Compositions and Methods Relating to Alzheimer's Disease

II. REFERENCE TO THE SEQUENCE LISTING

The Sequence Listing submitted 12 Oct. 2021 as a text file named “20_940_WO_Sequence_Listing”, created on 12 Oct. 2021 and having a size of 581 kilobytes is hereby incorporated by reference pursuant to 37 C.F.R. § 1.52(e)(5).

III. BACKGROUND

Alzheimer's disease (AD) is sixth leading cause of death in the US and the most common cause of dementia in aging. With a rapidly growing aging population the number of AD cases is growing fast and projected to rise drastically over the next three decades. Today, more than 5 million people are living with AD in the United States alone, and by 2050, this number is projected to reach 14 million. Therefore, AD poses a huge economic burden on society placing overwhelming strain on the healthcare system. In 2020, the cost of AD to the US was $301 billion, including $206 billion in Medicare and Medicaid payments, while the caregivers provided $244 billions worth of care (Alzheimer's Association, Alzheimer's Impact Movement: Factsheet 2020). These trends will only worsen with time because there are no therapies to halt or prevent AD (i.e., projected to cost more than $1.1 trillion annually by 2050). Despite all the research effort, money and commitment, clinical trials to identify disease modifying therapies (DMT) for AD have repeatedly failed. To date, there is no cure and no DMT for AD and there are no methods to delay the onset and/or progression of the disease. Most available treatments are palliative and aimed at relieving symptom (Sharma K. (2019) Mol Med Rep. 20:1479-1487; Olivares D, et al. (2012) Curr Alzheimer Res. 9:746-758).

Late onset AD (LOAD) is a heterogenous disease with various genetic etiologies (Lo M T, et al. (2019). Neurobiol Aging. 84:243 e1-243.e9; Nacmias B, et al. (2018) J Alzheimers Dis. 62:903-911). A major reason for the failure to identify an effective treatment is likely the inaccurate consideration of LOAD as a homogeneous disease. In this respect, increasing evidence demonstrate the heterogeneity in the underlying pathophysiologic processes of LOAD and show variability in the genetic risk and molecular profiles amongst AD patients (Reitz C. (2016) Ann Transl Med. 4:107; Chiba-Falek O, et al. (2017) Expert Rev Precis Med Drug Dev. 2:47-55). Thus, AD remains an unmet medical need underscoring the urgent need for a paradigm shift in AD clinical research.

Accordingly, any advancement in LOAD therapy will require the development and validation of new therapeutic targets including those tailored to sub-groups of patients with specific risk factors. Thus, to date many investigators and funding bodies recognize the need to shift the focus to new potential culprits including candidate gene-targets such as LOAD risk genes and rare mutations (Guerreiro R, et al. (2013) Neurobiol Aging. 34:2890 e1-5). Consistently, recently, alternative targets such as APOE have emerged as potential promising targets for LOAD treatment (Huynh T V, et al. (2017) Neuron. 96:1013-1023 e4; Brody D L, et al. (2008) Annu Rev Neurosci 31:175-193; Kim J, et al. (2012) J Exp Med. 209:2149-2156).

Accordingly, there is a need for a disease modifying therapy for Alzheimer's disease, particularly LOAD.

IV. BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows the effect of APOE genotypes on APOE-mRNA levels. FIG. 1A shows a schematic model describing the mechanisms that lead to increased ApoE activity and by that mediate the pathogenic effect of APOE e4 and APOE e3 (differ in amino acid at position 112 Arg and Cys, respectively) on LOAD. FIG. 1B shows a diagram of the different technologies to target ApoE, including antisense oligonucleotide (ASO), monoclonal antibody (mAbs), and CRISPR/Cas9 gene editing technologies. The fold levels of human APOE mRNA were assayed using qRT-PCR in temporal tissues (FIG. 1C) and in occipital tissues (FIG. 1D). FIG. 1E shows the level of human APOE-mRNA in whole brain tissues from humanized mice assayed by qRT-PCR.

FIG. 2 shows a schematic representation of APOE gene. The APOE gene is located at chromosome 19q13.2. The SNP rs429358 changes amino acid in position 112 and defines APOE e4 allele. The SNP rs7412 changes amino acid in position 158 and defines the APOE e2 allele. The CpG island in exon 4 is highlighted. DMRI and DMR2 regions are defined by two CGIs, which are marked in a yellow box. Exons 1-4 are designated in boxes. The translated exons are highlighted in dark blue. 5′-UTR and 3′-UTR of the gene are highlighted in light blue.

FIG. 3 shows the DNA-methylation profile of the APOE LD region in FANS-sorted neuronal and non-neuronal nuclei. FIG. 3A shows a map of MethylEPIC array probes in the chr19: 45,393,000-45,424,000; hg19. The red circles represent probes with >0.5 methylation levels while the blue circles represent probes with <0.5 methylation levels. The APOE promoter region is hypomethylated and is an excellent target region for enhancement of DNA-methylation. FIG. 3B shows that probes showed significant differences in methylation levels between NeuN⁺ (n=16), NeuN⁻ sorted nuclei (n=16), or LOAD (n=8) vs. Normal (n=8). Solid bars represent neuronal population while the hatched bars represent the non-neuronal population. The accompanying table summarizes the p-values for each of the significant probes.

FIG. 4 show the structure of human APOE gene and the design of spCas9 gRNAs to target the promoter region of the gene. FIG. 4 shows the genomic organization of the gene including the two SNPs in exon 4 and the gRNA targeting of the promoter region of the gene. The 5′-UTR and 3′-UTR of the gene are also shown.

FIG. 5 shows the schematic representation of lentiviral vector system carrying DNMT3A to target the promoter and exon 4 regions of APOE gene. The 5′-LTR and the 3′-LTR represent long terminal repeats. Phi represents the packaging signal of the vector. RRE represents the rev responsive element responsible for binding REV protein of the virus. The Sp1 responsive element inclusion (Ortiniski et al. (2017); Kantor et al. (2018)) demonstrated high production yield. The hU6 promoter drives expression of the gRNA and the EFS-NC promoter drives the expression of dCAS9 (to target promoter of APOE) or dVRER to target SNP (112) at the exon 4 region. The p2A signal separates the effector molecule from GFP/Puro reporters. WPRE is the Woodchuck Hepatitis Virus (WHP) Post-Transcriptional Regulatory Element (WPRE), which is a DNA sequence that when transcribed creates a tertiary structure enhancing expression. The arrow pointing to the promoter region highlights the binding of the dCas9-DNMT3A-gRNA to the promoter region or the SNP region that results in the DNA methylation (red lollipops) and downregulation of gene expression (represented with the red cross sign).

