



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_9FE_





PB13







235
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25114



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_9FE_





PB12







236
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25115



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_9FE_





PB11







237
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25116



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_9FE_





PB10







238
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25117



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAG




SpRY_





ED3-_





9FE_





PB9







239
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25118



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_





9FE_





PB8







240
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25119



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_7FE_





PB17







241
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25120



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_7FE_





PB16







242
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25121



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_7FE_





PB15







243
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25122



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_7FE_





PB14







244
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25123



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_7FE_





PB13







245
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25124



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_7FE_





PB12







246
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25125



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_7FE_





PB11







247
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25126



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_7FE_





PB10







248
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25127



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAG




SpRY_





ED3-_





7FE_





PB9







249
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25128



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_





7FE_





PB8







250
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25129



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_5FE_





PB17







251
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25130



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_5FE_





PB16







252
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25131



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_5FE_





PB15







253
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25132



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_5FE_





PB14







254
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25133



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_5FE_





PB13







255
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25134



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_5FE_





PB12







256
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25135



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_5FE_





PB11







257
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25136



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGC




SpRY_





ED3-_5FE_





PB10







258
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25137



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAG




SpRY_





ED3-_





5FE_





PB9







259
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25138



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCA




SpRY_





ED3-_





5FE_





PB8







260
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25139



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCU




SpRY_





ED3-_3FE_





PB17







261_
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25140





1
ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCC




SpRY_





ED3-_3FE_





PB16







262
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25141



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGC




SpRY_





ED3-_3FE_





PB15







263
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25142



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAG




SpRY_





ED3-_3FE_





PB14







264
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25143



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACA




SpRY_





ED3-_3FE_





PB13







265
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25144



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCAC




SpRY_





ED3-_3FE_





PB12







266
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25145



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCA




SpRY_





ED3-_3FE_





PB11







267
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25146



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGC




SpRY_





ED3-_3FE_





PB10







268
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25147



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAG




SpRY_





ED3-_





3FE_





PB9







269
A1AT_
AGGCUGUGCUGACCAUCGACGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25148



ScaCas9++_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCA




SpRY_





ED3-_





3FE_





PB8







270
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25149



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCACAGCCUU




30FE_





PB17







271
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25150



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCACAGCCU




30FE_





PB16







272
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25151



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCACAGCC




30FE_





PB15







273
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25152



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCACAGC




30FE_





PB14







274
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25153



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCACAG




30FE_





PB13







275
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25154



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCACA




30FE_





PB12







276
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25155



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCAC




30FE_





PB11







277
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25156



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGCA




30FE_





PB10







278
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25157



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAGC




30FE_





PB9







279
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25158



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGA




ED4-_G_
UGGUCAG




30FE_





PB8







280
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25159



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCACAGCCUU




25FE_





PB17







281
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25160



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCACAGCCU




25FE_





PB16







282
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25161



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCACAGCC




25FE_





PB15







283
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25162



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCACAGC




25FE_





PB14







284
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25163



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCACAG




25FE_





PB13







285
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25164



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCACA




25FE_





PB12







286
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25165



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCAC




25FE_





PB11







287
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25166



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGCA




25FE_





PB10







288
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25167



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AGC




25FE_





PB9







289
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25168



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUC




ED4-_G_
AG




25FE_





PB8







290
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25169



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_
AGCCUU




20FE_





PB17







291
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25170



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_
AGCCU




20FE_





PB16







292
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25171



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_
AGCC




20FE_





PB15







293
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25172



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_
AGC




20FE_





PB14







294
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25173



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_
AG




20FE_





PB13







295
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25174



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_
A




20FE_





PB12







296
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25175



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





20FE_





PB11







297
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25176



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




ED4-_G_





20FE_





PB10







298
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25177



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGC




ED4-_G_





20FE_





PB9







299
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25178



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED4-_G_





20FE_





PB8







300
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25179



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUU




ED4-_G_





14FE_





PB17







301
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25180



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




ED4-_G_





14FE_





PB16







302
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25181



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




ED4-_G_





14FE_





PB15







303
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25182



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




ED4-_G_





14FE_





PB14







304
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25183



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED4-_G_





14FE_





PB13







305
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25184



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCACA




ED4-_G_





14FE_





PB12







306
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25185



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





14FE_





PB11







307
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25186



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGCA




ED4-_G_





14FE_





PB10







308
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25187



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAGC




ED4-_G_





14FE_





PB9







309
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25188



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED4-_G_





14FE_





PB8







310
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25189



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUU




ED4-_G_





11FE_





PB17







311
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25190



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




ED4-_G_





11FE_





PB16







312
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25191



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCACAGCC




ED4-_G_





11FE_





PB15







313
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25192



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCACAGC




ED4-_G_





11FE_





PB14







314
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25193



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED4-_G_





11FE_





PB13







315
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25194



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCACA




ED4-_G_





11FE_





PB12







316
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25195



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





11FE_





PB11







317
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25196



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGCA




ED4-_G_





11FE_





PB10







318
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25197



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAGC




ED4-_G_





11FE_





PB9







319
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25198



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUAGUCCCUUUCUCGUCGAUGGUCAG




ED4-_G_





11FE_





PB8







320
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25199



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCACAGCCUU




ED4-_G_





9FE_





PB17







321
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25200



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCACAGCCU




ED4-_G_





9FE_





PB16







322
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25201



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCACAGCC




ED4-_G_





9FE_





PB15







323
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25202



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCACAGC




ED4-_G_





9FE_





PB14







324
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25203



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCACAG




ED4-_G_





9FE_





PB13







325
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25204



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCACA




ED4-_G_





9FE_





PB12







326
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25205



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





9FE_





PB11







327
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25206



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGCA




ED4-_G_





9FE_





PB10







328
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25207



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAGC




ED4-_G_





9FE_





PB9







329
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25208



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCCCUUUCUCGUCGAUGGUCAG




ED4-_G_





9FE_





PB8







330
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25209



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCACAGCCUU




ED4-_G_





7FE_





PB17







331
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25210



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCACAGCCU




ED4-_G_





7FE_





PB16







332
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25211



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCACAGCC




ED4-_G_





7FE_





PB15







333
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25212



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCACAGC




ED4-_G_





7FE_





PB14







334
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25213



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCACAG




ED4-_G_





7FE_





PB13







335
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25214



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCACA




ED4-_G_





7FE_





PB12







336
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25215



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





7FE_





PB11







337
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25216



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGCA




ED4-_G_





7FE_





PB10







338
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25217



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAGC




ED4-_G_





7FE_





PB9







339
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25218



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUCCUUUCUCGUCGAUGGUCAG




ED4-_G_





7FE_





PB8







340
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25219



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCACAGCCUU




ED4-_G_





5FE_





PB17







341
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25220



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCACAGCCU




ED4-_G_





5FE_





PB16







342
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25221



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCACAGCC




ED4-_G_





5FE_





PB15







343
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25222



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCACAGC




ED4-_G_





5FE_





PB14







344
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25223



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCACAG




ED4-_G_





5FE_





PB13







345
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25224



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCACA




ED4-_G_





5FE_





PB12







346
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25225



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





5FE_





PB11







347
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25226



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGCA




ED4-_G_





5FE_





PB10







348
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25227



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAGC




ED4-_G_





5FE_





PB9







349
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25228



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUUUCUCGUCGAUGGUCAG




ED4-_G_





5FE_





PB8







350
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25229



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCACAGCCUU




ED4-_G_





3FE_





PB17







351
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25230



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCACAGCCU




ED4-_G_





3FE_





PB16







352
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25231



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCACAGCC




ED4-_G_





3FE_





PB15







353
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25232



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCACAGC




ED4-_G_





3FE_





PB14







354
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25233



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCACAG




ED4-_G_





3FE_





PB13







355
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25234



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCACA




ED4-_G_





3FE_





PB12







356
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25235



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCAC




ED4-_G_





3FE_





PB11







357
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25236



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGCA




ED4-_G_





3FE_





PB10







358
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25237



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAGC




ED4-_G_





3FE_





PB9







359
A1AT_
AAGGCUGUGCUGACCAUCGAGUCUUUGUACUCUGGUACCAGAAGCUACAAAGAUAAGGCUUCAUGCCGAA
25238



St1_
AUCAACACCCUGUCAUUUUAUGGCAGGGUGUUUUUCUCGUCGAUGGUCAG




ED4-_G_





3FE_





PB8







360
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25239



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCCAUGUUU




30FE_





PB17







361
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25240



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCCAUGUU




30FE_





PB16







362
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25241



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCCAUGU




30FE_





PB15







363
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25242



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCCAUG




30FE_





PB14







364
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25243



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCCAU




30FE_





PB13







365
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25244



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCCA




30FE_





PB12







366
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25245



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGCC




30FE_





PB11







367
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25246



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGGC




30FE_





PB10







368
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25247



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGGG




30FE_





PB9







369
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25248



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCGUGCAUAAGGCUGUGCUGA




G_
CCAUCGACGAGAAAGGGACUGAAGCUGCUGGG




30FE_





PB8







370
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25249



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCCAUGUUU




25FE_





PB17







371
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25250



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCCAUGUU




25FE_





PB16







372
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25251



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCCAUGU




25FE_





PB15







373
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25252



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCCAUG




25FE_





PB14







374
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25253



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCCAU




25FE_





PB13







375
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25254



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCCA




25FE_





PB12







376
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25255



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGCC




25FE_





PB11







377
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25256