FIG. 6 shows the targeting of the promoter region of APOE with gRNA-dCas9-DNMT3A lentiviral vector system. Human hepatocytes HEPG2 cell (having APOEe3/3 genotype) were stably transduced with lentiviral vector carrying 4 different gRNA paired with dCas9-DNMT3A or dCAS9-DNMT3A null vectors. In FIG. 6, the levels of the mRNA and protein downregulation were compared to untransduced naïve HEPG2 cells. The vectors delivering the active version of DNMT3A represented in white bars while the null mutants are shown in black bars. The experiments were repeated three time and the SD bars are highlighted. FIG. 6A shows the levels of RNA knockdown following the transduction with a lentiviral vector as assessed by real-time PCR. gRNA1 showed the most robust reduction in APOE-mRNA. FIG. 6B shows the levels of protein knockdown following the transduction with a lentiviral vector as assessed by western blot. The effects on the protein levels were comparable with the effects on the mRNA shown in FIG. 6A, demonstrating the most robust decrease in protein levels was driven by gRNA1. In FIG. 6, gRNA1 was gacagggggagccctataat (SEQ ID NO:25), gRNA3 was actgggatgtaagccatagc (SEQ ID NO:27), and gRNA4 was gttggagcttagaatgtgaa (SEQ ID NO:28).

FIG. 7 shows the structure of humanAPOE gene and VRER gRNAs design relative to the spCas9 gRNAs positions targeting the promoter region of the gene. Genomic organization of the gene outlined in the lower panel highlighting the 2 SNPs within exon 4. gRNA targeting promoter region of the gene are outlined. spCas9 gRNAs (in green) and VRER gRNAs (in yellow) positions. The 5′-UTR and the 3′-UTR of the gene are indicated in boxes. Structure of a human APOE gene and VRER vs spCas9 gRNAs locations are shown.

FIG. 8A-FIG. 8B show the validation of VRER system using GFP-reporter cells. A GFP-reporter 293T cell line was created by stable transduction using lentiviral vector. GFP was subjected to site-directed mutagenesis to change the PAM motif for VRER enzyme NGCG to GGG, which is recognized by SpCas9. The cells identified as 1003GFP⁻ are generated to include this modification. The target cells were transduced with SpCas9-gRNA-to-GFP vector VRER-gRNA-to-GFP vector to assess the specificity and efficacy of the corresponding enzymes. The gRNA sequence selected for targeting is highlighted. The cells identified as 201A GFP cells (FIG. 8A) contained the “naïve” GFP sequence, while 1003 GFP cells (FIG. 8B) were introduced with point-substitution (as above) without changing amino acid residues. A score of 5+ highlights the high efficiency of the GFP cleavage, while a score of 5− highlights incapacity of the enzyme to digest DNA. The specificity of VRER was found to be comparable to that of Cas9 while the efficacy was demonstrated to be significantly lower.

FIG. 9 shows the effect of targeting the promoter region of APOE with a gRNA-dVRER-DNMT3A lentiviral vector system. Human hepatocytes HEPG2 cells were stably transduced with lentiviral vector carrying 4 different gRNA paired with dVRER-DNMT3A or dVRER-DNMT3A null vectors. FIG. 9 shows the level of RNA knockdown following the transduction using real-time PCR. The levels of the mRNA downregulation was compared to untransduced, naïve HEPG2 cells. The vectors delivering the active version of DNMT3A are represented in white bars while the null mutants are represented in black bars. The experiments were repeated three times and the SD bars are highlighted.

FIG. 10A-FIG. 10F shows the differentiation and characterization of hiPSC-derived neurons. FIG. 10A shows the timeline for neuronal differentiation. FIG. 10B shows representative immunocytochemistry of hiPSC-derived neurons. FIG. 10C shows the FACS-analysis showing co-expression of TUBB3 and VachT (36.4%) while FIG. 10D shows the absence of GFAP signal. FIG. 10E shows the relative expression levels of the neuronal-specific markers (TUBB3 and CHAT) and the astrocyte specific marker (GFAP). FIG. 10F shows APOE-mRNA expression in isogenic APOE 3/3 and 4/4 hiPSC-derived neurons. APOE-mRNA 3/3>4/4 consistent with the observation in human brain, which demonstrated the suitability of the system for drug discovery.

FIG. 11A-FIG. 11C show expression levels and immunohistochemical staining of isogenic APOE-hiPSC. FIG. 11A shows the fold levels of human APOE mRNA assayed by qRT-PCR using TaqMan assay. FIG. 11B (APOE 3/3) and FIG. 11C (APOE 4/4) show hiPSC shows cells stained with pluripotency markers OCT 4 and NANOG. (FROM GRANT)

FIG. 12A-FIG. 12M show the nuclear envelope markers in isogenic APOE 3/3 and 4/4 hiPSC-derived neurons. FIG. 12A shows the immunocytochemistry for lamin B1 in APOE 3/3 hiPSC-derived neurons while FIG. 12B shows lamin B1 staining in APOE 4/4 hiPSC-derived neurons. As demonstrated in FIG. 12C, the quantification of the nuclear envelope circularity showed loss circularity in the APOE 4/4 hiPSC-derived neurons vs. the APOE 3/3 hiPSC-derived neurons before heat treatment while FIG. 12D shows the same comparison after heat treatment. FIG. 12E shows the immunocytochemistry for lamin AC in APOE 3/3 hiPSC-derived neurons while FIG. 12F shows lamin B1 staining in APOE 4/4 hiPSC-derived neurons. FIG. 12G shows the proportion of cells with abnormal nuclear morphology in the APOE 4/4 hiPSC-derived neurons vs. the APOE 3/3 hiPSC-derived neurons before heat treatment while FIG. 12H shows the same comparison after heat treatment. As shown in FIG. 12I, osmotic stress showed an increased sensitivity of the nuclear envelope in the APOE 4/4 neurons compared to the APOE 3/3. FIG. 13J shows the decrease in global 5-mC % in APOE 4/4 hiPSC-derived neurons as compared to APOE 3/3 hiPSC-derived neurons. FIG. 12K and FIG. 12L shows the nuclear leakage as assessed by a dextran assay using 155 kDa fluorescently-label molecule APOE 3/3 hiPSC-derived neurons and 4/4 hiPSC-derived neurons, respectively. FIG. 12M shows the percentage of leaky nuclei for both APOE 3/3 and APO 4/4 hiPSC-derived neurons.

FIG. 13A-FIG. 13E shows the methylation profile of the APOE LD region in isogenic APOE hiPSC-derived neurons. FIG. 13A shows a map of MethylEPIC array probes in chromosome 19 from 45,393,000-45,424,000 (hg19). Those probes with >0.5 methylation levels are highlighted in red. Those probes with <0.5 methylation levels are highlighted in blue. Significant differences in methylation between the APOE neuronal lines are shown using asterisks as follows: black asterisk (>0.1) and red asterisk (>0.2). Because the APOE promoter region was hypomethylated, it became an excellent target region for enhancement of DNA-methylation. FIG. 13B shows a schematic representation of the 27 CpG islands for pyrosequencing in the APOE region, i.e., chromosome 19 from 45,411,858-45,412,079 (hg19). FIG. 13C shows those probes that had significant differences in DNA-methylation levels between isogenic APOE hiPSC-derived neurons. FIG. 13D shows the methylation level (%) of the CpG 11-38 that was quantitatively determined in the isogenic hiPSC-derived neurons using pyrosequencing. FIG. 13E shows a comparison of the methylation level (%) of CpG 11-38 between hiPSC-derived neurons and NeuN⁺ FANS-sorted nuclei using pyrosequencing. Here, the DNA-methylation profiles of the hiPSC-derived neurons were comparable to those observed for the human brain sorted neuronal nuclei (indicating that the hiPSC-derived neuronal system was suitable for drug discovery studies aiming at DNA-methylation editing).

FIG. 14A-FIG. 14D show the AD-related phenotypes in isogenic APOE 3/3 and 4/4 hiPSC-derived neurons. FIG. 14A shows the ratio of extracellular A042:AD40 secreted from APO 3/3 and APOE 4/4 neurons measured by ELISA. FIG. 14B shows the total tau levels measured by ELISA. FIG. 14C shows the neurite outgrowth evaluated using TUBB3 immunostaining in APOE 3/3 hiPSC-derived neurons. and FIG. 14D shows the neurite outgrowth evaluated using TUBB3 immunostaining in APOE 4/4 hiPSC-derived neurons.

FIG. 15A-15B shows methylation in the target promoter region of APOE and the design of gRNA for targeting. FIG. 15A shows the genome browser view of a map of the targeted region using UCSC genome browser viewer. The black bars in the upper portion of the panel shows the positions of (i) the target region, (ii) the designed gRNAs, and (iii) MethylEpic probes. The lower panel of FIG. 15A shows the APOE gene structure including the promoter, exon 1, intron 1, and the TSS. FIG. 15B shows the analysis of DNA-methylation within the APOE-promoter target region. Relevant probes were those that overlapped the target region and showed differences in DNA-methylation levels between the isogenic APOE hiPSC-derived neurons.

FIG. 16 shows the targeting of the promoter region of APOE with gRNA-dCas9-DNMT3A lentiviral vector system. hiPSC-derived cholinergic neurons homozygote to the APOE e4 allele (APOE 4/4) were stably transduced with lentiviral vector carrying gRNA3 paired with either a dCas9-DNMT3A vector or a dCAS9-DNMT3A null vector. To assess the level of APOE mRNA knockdown following the transduction, qRT-PCR was used.

FIG. 17 shows the targeting of the promoter region of APOE with gRNA-dCas9-DNMT3A lentiviral vector system. hiPSC-derived cholinergic neurons homozygote to the APOE e4 allele (APOE 4/4) were stably transduced with lentiviral vector carrying gRNAs 1-4 paired with dCas9-DNMT3A or a dCas9-DNMT3A vector with no-gRNA. To assess the level of APOE mRNA knockdown following the transduction, qRT-PCR was used.

FIG. 18 shows the targeting of the promoter region of APOE with gRNA-dCas9-DNMT3A lentiviral vector system. hiPSC-derived cholinergic neurons homozygote to the APOE e3 allele (APOE 3/3) were stably transduced with lentiviral vector carrying gRNAs 1-4 paired with dCas9-DNMT3A compared to dCAS9-DNMT3A vector with no-gRNA. To assess the level of APOE mRNA knockdown following the transduction, qRT-PCR was used.

FIG. 19 shows the targeting exon 4 region of APOE with a gRNA-dVRER-DNMT3A lentiviral vector system. hiPSC-derived cholinergic neurons homozygote to the APOE e4 allele (APOE 4/4) were stably transduced with lentiviral vector carrying a gRNA 2′-paired with dVRER-DNMT3A compared to a dVRER-DNMT3A vector with no-gRNA. Real-time PCR assessed the level of mRNA knockdown following the transduction. A 15% reduction in the level of APOE-mRNA was observed following transduction with the lentiviral vector carrying the gRNA.

FIG. 20 shows the targeting exon 4 region of APOE with a gRNA-dVRER-DNMT3A lentiviral vector system. hiPSC-derived cholinergic neurons homozygote to the APOE e3 allele (APOE 3/3) were stably transduced with lentiviral vector carrying a gRNA 2′-paired with dVRER-DNMT3A compared to a dVRER-DNMT3A vector with no-gRNA. Real-time PCR assessed the level of mRNA knockdown following the transduction. No changes in APOE-mRNA were observed.