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGGC




25FE_





PB10







378
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25257



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGGG




25FE_





PB9







379
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25258



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCAUAAGGCUGUGCUGACCAUC




G_
GACGAGAAAGGGACUGAAGCUGCUGGG




25FE_





PB8







380
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25259



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCCAUGUUU




20FE_





PB17







381
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25260



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCCAUGUU




20FE_





PB16







382
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25261



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCCAUGU




20FE_





PB15







383
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25262



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCCAUG




20FE_





PB14







384
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25263



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCCAU




20FE_





PB13







385
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25264



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCCA




20FE_





PB12







386
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25265



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGCC




20FE_





PB11







387
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25266



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGGC




20FE_





PB10







388
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25267



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGGG




20FE_





PB9







389
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25268



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGGCUGUGCUGACCAUCGACGA




G_
GAAAGGGACUGAAGCUGCUGGG




20FE_





PB8







390
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25269



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCCAUGUUU




14FE_





PB17







391
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25270



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCCAUGUU




14FE_





PB16







392
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25271



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCCAUGU




14FE_





PB15







393
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25272



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCCAUG




14FE_





PB14







394
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25273



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCCAU




14FE_





PB13







395
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25274



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCCA




14FE_





PB12







396
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25275



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGCC




14FE_





PB11







397
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25276



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGGC




14FE_





PB10







398
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25277



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGGG




14FE_





PB9







399
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25278



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGCUGACCAUCGACGAGAAAGG




G_
GACUGAAGCUGCUGGG




14FE_





PB8







400
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25279



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCCAUGUUU




11FE_





PB17







401
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25280



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCCAUGUU




11FE_





PB16







402
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25281



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCCAUGU




11FE_





PB15







403
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25282



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCCAUG




11FE_





PB14







404
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25283



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCCAU




11FE_





PB13







405
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25284



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCCA




11FE_





PB12







406
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25285



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGCC




11FE_





PB11







407
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25286



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGGC




11FE_





PB10







408
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25287



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGGG




11FE_





PB9







409
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25288



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUGACCAUCGACGAGAAAGGGAC




G_
UGAAGCUGCUGGG




11FE_





PB8







410
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25289



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCCAUGUUU




9FE_





PB17







411
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25290



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCCAUGUU




9FE_





PB16







412
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25291



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCCAUGU




9FE_





PB15







413
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25292



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCCAUG




9FE_





PB14







414
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25293



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCCAU




9FE_





PB13







415
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25294



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCCA




9FE_





PB12







416
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25295



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGCC




9FE_





PB11







417
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25296



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGGC




9FE_





PB10







418
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25297



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGGG




9FE_





PB9







419
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25298



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACCAUCGACGAGAAAGGGACUG




G_
AAGCUGCUGGG




9FE_





PB8







420
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25299



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCCAUGUUU




6FE_





PB17







421
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25300



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCCAUGUU




6FE_





PB16







422
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25301



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCCAUGU




6FE_





PB15







423
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25302



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCCAUG




6FE_





PB14







424
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25303



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCCAU




6FE_





PB13







425
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25304



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCCA




6FE_





PB12







426
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25305



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGCC




6FE_





PB11







427
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25306



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGGC




6FE_





PB10







428
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25307



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGGG




6FE_





PB9







429
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25308



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAUCGACGAGAAAGGGACUGAAG




G_
CUGCUGGG




6FE_





PB8







430
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25309



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCCAUGUUU




5FE_





PB17







431
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25310



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCCAUGUU




5FE_





PB16







432
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25311



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCCAUGU




5FE_





PB15







433
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25312



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCCAUG




5FE_





PB14







434
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25313



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCCAU




5FE_





PB13







435
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25314



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCCA




5FE_





PB12







436
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25315



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGCC




5FE_





PB11







437
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25316



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGGC




5FE_





PB10







438
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25317



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGGG




5FE_





PB9







439
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25318



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCGACGAGAAAGGGACUGAAGC




G_
UGCUGGG




5FE_





PB8







440
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25319



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCCAUGUUU




3FE_





PB17







441
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25320



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCCAUGUU




3FE_





PB16







442
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25321



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCCAUGU




3FE_





PB15







443
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25322



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCCAUG




3FE_





PB14







444
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25323



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCCAU




3FE_





PB13







445
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25324



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCCA




3FE_





PB12







446
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25325



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGCC




3FE_





PB11







447
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25326



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGGC




3FE_





PB10







448
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25327



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGGG




3FE_





PB9







449
A1AT_
UAAAAACAUGGCCCCAGCAGCUUGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25328



Nme2_ED15+_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGACGAGAAAGGGACUGAAGCUG




G_
CUGGG




3FE_





PB8







450
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25329



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCACGGCCU




ED15-_30FE_





PB17







451
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25330



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCACGGCC




ED15-_30FE_





PB16







452
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25331



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCACGGC




ED15-_30FE_





PB15







453
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25332



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCACGG




ED15-_30FE_





PB14







454
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25333



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCACG




ED15-_30FE_





PB13







455
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25334



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCAC




ED15-_30FE_





PB12







456
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25335



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGCA




ED15-_30FE_





PB11







457
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25336



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUGC




ED15-_30FE_





PB10







458
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25337



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAUG




ED15-_30FE_





PB9







459
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25338



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




SpRY_
CACAGCCUUAU




ED15-_30FE_





PB8







460
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25339



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCACGGCCU




ED15-_25FE_





PB17







461
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25340



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCACGGCC




ED15-_25FE_





PB16







462
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25341



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCACGGC




ED15-_25FE_





PB15







463
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25342



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCACGG




ED15-_25FE_





PB14







464
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25343



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCACG




ED15-_25FE_





PB13







465
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25344



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCAC




ED15-_25FE_





PB12







466
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25345



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGCA




ED15-_25FE_





PB11







467
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25346



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUGC




ED15-_25FE_





PB10







468
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25347



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAUG




ED15-_25FE_





PB9







469
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25348



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




SpRY_
CCUUAU




ED15-_25FE_





PB8







470
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25349



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCACGGCCU




ED15-_20FE_





PB17







471
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25350



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCACGGCC




ED15-_20FE_





PB16







472
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25351



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCACGGC




ED15-_20FE_





PB15







473
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25352



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCACGG




ED15-_20FE_





PB14







474
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25353



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCACG




ED15-_20FE_





PB13







475
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25354



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCAC




ED15-_20FE_





PB12







476
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25355



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGCA




ED15-_20FE_





PB11







477
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25356



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UGC




ED15-_20FE_





PB10







478
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25357



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
UG




ED15-_20FE_





PB9







479
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25358



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




SpRY_
U




ED15-_20FE_





PB8







480
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25359



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_
GCCU




ED15-_14FE_





PB17







481
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25360



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_
GCC




ED15-_14FE_





PB16







482
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25361



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_
GC




ED15-_14FE_





PB15







483
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25362



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_
G




ED15-_14FE_





PB14







484
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25363



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_





ED15-_14FE_





PB13







485
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25364



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




SpRY_





ED15-_14FE_





PB12







486
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25365



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




SpRY_





ED15-_14FE_





PB11







487
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25366



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




SpRY_





ED15-_14FE_





PB10







488
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25367



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUG




SpRY_





ED15-_14FE_





PB9







489
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25368



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAU




SpRY_





ED15-_14FE_





PB8







490
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25369



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




SpRY_
U




ED15-_11FE_





PB17







491
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25370



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




SpRY_





ED15-_11FE_





PB16







492
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25371



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




SpRY_





ED15-_11FE_





PB15







493
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25372



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




SpRY_





ED15-_11FE_





PB14







494
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25373



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_





ED15-_11FE_





PB13







495
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25374



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




SpRY_





ED15-_11FE_





PB12







496
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25375



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




SpRY_





ED15-_11FE_





PB11







497
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25376



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




SpRY_





ED15-_11FE_





PB10







498
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25377



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUG




SpRY_





ED15-_11FE_





PB9







499
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25378



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAU




SpRY_





ED15-_11FE_





PB8







500
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25379



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




SpRY_





ED15-_9FE_





PB17







501
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25380



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




SpRY_





ED15-_9FE_





PB16







502
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25381



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




SpRY_





ED15-_9FE_





PB15







503
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25382



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




SpRY_





ED15-_9FE_





PB14







504
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25383



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_





ED15-_9FE_





PB13







505
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25384



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




SpRY_





ED15-_9FE_





PB12







506
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25385



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




SpRY_





ED15-_9FE_





PB11







507
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25386



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




SpRY_





ED15-_9FE_





PB10







508
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25387



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUG




SpRY_





ED15-_9FE_





PB9







509
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25388



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAU




SpRY_





ED15-_9FE_





PB8







510
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25389



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




SpRY_





ED15-_7FE_





PB17







511
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25390



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




SpRY_





ED15-_7FE_





PB16







512
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25391



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




SpRY_





ED15-_7FE_





PB15







513
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25392



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




SpRY_





ED15-_7FE_





PB14







514
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25393



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_





ED15-_7FE_





PB13







515
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25394



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




SpRY_





ED15-_7FE_





PB12







516
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25395



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




SpRY_





ED15-_7FE_





PB11







517
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25396



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




SpRY_





ED15-_7FE_





PB10







518
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25397



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUG




SpRY_





ED15-_7FE_





PB9







519
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25398



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAU




SpRY_





ED15-_7FE_





PB8







520
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25399



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




SpRY_





ED15-_5FE_





PB17







521
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25400



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




SpRY_





ED15-_5FE_





PB16







522
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25401



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




SpRY_





ED15-_5FE_





PB15







523
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25402



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




SpRY_





ED15-_5FE_





PB14







524
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25403



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_





ED15-_5FE_





PB13







525
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25404



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




SpRY_





ED15-_5FE_





PB12







526
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25405



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




SpRY_





ED15-_5FE_





PB11







527
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25406



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




SpRY_





ED15-_5FE_





PB10







528
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25407



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUG




SpRY_





ED15-_5FE_





PB9







529
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25408



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAU




SpRY_





ED15-_5FE_





PB8







530
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25409



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




SpRY_





ED15-_3FE_





PB17







531
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25410



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




SpRY_





ED15-_3FE_





PB16







532
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25411



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




SpRY_





ED15-_3FE_





PB15







533
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25412



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




SpRY_





ED15-_3FE_





PB14







534
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25413



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




SpRY_





ED15-_3FE_





PB13







535
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25414



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




SpRY_





ED15-_3FE_





PB12







536
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25415



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




SpRY_





ED15-_





3FE_





PB11







537
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25416



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGC




SpRY_





ED15-_3FE_





PB10







538
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25417



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUG




SpRY_





ED15-_3FE_





PB9







539
A1AT_
AGGCCGUGCAUAAGGCUGUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25418



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAU




SpRY_





ED15-_3FE_





PB8







540
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25419



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




PB17







541
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25420



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




PB16







542
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25421



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




PB15







543
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25422



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




PB14







544
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25423