FIG. 21A-FIG. 21B show the schematic strategy to silence APOEe4 allele using DNMT3A-DNMT3L enzymes and KRAB repressor as the effector molecules. FIG. 21 shows a schematic representation of the APOE gene including promoter region and exons 1-4. As shown on the right, two lentiviral vector systems were established. The first system carried dCAS9-gRNA-to-promoter. This vector also contained a SunTag epitope that was recognized by single-chain scFv protein. The second system carried dVRER and a gRNA for specific targeting of SNP rs429358 in the exon 4 (on the e4) and DNMT3A-DNMT3L effectors. gRNA introduced with MS2 binding sites allowed the recruitment of KRAB repressor via the MS2-protein (fusion). FIG. 21B shows that following lentiviral vector delivery of dCAS9-gRNA-SunTag binds to the promoter region on both alleles. However, it was inactive on the e3-allele as it lacked the effector molecules. The recruitment of dVRER via specific binding mediated throughout the recognition of the PAM (NGCG) brings the effector molecules in the action. Following interaction between SunTag-scFv DNA on the e4 will be looped out and two the effector molecules, KRAB and DNMT3A-L repress and methylate the promoter of the e4. This SunTag-MS2-bridging system allows specific repression of the e4 allele.

FIG. 22 shows the schematic of a lentiviral vector carrying gRNA-dCas9/dVRER-repressor transgene. the targeting exon 4 region of APOE with a gRNA-dVRER-DNMT3A lentiviral vector system. The vector backbone was optimized by inclosing Sp1 binding sites. dCas9-KRAB/MeCP2/KRAB-MeCP2 fusion was expressed from EFS-NC promoter. Human U6 promoter drove the gRNA expression. The vector carried gRNA to target the regulatory element within exon 4 overlapping the e4-SNP (i.e., specifically target the ApoE4 allele).

FIG. 23A-FIG. 23B show the targeting exon 4 region of APOE with a gRNA-dVRER-DNMT3A lentiviral vector system. FIG. 23A shows that the construct was identical to that of FIG. 5 but for the addition of the repressor to the fused domains of KRAB-MeCP2. FIG. 23B shows the mRNA level in hiPSC-derived cholinergic neurons homozygote to the APOE e4 allele following stable transduction with lentiviral vector carrying a gRNA 2′-paired with dVRER-CRAB MeCp2 or a lentiviral vector carrying a dVRER-KRAB MeCp2 vector with no gRNA. Real-time PCR assessed the levels of mRNA knockdown following the transduction. The vector harboring gRNA2 caused a >50% reduction in the level of APOE mRNA.

V. BRIEF SUMMARY

Disclosed herein is an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA. Disclosed herein is an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is a viral vector comprising a disclosed isolated nucleic acid molecule. Disclosed herein is a viral vector comprising a disclosed isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA. Disclosed herein is a viral vector comprising a disclosed isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is a lentiviral vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA. Disclosed herein is a lentiviral vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is pharmaceutical formulation comprising a disclosed isolated nucleic acid molecule and a pharmaceutically acceptable carrier. Disclosed herein is pharmaceutical formulation comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and a pharmaceutically acceptable carrier. Disclosed herein is pharmaceutical formulation comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA, and a pharmaceutically acceptable carrier. Disclosed herein is pharmaceutical formulation comprising a disclosed vector and a pharmaceutically acceptable carrier. Disclosed herein is pharmaceutical formulation comprising a disclosed lentiviral vector and a pharmaceutically acceptable carrier.

Disclosed herein is a host cell comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA.

Disclosed herein is a host cell comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is a host cell comprising a lentiviral vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA. Disclosed herein is a host cell comprising a lentiviral vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is a host cell comprising plasmid comprising the sequence set forth in any one of SEQ ID NO:21-24, SEQ ID NO:29-36, SEQ ID NO:43-50, SEQ ID NO:53-56, SEQ ID NO:59-61. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:05-SEQ ID NO:14. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:25-SEQ ID NO:28. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:39-SEQ ID NO:42. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:51-SEQ ID NO:52. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:21-SEQ ID NO:24.

Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:29-SEQ ID NO:36. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:43-SEQ ID NO:50. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:53-SEQ ID NO:56. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:59-SEQ ID NO:61.

Disclosed herein is a method of administering precision gene therapy, the method comprising contacting one or more cells with a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA, wherein the fusion protein comprises a Cas endonuclease and a polypeptide having an enzymatic activity, and reducing expression of APOE in one or more cells.

Disclosed herein is a method of administering precision gene therapy, the method comprising contacting one or more cells with a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of APOE in one or more cells.

Disclosed herein is a method of administering precision gene therapy, the method comprising contacting one or more cells with a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of the APOE e4 allele in one or more cells.

Disclosed herein is a method of administering precision gene therapy, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA, wherein the fusion protein comprises a Cas endonuclease and a polypeptide having an enzymatic activity, and reducing expression of the APOE e4 allele.

Disclosed herein is a method of administering precision gene therapy, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of the APOE e4 allele.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein comprising a Cas endonuclease and a polypeptide having an enzymatic activity and (ii) at least one guide RNA, and reducing expression of APOE, thereby reducing the pathological phenotype associated with Alzheimer's disease.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising reducing the pathological phenotype associated with Alzheimer's disease by administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein comprising a Cas endonuclease and a polypeptide having an enzymatic activity and (ii) at least one guide RNA, and reducing expression of APOE.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of APOE, thereby reducing the pathological phenotype associated with Alzheimer's disease.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising reducing the pathological phenotype associated with Alzheimer's disease by administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of APOE.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein comprising a Cas endonuclease and a polypeptide having an enzymatic activity and (ii) at least one guide RNA, and reducing expression of the APOE e4 allele, thereby reducing the pathological phenotype associated with Alzheimer's disease.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of the APOE e4 allele, thereby reducing the pathological phenotype associated with Alzheimer's disease.

Disclosed herein is a method of treating and/or preventing Alzheimer's disease progression in a subject, the method comprising reducing the pathological phenotype associated with Alzheimer's disease by administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of the APOE e4 allele.

Disclosed herein is a method of reducing expression of APOE, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein comprising a Cas endonuclease and a polypeptide having an enzymatic activity and (ii) at least one guide RNA, thereby reducing expression of APOE.

Disclosed herein is a method of reducing expression of APOE, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, thereby reducing expression of APOE.

Disclosed herein is a method of reducing expression of APOE e4, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein comprising a Cas endonuclease and a polypeptide having an enzymatic activity and (ii) at least one guide RNA, thereby reducing expression of the APOE e4 allele.

Disclosed herein is a method of reducing expression of APOE e4, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, thereby reducing expression of the APOE e4 allele.

Disclosed herein is a kit comprising one or more disclosed isolated nucleic acid molecules, disclosed vectors, disclosed lentiviral vectors, disclosed pharmaceutical formulations, disclosed host cells, disclosed guide RNAs, disclosed plasmids, or any combination thereof with or without additional therapeutic agents to treat, prevent, inhibit, and/or ameliorate one or more symptoms or complications associated AD or LOAD.

VI. DETAILED DESCRIPTION

The present disclosure describes formulations, compounded compositions, kits, capsules, containers, and/or methods thereof. It is to be understood that the inventive aspects of which are not limited to specific synthetic methods unless otherwise specified, or to particular reagents unless otherwise specified, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and is not intended to be limiting. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, example methods and materials are now described.

All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited. The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention.

A. Relevant Definitions

Before the present compounds, compositions, articles, systems, devices, and/or methods are disclosed and described, it is to be understood that they are not limited to specific synthetic methods unless otherwise specified, or to particular reagents unless otherwise specified, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and is not intended to be limiting. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, example methods and materials are now described.

This disclosure describes inventive concepts with reference to specific examples. However, the intent is to cover all modifications, equivalents, and alternatives of the inventive concepts that are consistent with this disclosure.

As used in the specification and the appended claims, the singular forms “a”, “an”, and “the” include plural referents unless the context clearly dictates otherwise.

The phrase “consisting essentially of” limits the scope of a claim to the recited components in a composition or the recited steps in a method as well as those that do not materially affect the basic and novel characteristic or characteristics of the claimed composition or claimed method. The phrase “consisting of” excludes any component, step, or element that is not recited in the claim. The phrase “comprising” is synonymous with “including”, “containing”, or “characterized by”, and is inclusive or open-ended. “Comprising” does not exclude additional, unrecited components or steps.

As used herein, when referring to any numerical value, the term “about” means a value falling within a range that is +10% of the stated value.

Ranges can be expressed herein as from “about” one particular value, and/or to “about” another particular value. When such a range is expressed, a further aspect includes from the one particular value and/or to the other particular value. Similarly, when values are expressed as approximations, by use of the antecedent “about,” it will be understood that the particular value forms a further aspect. It will be further understood that the endpoints of each of the ranges are significant both in relation to the other endpoint and independently of the other endpoint. It is also understood that there are a number of values disclosed herein, and that each value is also herein disclosed as “about” that particular value in addition to the value itself. For example, if the value “10” is disclosed, then “about 10” is also disclosed. It is also understood that each unit between two particular units are also disclosed. For example, if 10 and 15 are disclosed, then 11, 12, 13, and 14 are also disclosed.

References in the specification and concluding claims to parts by weight of a particular element or component in a composition denotes the weight relationship between the element or component and any other elements or components in the composition or article for which a part by weight is expressed. Thus, in a compound containing 2 parts by weight component X and 5 parts by weight component Y, X and Y are present at a weight ratio of 2:5, and are present in such ratio regardless of whether additional components are contained in the compound.

As used herein, the terms “optional” or “optionally” means that the subsequently described event or circumstance can or cannot occur, and that the description includes instances where said event or circumstance occurs and instances where it does not. In an aspect, a disclosed method can optionally comprise one or more additional steps, such as, for example, repeating an administering step or altering an administering step.

As used herein, the term “subject” refers to the target of administration, e.g., a human being. The term “subject” also includes domesticated animals (e.g., cats, dogs, etc.), livestock (e.g., cattle, horses, pigs, sheep, goats, etc.), and laboratory animals (e.g., mouse, rabbit, rat, guinea pig, fruit fly, etc.). Thus, the subject of the herein disclosed methods can be a vertebrate, such as a mammal, a fish, a bird, a reptile, or an amphibian. Alternatively, the subject of the herein disclosed methods can be a human, non-human primate, horse, pig, rabbit, dog, sheep, goat, cow, cat, guinea pig, or rodent. The term does not denote a particular age or sex, and thus, adult and child subjects, as well as fetuses, whether male or female, are intended to be covered. In an aspect, a subject can be a human patient. In an aspect, a subject can have Alzheimer's disease (e.g., LOAD), be suspected of having Alzheimer's disease, or be at risk of developing and/or acquiring Alzheimer's disease.

As used herein, the term “diagnosed” means having been subjected to an examination by a person of skill, for example, a physician, and found to have a condition that can be diagnosed or treated by one or more of the disclosed agents, disclosed therapeutic agents, disclosed pharmaceutical formulations, or a combination thereof, or by one or more of the disclosed methods. For example, “diagnosed with Alzheimer's disease or LOAD” means having been subjected to an examination by a person of skill, for example, a physician, and found to have a condition that can be treated by one or more of the disclosed isolated nucleic acid molecules, disclosed viral vectors, disclosed lentiviral vectors, disclosed pharmaceutical formulations, disclosed host cells, disclosed gRNAs, disclosed plasmids, or any combination thereof, or by one or more of the disclosed methods. For example, “suspected of having Alzheimer's disease” can mean having been subjected to an examination by a person of skill, for example, a physician, and found to have a condition that can likely be treated by one or more of the disclosed isolated nucleic acid molecules, disclosed viral vectors, disclosed lentiviral vectors, disclosed pharmaceutical formulations, disclosed host cells, disclosed gRNAs, disclosed plasmids, or any combination thereof, or by one or more of the disclosed methods. In an aspect, an examination can be physical, can involve various tests (e.g., blood tests, genotyping, biopsies, etc.) and assays (e.g., enzymatic assay), or a combination thereof.

A “patient” can refer to a subject that has been diagnosed with or is suspected of having Alzheimer's disease (AD) or late-onset Alzheimer's disease (LOAD). In an aspect, a patient can refer to a subject that has been diagnosed with or is suspected of having AD such as for example, LOAD, and is seeking treatment or receiving treatment for AD or LOAD.