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




PB13







545
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25424



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




PB12







546
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25425



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




PB11







547
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25426



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




PB10







548
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25427



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUG




PB9







549
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25428



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUACAUGGCCCCAGCAGCUUCAGU




30FE_
CCCUUUCUCGUCGAUGGUCAGCACAGCCUUAU




PB8







550
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25429



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




PB17







551
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25430



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




PB16







552
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25431



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




PB15







553
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25432



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




PB14







554
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25433



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




PB13







555
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25434



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




PB12







556
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25435



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGCA




PB11







557
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25436



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUGC




PB10







558
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25437



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAUG




PB9







559
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25438



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUGCCCCAGCAGCUUCAGUCCCUU




25FE_
UCUCGUCGAUGGUCAGCACAGCCUUAU




PB8







560
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25439



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCACGGCCU




PB17







561
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25440



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCACGGCC




PB16







562
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25441



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCACGGC




PB15







563
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25442



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCACGG




PB14







564
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25443



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCACG




PB13







565
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25444



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCAC




PB12







566
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25445



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGCA




PB11







567
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25446



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUGC




PB10







568
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25447



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAUG




PB9







569
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25448



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGCAGCUUCAGUCCCUUUCUCG




20FE_
UCGAUGGUCAGCACAGCCUUAU




PB8







570
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25449



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCACGGCCU




PB17







571
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25450



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCACGGCC




PB16







572
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25451



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCACGGC




PB15







573
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25452



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCACGG




PB14







574
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25453



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCACG




PB13







575
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25454



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCAC




PB12







576
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25455



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUGCA




PB11







577
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25456



Nme2_ED_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




15-_G_
GUCAGCACAGCCUUAUGC




14FE_





PB10







578
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25457



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAUG




PB9







579
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25458



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUCAGUCCCUUUCUCGUCGAUG




14FE_
GUCAGCACAGCCUUAU




PB8







580
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25459



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCACGGCCU




PB17







581
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25460



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCACGGCC




PB16







582
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25461



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCACGGC




PB15







583
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25462



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCACGG




PB14







584
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25463



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCACG




PB13







585
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25464



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCAC




PB12







586
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25465



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGCA




PB11







587
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25466



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUGC




PB10







588
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25467



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAUG




PB9







589
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25468



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUAGUCCCUUUCUCGUCGAUGGUC




11FE_
AGCACAGCCUUAU




PB8







590
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25469



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGCACGGCCU




PB17







591
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25470



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGCACGGCC




PB16







592
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25471



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGCACGGC




PB15







593
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25472



Nme2_ED_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




15-_G_
CACAGCCUUAUGCACGG




9FE_





PB14







594
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25473



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGCACG




PB13







595
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25474



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGCAC




PB12







596
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25475



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGCA




PB11







597
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25476



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUGC




PB10







598
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25477



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAUG




PB9







599
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25478



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCCCUUUCUCGUCGAUGGUCAG




9FE_
CACAGCCUUAU




PB8







600
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25479



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCACGGCCU




PB17







601
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25480



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCACGGCC




PB16







602
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25481



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCACGGC




PB15







603
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25482



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCACGG




PB14







604
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25483



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCACG




PB13







605
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25484



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCAC




PB12







606
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25485



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGCA




PB11







607
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25486



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUGC




PB10







608
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25487



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAUG




PB9







609
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25488



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUCCUUUCUCGUCGAUGGUCAGCA




7FE_
CAGCCUUAU




PB8







610
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25489



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCACGGCCU




PB17







611
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25490



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCACGGCC




PB16







612
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25491



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCACGGC




PB15







613
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25492



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCACGG




PB14







614
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25493



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCACG




PB13







615
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25494



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCAC




PB12







616
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25495



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGCA




PB11







617
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25496



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUGC




PB10







618
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25497



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAUG




PB9







619
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25498



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUUUCUCGUCGAUGGUCAGCACA




5FE_
GCCUUAU




PB8







620
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25499



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCACGGCCU




PB17







621
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25500



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCACGGCC




PB16







622
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25501



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCACGGC




PB15







623
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25502



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCACGG




PB14







624
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25503



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCACG




PB13







625
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25504



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCAC




PB12







626
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25505



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGCA




PB11







627
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25506



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUGC




PB10







628
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25507



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAUG




PB9







629
A1AT_
UCCAGGCCGUGCAUAAGGCUGUGGUUGUAGCUCCCGAAACGUUGCUACAAUAAGGCCGUCUGAAAAGAUG
25508



Nme2_ED15-_G_
UGCCGCAACGCUCUGCCCCUUAAAGCUUCUGCUUUAAGGGGCAUCGUUUCUCGUCGAUGGUCAGCACAGC




3FE_
CUUAU




PB8







630
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25509



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACGGCCUGG




PB17







631
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25510



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACGGCCUG




PB16







632
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25511



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACGGCCU




PB15







633
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25512



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACGGCC




PB14







634
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25513



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACGGC




PB13







635
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25514



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACGG




PB12







636
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25515



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCACG




PB11







637
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25516



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCAC




PB10







638
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25517



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGCA




PB9







639
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25518



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED17-_30FE_
CACAGCCUUAUGC




PB8







640
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25519



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACGGCCUGG




PB17







641
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25520



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACGGCCUG




PB16







642
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25521



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACGGCCU




PB15







643
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25522



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACGGCC




PB14







644
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25523



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACGGC




PB13







645
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25524



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACGG




PB12







646
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25525



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCACG




PB11







647
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25526



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCAC




PB10







648
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25527