As used herein, the phrase “identified to be in need of treatment for a disorder,” or the like, refers to selection of a subject based upon need for treatment of the disorder. For example, a subject can be identified as having a need for treatment of a disorder (e.g., such as Alzheimer's disease) based upon an earlier diagnosis by a person of skill and thereafter subjected to treatment for the disorder (e.g., AD or LOAD). In an aspect, the identification can be performed by a person different from the person making the diagnosis. In an aspect, the administration can be performed by one who performed the diagnosis.

As used herein, “inhibit,” “inhibiting”, and “inhibition” mean to diminish or decrease an activity, level, response, condition, severity, disease, or other biological parameter. This can include, but is not limited to, the complete ablation of the activity, level, response, condition, severity, disease, or other biological parameter. This can also include, for example, a 10% inhibition or reduction in the activity, level, response, condition, severity, disease, or other biological parameter as compared to the native or control level (e.g., a subject not having Alzheimer's disease). Thus, in an aspect, the inhibition or reduction can be a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or any amount of reduction in between as compared to native or control levels. In an aspect, the inhibition or reduction can be 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100% as compared to native or control levels. In an aspect, the inhibition or reduction can be 0-25%, 25-50%, 50-75%, or 75-100% as compared to native or control levels. In an aspect, a native or control level can be a pre-disease or pre-disorder level.

The words “treat” or “treating” or “treatment” include palliative treatment, that is, treatment designed for the relief of symptoms rather than the curing of the disease, pathological condition, or disorder; preventative treatment, that is, treatment directed to minimizing or partially or completely inhibiting the development of the associated disease, pathological condition, or disorder; and supportive treatment, that is, treatment employed to supplement another specific therapy directed toward the improvement of the associated disease, pathological condition, or disorder (such as Alzheimer's disease). In an aspect, the terms cover any treatment of a subject, including a mammal (e.g., a human), and includes: (i) preventing the undesired physiological change, disease, pathological condition, or disorder from occurring in a subject that can be predisposed to the disease but has not yet been diagnosed as having it; (ii) inhibiting the physiological change, disease, pathological condition, or disorder, i.e., arresting its development; or (iii) relieving the physiological change, disease, pathological condition, or disorder, i.e., causing regression of the disease. For example, in an aspect, treating Alzheimer's disease or LOAD can reduce the severity of an established disease in a subject by 1%-100% as compared to a control (such as, for example, an individual not having AD or LOAD). In an aspect, treating can refer to a 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% reduction in the severity of AD or LOAD. For example, treating Alzheimer's disease can reduce one or more symptoms of AD or LOAD in a subject by 1%-100% as compared to a control (such as, for example, an individual not having AD or LOAD). In an aspect, treating can refer to 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% reduction of one or more symptoms of an established AD (such as LOAD). It is understood that treatment does not necessarily refer to a cure or complete ablation or eradication of AD. However, in an aspect, treatment can refer to a cure or complete ablation or eradication of AD or LOAD.

As used herein, “SunTag” refers to a tag that allows numerous copies of GFP to be recruited to a protein of interest for bright signals. The SunTag can be used for amplification of a fluorescence signal (Tanenbaum M E, et al. (2014) Cell. 159(3):635-646).

As used herein, a “biomarker” refers to a defined characteristic that is measured as an indicator of normal biological processes, pathogenic processes, or response to an exposure of intervention. In an aspect, a biomarker can be diagnostic (i.e., detects or classifies a pathological condition), prognostic (i.e., predicts the probability of disease occurrence or progression), pharmacodynamic/responsive (i.e., identifies a change in response to a therapeutic intervention), predictive (i.e., predicts how an individual or subject might respond to a particular intervention or event). In an aspect, a biomarker can be diagnostic, prognostic, pharmacodynamic/responsive, and/or predictive at the same time. In an aspect, a biomarker can be diagnostic, prognostic, pharmacodynamic/responsive, and/or predictive at different times (e.g., first a biomarker can be diagnostic and then later, the same biomarker can be prognostic, pharmacodynamic/responsive, and/or predictive). A biomarker can be an objective measure that can be linked to a clinical outcome assessment. A biomarker can be used by the skilled person to make a clinical decision based on its context of use.

As used herein, “operably linked” means that expression of a gene is under the control of a promoter with which it is spatially connected. A promoter can be positioned 5′ (upstream) or 3′ (downstream) of a gene under its control. The distance between the promoter and a gene can be approximately the same as the distance between that promoter and the gene it controls in the gene from which the promoter is derived. As is known in the art, variation in this distance can be accommodated without loss of promoter function.

As used herein, the term “prevent” or “preventing” or “prevention” refers to precluding, averting, obviating, forestalling, stopping, or hindering something from happening, especially by advance action. It is understood that where reduce, inhibit, or prevent are used herein, unless specifically indicated otherwise, the use of the other two words is also expressly disclosed. In an aspect, preventing Alzheimer's disease (AD) or LOAD and/or AD or LOAD progression is intended. The words “prevent” and “preventing” and “prevention” also refer to prophylactic or preventative measures for protecting or precluding a subject (e.g., an individual) not having AD or LOAD or an AD or LOAD complication from progressing to that complication. In an aspect, preventing or reducing APOE expression and/or activity is intended.

As used herein, the terms “administering” and “administration” refer to any method of providing one or more of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof to a subject. Such methods are well known to those skilled in the art and include, but are not limited to, the following routes: oral administration, transdermal administration, administration by inhalation, nasal administration, topical administration, in utero administration, intrahepatic administration, intravaginal administration, ophthalmic administration, intraaural administration, otic administration, intracerebral administration, rectal administration, sublingual administration, buccal administration, and parenteral administration, including injectable such as intravenous administration, intra-CSF administration, intra-arterial administration, intramuscular administration, and subcutaneous administration. Administration can also include hepatic intra-arterial administration or administration through the hepatic portal vein (HPV). Administration of a disclosed therapeutic agent, a disclosed pharmaceutical composition, or a combination thereof can comprise administration directly into the CNS (e.g., intraparenchymal, intracerebroventriular, inthrathecal cisternal, intrathecal (lumbar), deep gray matter delivery, convection-enhanced delivery to deep gray matter) or the PNS. Administration can be continuous or intermittent.

In an aspect, a “therapeutic agent” can be a “biologically active agent” or “biologic active agent” or “bioactive agent”, which refers to an agent that is capable of providing a local or systemic biological, physiological, or therapeutic effect in the biological system to which it is applied. For example, the bioactive agent can act to control infection or inflammation, enhance cell growth and tissue regeneration, control tumor growth, act as an analgesic, promote anti-cell attachment, and enhance bone growth, among other functions. Other suitable bioactive agents can include anti-viral agents, vaccines, hormones, antibodies (including active antibody fragments sFv, Fv, and Fab fragments), aptamers, peptide mimetics, functional nucleic acids, therapeutic proteins, peptides, or nucleic acids. Other bioactive agents include prodrugs, which are agents that are not biologically active when administered but, upon administration to a subject are converted to bioactive agents through metabolism or some other mechanism. Additionally, any of the compositions of the invention can contain combinations of two or more bioactive agents. It is understood that a biologically active agent can be used in connection with administration to various subjects, for example, to humans (i.e., medical administration) or to animals (i.e., veterinary administration). As used herein, the recitation of a biologically active agent inherently encompasses the pharmaceutically acceptable salts thereof.

In an aspect, a “therapeutic agent” can be any agent that effects a desired clinical outcome in a subject having AD or LOAD, suspected of having AD or LOAD, and/or likely to develop or acquire AD or LOAD. In an aspect, a disclosed therapeutic agent can be an oligonucleotide therapeutic agent. A disclosed oligonucleotide therapeutic agent can comprise a single-stranded or double-stranded DNA, iRNA, shRNA, siRNA, mRNA, non-coding RNA (ncRNA), an antisense molecule, miRNA, a morpholino, a peptide-nucleic acid (PNA), or an analog or conjugate thereof. In an aspect, a disclosed oligonucleotide therapeutic agent can be an ASO or an RNAi. In an aspect, a disclosed oligonucleotide therapeutic agent can comprise one or more modifications at any position applicable.

In an aspect, a therapeutic agent can be a “drug” or a “vaccine” and means a molecule, group of molecules, complex or substance administered to an organism for diagnostic, therapeutic, preventative medical, or veterinary purposes. This term includes externally and internally administered topical, localized and systemic human and animal pharmaceuticals, treatments, remedies, nutraceuticals, cosmeceuticals, biologicals, devices, diagnostics and contraceptives, including preparations useful in clinical and veterinary screening, prevention, prophylaxis, healing, wellness, detection, imaging, diagnosis, therapy, surgery, monitoring, cosmetics, prosthetics, forensics and the like. This term may also be used in reference to agriceutical, workplace, military, industrial and environmental therapeutics or remedies comprising selected molecules or selected nucleic acid sequences capable of recognizing cellular receptors, membrane receptors, hormone receptors, therapeutic receptors, microbes, viruses or selected targets comprising or capable of contacting plants, animals and/or humans. Examples include but are not limited to a radiosensitizer, the combination of a radiosensitizer and a chemotherapeutic, a steroid, a xanthine, a beta-2-agonist bronchodilator, an anti-inflammatory agent, an analgesic agent, a calcium antagonist, an angiotensin-converting enzyme inhibitors, a beta-blocker, a centrally active alpha-agonist, an alpha-1-antagonist, carbonic anhydrase inhibitors, prostaglandin analogs, a combination of an alpha agonist and a beta blocker, a combination of a carbonic anhydrase inhibitor and a beta blocker, an anticholinergic/antispasmodic agent, a vasopressin analogue, an antiarrhythmic agent, an antiparkinsonian agent, an antiangina/antihypertensive agent, an anticoagulant agent, an antiplatelet agent, a sedative, an ansiolytic agent, a peptidic agent, a biopolymeric agent, an antineoplastic agent, a laxative, an antidiarrheal agent, an antimicrobial agent, an antifungal agent, or a vaccine. In a further aspect, the pharmaceutically active agent can be coumarin, albumin, bromolidine, steroids such as betamethasone, dexamethasone, methylprednisolone, prednisolone, prednisone, triamcinolone, budesonide, hydrocortisone, and pharmaceutically acceptable hydrocortisone derivatives; xanthines such as theophylline and doxophylline; beta-2-agonist bronchodilators such as salbutamol, fenterol, clenbuterol, bambuterol, salmeterol, fenoterol; antiinflammatory agents, including antiasthmatic anti-inflammatory agents, antiarthritis antiinflammatory agents, and non-steroidal antiinflammatory agents, examples of which include but are not limited to sulfides, mesalamine, budesonide, salazopyrin, diclofenac, pharmaceutically acceptable diclofenac salts, nimesulide, naproxene, acetominophen, ibuprofen, ketoprofen and piroxicam; analgesic agents such as salicylates; calcium channel blockers such as nifedipine, amlodipine, and nicardipine; angiotensin-converting enzyme inhibitors such as captopril, benazepril hydrochloride, fosinopril sodium, trandolapril, ramipril, lisinopril, enalapril, quinapril hydrochloride, and moexipril hydrochloride; beta-blockers (i.e., beta adrenergic blocking agents) such as sotalol hydrochloride, timolol maleate, timol hemihydrate, levobunolol hydrochloride, esmolol hydrochloride, carteolol, propanolol hydrochloride, betaxolol hydrochloride, penbutolol sulfate, metoprolol tartrate, metoprolol succinate, acebutolol hydrochloride, atenolol, pindolol, and bisoprolol fumarate; centrally active alpha-2-agonists (i.e., alpha adrenergic receptor agonist) such as clonidine, brimonidine tartrate, and apraclonidine hydrochloride; alpha-1-antagonists such as doxazosin and prazosin; anticholinergic/antispasmodic agents such as dicyclomine hydrochloride, scopolamine hydrobromide, glycopyrrolate, clidinium bromide, flavoxate, and oxybutynin; vasopressin analogues such as vasopressin and desmopressin; prostaglandin analogs such as latanoprost, travoprost, and bimatoprost; cholinergics (i.e., acetylcholine receptor agonists) such as pilocarpine hydrochloride and carbachol; glutamate receptor agonists such as the N-methyl D-aspartate receptor agonist memantine; anti-Vascular endothelial growth factor (VEGF) aptamers such as pegaptanib; anti-VEGF antibodies (including but not limited to anti-VEGF-A antibodies) such as ranibizumab and bevacizumab; carbonic anhydrase inhibitors such as methazolamide, brinzolamide, dorzolamide hydrochloride, and acetazolamide; antiarrhythmic agents such as quinidine, lidocaine, tocainide hydrochloride, mexiletine hydrochloride, digoxin, verapamil hydrochloride, propafenone hydrochloride, flecaimide acetate, procainamide hydrochloride, moricizine hydrochloride, and diisopyramide phosphate; antiparkinsonian agents, such as dopamine, L-Dopa/Carbidopa, selegiline, dihydroergocryptine, pergolide, lisuride, apomorphine, and bromocryptine; antiangina agents and antihypertensive agents such as isosorbide mononitrate, isosorbide dinitrate, propranolol, atenolol and verapamil; anticoagulant and antiplatelet agents such as coumadin, warfarin, acetylsalicylic acid, and ticlopidine; sedatives such as benzodiazapines and barbiturates; ansiolytic agents such as lorazepam, bromazepam, and diazepam; peptidic and biopolymeric agents such as calcitonin, leuprolide and other LHRH agonists, hirudin, cyclosporin, insulin, somatostatin, protirelin, interferon, desmopressin, somatotropin, thymopentin, pidotimod, erythropoietin, interleukins, melatonin, granulocyte/macrophage-CSF, and heparin; antineoplastic agents such as etoposide, etoposide phosphate, cyclophosphamide, methotrexate, 5-fluorouracil, vincristine, doxorubicin, cisplatin, hydroxyurea, leucovorin calcium, tamoxifen, flutamide, asparaginase, altretamine, mitotane, and procarbazine hydrochloride; laxatives such as senna concentrate, casanthranol, bisacodyl, and sodium picosulphate; antidiarrheal agents such as difenoxine hydrochloride, loperamide hydrochloride, furazolidone, diphenoxylate hydrochloride, and microorganisms; vaccines such as bacterial and viral vaccines; antimicrobial agents such as penicillins, cephalosporins, and macrolides, antifungal agents such as imidazolic and triazolic derivatives; and nucleic acids such as DNA sequences encoding for biological proteins, and antisense oligonucleotides. It is understood that a pharmaceutically active agent can be used in connection with administration to various subjects, for example, to humans (i.e., medical administration) or to animals (i.e., veterinary administration). As used herein, the recitation of a pharmaceutically active agent inherently encompasses the pharmaceutically acceptable salts thereof.