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGCA




PB9







649
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25528



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED17-_25FE_
CCUUAUGC




PB8







650
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25529



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACGGCCUGG




PB17







651
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25530



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACGGCCUG




PB16







652
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25531



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACGGCCU




PB15







653
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25532



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACGGCC




PB14







654
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25533



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACGGC




PB13







655
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25534



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACGG




PB12







656
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25535



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCACG




PB11







657
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25536



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCAC




PB10







658
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25537



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGCA




PB9







659
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25538



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED17-_20FE_
UGC




PB8







660
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25539



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_
GCCUGG




PB17







661
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25540



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_
GCCUG




PB16







662
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25541



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_
GCCU




PB15







663
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25542



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_
GCC




PB14







664
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25543



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_
GC




PB13







665
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25544



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_
G




PB12







666
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25545



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_14FE_





PB11







667
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25546



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED17-_14FE_





PB10







668
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25547



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




ED17-_14FE_





PB9







669
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25548



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




ED17-_14FE_





PB8







670
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25549



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_11FE_
UGG




PB17







671
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25550



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_11FE_
UG




PB16







672
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25551



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_11FE_
U




PB15







673
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25552



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_11FE_





PB14







674
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25553



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




ED17-_11FE_





PB13







675
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25554



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




ED17-_11FE_





PB12







676
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25555



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_11FE_





PB11







677
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25556



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED17-_11FE_





PB10







678
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25557



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




ED17-_11FE_





PB9







679
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25558



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




ED17-_11FE_





PB8







680
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25559



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




ED17-_9FE_
G




PB17







681
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25560



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




ED17-_9FE_





PB16







682
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25561



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




ED17-_9FE_





PB15







683
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25562



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_9FE_





PB14







684
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25563



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




ED17-_9FE_





PB13







685
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25564



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




ED17-_9FE_





PB12







686
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25565



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_9FE_





PB11







687
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25566



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED17-_9FE_





PB10







688
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25567



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




ED17-_9FE_





PB9







689
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25568



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




ED17-_9FE_





PB8







690
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25569



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGG




ED17-_7FE_





PB17







691
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25570



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




ED17-_7FE_





PB16







692
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25571



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




ED17-_7FE_





PB15







693
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25572



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_7FE_





PB14







694
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25573



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




ED17-_7FE_





PB13







695
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25574



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




ED17-_7FE_





PB12







696
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25575



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_7FE_





PB11







697
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25576



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED17-_7FE_





PB10







698
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25577



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




ED17-_7FE_





PB9







699
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25578



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




ED17-_7FE_





PB8







700
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25579



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGG




ED17-_5FE_





PB17







701
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25580



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




ED17-_5FE_





PB16







702
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25581



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




ED17-_5FE_





PB15







703
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25582



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_5FE_





PB14







704
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25583



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




ED17-_5FE_





PB13







705
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25584



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




ED17-_5FE_





PB12







706
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25585



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_5FE_





PB11







707
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25586



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED17-_5FE_





PB10







708
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25587



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




ED17-_5FE_





PB9







709
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25588



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGC




ED17-_5FE_





PB8







710
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25589



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGG




ED17-_3FE_





PB17







711
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25590



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




ED17-_3FE_





PB16







712
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25591



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




ED17-_3FE_





PB15







713
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25592



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




ED17-_3FE_





PB14







714
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25593



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




ED17-_3FE_





PB13







715
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25594



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




ED17-_3FE_





PB12







716
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25595



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




ED17-_3FE_





PB11







717
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25596



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED17-_3FE_





PB10







718
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25597



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




ED17-_3FE_





PB9







719
A1AT_
CCAGGCCGUGCAUAAGGCUGGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25598



SpCas9-_NG_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGC




ED17-_3FE_





PB8







720
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25599



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACGGCCUGGA




PB17







721
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25600



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACGGCCUGG




PB16







722
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25601



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACGGCCUG




PB15







723
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25602



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACGGCCU




PB14







724
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25603



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACGGCC




PB13







725
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25604



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACGGC




PB12







726
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25605



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




ED18-_G_
CACAGCCUUAUGCACGG




30FE_





PB11







727
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25606



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCACG




PB10







728
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25607



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCAC




PB9







729
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25608



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCACAUGGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAG




30FE_
CACAGCCUUAUGCA




PB8







730
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25609



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGGCCUGGA




PB17







731
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25610



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGGCCUGG




PB16







732
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25611



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGGCCUG




PB15







733
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25612



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGGCCU




PB14







734
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25613



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGGCC




PB13







735
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25614



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGGC




PB12







736
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25615



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCACGG




PB11







737
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25616



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




ED18-_G_
CCUUAUGCACG




25FE_





PB10







738
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25617



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCAC




PB9







739
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25618



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCGCCCCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAG




25FE_
CCUUAUGCA




PB8







740
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25619



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACGGCCUGGA




PB17







741
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25620



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACGGCCUGG




PB16







742
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25621



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACGGCCUG




PB15







743
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25622



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACGGCCU




PB14







744
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25623



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACGGCC




PB13







745
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25624



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




ED18-_G_
UGCACGGC




20FE_





PB12







746
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25625



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACGG




PB11







747
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25626



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCACG




PB10







748
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25627



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCAC




PB9







749
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25628



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGCAGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUA




20FE_
UGCA




PB8







750
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25629



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
GCCUGGA




PB17







751
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25630



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
GCCUGG




PB16







752
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25631



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
GCCUG




PB15







753
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25632



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
GCCU




PB14







754
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25633



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
GCC




PB13







755
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25634



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
GC




PB12







756
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25635



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_
G




PB11







757
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25636



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




14FE_





PB10







758
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25637



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




14FE_





PB9







759
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25638



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




14FE_





PB8







760
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25639



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




11FE_
UGGA




PB17







761
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25640



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




11FE_
UGG




PB16







762
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25641



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




11FE_
UG




PB15







763
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25642



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




11FE_
U




PB14







764
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25643



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




11FE_





PB13







765
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25644



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




11FE_





PB12







766
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25645



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




11FE_





PB11







767
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25646



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




11FE_





PB10







768
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25647



SpRY_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




ED18-_G_





11FE_





PB9







769
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25648



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCAGUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




11FE_





PB8







770
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25649



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




9FE_
GA




PB17







771
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25650



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




9FE_
G




PB16







772
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25651



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




9FE_





PB15







773
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25652



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




9FE_





PB14







774
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25653



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




9FE_





PB13







775
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25654



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




9FE_





PB12







776
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25655



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




9FE_





PB11







777
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25656



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




9FE_





PB10







778
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25657



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




9FE_





PB9







779
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25658



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




9FE_





PB8







780
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25659



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGGA




7FE_





PB17







781
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25660



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGG




7FE_





PB16







782
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25661



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




7FE_





PB15







783
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25662



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




7FE_





PB14







784
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25663



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




7FE_





PB13







785
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25664



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




7FE_





PB12







786
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25665



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




7FE_





PB11







787
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25666



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




7FE_





PB10







788
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25667



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




7FE_





PB9







789
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25668



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCCCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




7FE_





PB8







790
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25669



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGGA




5FE_





PB17







791
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25670



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGG




5FE_





PB16







792
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25671



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




5FE_





PB15







793
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25672



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




5FE_





PB14







794
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25673



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




5FE_





PB13







795
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25674



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




5FE_





PB12







796
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25675



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




5FE_





PB11







797
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25676



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




5FE_





PB10







798
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25677



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




5FE_





PB9







799
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25678



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUUUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




5FE_





PB8







800
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25679



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGGA




3FE_





PB17







801
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25680



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUGG




3FE_





PB16







802
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25681



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCUG




3FE_





PB15







803
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25682



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCCU




3FE_





PB14







804
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25683



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGCC




3FE_





PB13







805
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25684



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGGC




3FE_





PB12







806
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25685



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACGG




3FE_





PB11







807
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25686



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCACG




3FE_





PB10







808
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25687



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCAC




3FE_





PB9







809
A1AT_
UCCAGGCCGUGCAUAAGGCUGUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCGUUAUCAA
25688



SpRY_ED18-_G_
CUUGAAAAAGUGGCACCGAGUCGGUGCUCUCGUCGAUGGUCAGCACAGCCUUAUGCA




3FE_





PB8









It should be understood that for all numerical bounds describing some parameter in this application, such as “about,” “at least,” “less than,” and “more than,” the description also necessarily encompasses any range bounded by the recited values. Accordingly, for example, the description “at least 1, 2, 3, 4, or 5” also describes, inter alia, the ranges 1-2, 1-3, 1-4, 1-5, 2-3, 2-4, 2-5, 3-4, 3-5, and 4-5, et cetera.


For all patents, applications, or other reference cited herein, such as non-patent literature and reference sequence information, it should be understood that they are incorporated by reference in their entirety for all purposes as well as for the proposition that is recited. Where any conflict exists between a document incorporated by reference and the present application, this application will control. All information associated with reference gene sequences disclosed in this application, such as GeneIDs or accession numbers (typically referencing NCBI accession numbers), including, for example, genomic loci, genomic sequences, functional annotations, allelic variants, and reference mRNA (including, e.g., exon boundaries or response elements) and protein sequences (such as conserved domain structures), as well as chemical references (e.g., PubChem compound, PubChem substance, or PubChem Bioassay entries, including the annotations therein, such as structures and assays, et cetera), are hereby incorporated by reference in their entirety.


Headings used in this application are for convenience only and do not affect the interpretation of this application.










LENGTHY TABLES




The patent application contains a lengthy table section. A copy of the table is available in electronic form from the USPTO web site (). An electronic copy of the table will also be available from the USPTO upon request and payment of the fee set forth in 37 CFR 1.19(b)(3).





Claims
  • 1. A template RNA comprising, from 5′ to 3′: a) a gRNA spacer that is complementary to a first portion of the human SERPINA1 gene, wherein the gRNA spacer comprises an RNA sequence according to SEQ ID NO: 20,623;b) a gRNA scaffold that binds a SpyCas9-SpRY domain;c) a heterologous object sequence comprising a mutation region to correct a mutation in a second portion of the human SERPINA1 gene; andd) a primer binding site (PBS) sequence comprising at least 3 bases with 100% identity to a third portion of the human SERPINA1 gene.
  • 2. The template RNA of claim 1, wherein the mutation to be corrected in the human SERPINA1 gene is E342K.
  • 3. The template RNA of claim 1, wherein the gRNA spacer has a length of 20 nucleotides.
  • 4. The template RNA of claim 1, wherein the heterologous object sequence has a length of 6-16 nucleotides.
  • 5. The template RNA of claim 1, wherein the heterologous object sequence has a length of 6 nucleotides.
  • 6. The template RNA of claim 1, wherein the heterologous object sequence comprises, from 5′ to 3′, a post-edit homology region, a mutation region, and a pre-edit homology region.
  • 7. The template RNA of claim 1, wherein the heterologous object sequence has an RNA sequence of TTTCTC.
  • 8. The template RNA of claim 1, wherein the PBS sequence has a length of 8-12 nucleotides.
  • 9. The template RNA of claim 1, wherein the PBS sequence has a length of 10 nucleotides.
  • 10. The template RNA of claim 1, wherein the PBS sequence has an RNA sequence according to SEQ ID NO: 21433.
  • 11. The template RNA of claim 1, wherein the gRNA scaffold comprises an RNA sequence having at least 90% identity to SEQ ID NO: 20427.
  • 12. The template RNA of claim 1, wherein the gRNA scaffold comprises an RNA sequence according to SEQ ID NO: 20427.
  • 13. The template RNA of claim 1, which comprises an RNA sequence having at least 90% identity to SEQ ID NO: 24956.
  • 14. The template RNA of claim 1, which comprises an RNA sequence according to SEQ ID NO: 24956.
  • 15. The template RNA of claim 1, which comprises one or more chemically modified nucleotides.
  • 16. The template RNA of claim 15, which comprises the RNA sequence and chemical modifications set out in SEQ ID NO: 24146.
  • 17. A gene modifying system comprising: a template RNA of claim 1, anda gene modifying polypeptide, or a nucleic acid encoding the gene modifying polypeptide.
  • 18. The gene modifying system of claim 17, which comprises the nucleic acid encoding the gene modifying polypeptide, wherein the nucleic acid comprises RNA.
  • 19. The gene modifying system of claim 17, wherein the gene modifying polypeptide comprises: a reverse transcriptase (RT) domain;a Cas domain; anda linker disposed between the RT domain and the Cas domain.
  • 20. The gene modifying system of claim 19, wherein the Cas domain is a SpyCas9-SpRY domain.
  • 21. The gene modifying system of claim 19, wherein the RT domain is an RT domain from a murine leukemia virus (MMLV), a porcine endogenous retrovirus (PERV); Avian reticuloendotheliosis virus (AVIRE), a feline leukemia virus (FLV), simian foamy virus (SFV) (e.g., SFV3L), bovine leukemia virus (BLV), Mason-Pfizer monkey virus (MPMV), human foamy virus (HFV), or bovine foamy/syncytial virus (BFV/BSV).
  • 22. A pharmaceutical composition, comprising the gene modifying system of claim 17 and a pharmaceutically acceptable excipient or carrier.
  • 23. The pharmaceutical composition of claim 22, wherein the pharmaceutically acceptable excipient or carrier is selected from the group consisting of a plasmid vector, a viral vector, a vesicle, and a lipid nanoparticle.
  • 24. A method of making the template RNA of claim 1, the method comprising synthesizing the template RNA by in vitro transcription, solid-phase synthesis, or by introducing a DNA encoding the template RNA into a host cell under conditions that allow for production of the template RNA.
  • 25. A method for modifying a target site in the human SERPINA1 gene in a cell, the method comprising contacting the cell with the gene modifying system of claim 17, or DNA encoding the same, thereby modifying the target site in the human SERPINA1 gene in a cell.
  • 26. A method for treating a subject having a disease or condition associated with a mutation in the human SERPINA1 gene, the method comprising administering to the subject the gene modifying system of claim 17, or DNA encoding the same, thereby treating the subject having a disease or condition associated with a mutation in the human SERPINA1 gene.
  • 27. A template RNA comprising, from 5′ to 3′: (i) a gRNA spacer that is complementary to a first portion of the human SERPINA1 gene, wherein the gRNA spacer has a sequence comprising the core nucleotides of a gRNA spacer sequence of Table 1, or wherein the gRNA spacer has a sequence of a spacer chosen from Tables 6A, 6B, X2, X3, X3a, X5, or XX, wherein the gRNA spacer has a sequence other than SEQ ID NO: 20,623;(ii) a gRNA scaffold that binds a gene modifying polypeptide,(iii) a heterologous object sequence comprising a mutation region to correct a mutation in a second portion of the human SERPINA1 gene, and(iv) a primer binding site (PBS) sequence comprising at least 5, 6, 7, or 8 bases with 100% identity to a third portion of the human SERPINA1 gene.
  • 28. A gene modifying system comprising: a template RNA of claim 27, anda gene modifying polypeptide, or a nucleic acid encoding the gene modifying polypeptide.
  • 29. A method for modifying a target site in the human SERPINA1 gene in a cell, the method comprising contacting the cell with the gene modifying system of claim 28, or DNA encoding the same, thereby modifying the target site in the human SERPINA1 gene in a cell.
  • 30. A method for treating a subject having a disease or condition associated with a mutation in the human SERPINA1 gene, the method comprising administering to the subject the gene modifying system of claim 28, or DNA encoding the same, thereby treating the subject having a disease or condition associated with a mutation in the human SERPINA1 gene.
Provisional Applications (3)
Number Date Country
63303905 Jan 2022 US
63253087 Oct 2021 US
63241970 Sep 2021 US
Continuations (1)
Number Date Country
Parent PCT/US2022/076073 Sep 2022 US
Child 18469344 US