“Sequence identity” and “sequence similarity” can be determined by alignment of two peptide or two nucleotide sequences using global or local alignment algorithms. Sequences may then be referred to as “substantially identical” or “essentially similar” when they are optimally aligned. For example, sequence similarity or identity can be determined by searching against databases such as FASTA, BLAST, etc., but hits should be retrieved and aligned pairwise to compare sequence identity. Two proteins or two protein domains, or two nucleic acid sequences can have “substantial sequence identity” if the percentage sequence identity is at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, 99% or more, preferably 90%, 95%, 98%, 99% or more. Such sequences are also referred to as “variants” herein, e.g., other variants of glycogen branching enzymes and amylases. It should be understood that sequence with substantial sequence identity do not necessarily have the same length and may differ in length. For example, sequences that have the same nucleotide sequence but of which one has additional nucleotides on the 3′- and/or 5′-side are 100% identical.

In an aspect, the skilled person can determine an efficacious dose, an efficacious schedule, and an efficacious route of administration for one or more of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof so as to treat or prevent AD or LOAD. In an aspect, the skilled person can also alter, change, or modify an aspect of an administering step to improve efficacy of one or more of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof. In an aspect, the skilled person can determine an efficacious dose, an efficacious schedule, and an efficacious route of administration for any disclosed isolated nucleic acid molecule, disclosed pharmaceutical formulation, disclosed vector, disclosed therapeutic agent, or any combination thereof.

As used herein, “modifying the method” can comprise modifying or changing one or more features or aspects of one or more steps of a disclosed method. For example, in an aspect, a method can be altered by changing the amount of one or more of the disclosed isolated nucleic acid molecules, disclosed viral vectors, disclosed lentiviral vectors, disclosed pharmaceutical formulations, disclosed host cells, disclosed gRNAs, disclosed plasmids, or any combination thereof, or administered to a subject, or by changing the frequency of administration of one or more of the disclosed isolated nucleic acid molecules, disclosed viral vectors, disclosed lentiviral vectors, disclosed pharmaceutical formulations, disclosed host cells, disclosed gRNAs, disclosed plasmids, or any combination thereof, or by changing the duration of time that the one or more of the disclosed isolated nucleic acid molecules, disclosed viral vectors, disclosed lentiviral vectors, disclosed pharmaceutical formulations, disclosed host cells, disclosed gRNAs, disclosed plasmids, or any combination thereof, or are administered to a subject.

As used herein, “concurrently” means (1) simultaneously in time, or (2) at different times during the course of a common treatment schedule.

The term “contacting” as used herein refers to bringing one or more of disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof together with a target area or intended target area in such a manner that the one or more of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof exert an effect on the intended target or targeted area either directly or indirectly. A target area or intended target area can be one or more of a subject's organs (e.g., lungs, heart, liver, kidney, brain, etc.). In an aspect, a target area or intended target area can be any cell or any organ infected by AD or LOAD (such as cholinergic neurons). In an aspect, a target area or intended target area can be the brain or various neuronal populations.

As used herein, “determining” can refer to measuring or ascertaining the presence and severity of AD such as, for example, LOAD. Methods and techniques used to determine the presence and/or severity of AD are typically known to the medical arts. For example, the art is familiar with the ways to identify and/or diagnose the presence, severity, or both of AD (such as, for example, a LOAD. In an aspect, “determining” can also refer to measuring or ascertaining the level of one or more proteins or peptides in a biosample, or measuring or ascertaining the level or one or more RNAs or miRNAs in a biosample. Methods and techniques for determining the level of proteins/peptides and RNAs/miRNAs are known to the art and are disclosed herein.

As used herein, “effective amount” and “amount effective” can refer to an amount that is sufficient to achieve the desired result such as, for example, the treatment and/or prevention of AD or LOAD. As used herein, the terms “effective amount” and “amount effective” can refer to an amount that is sufficient to achieve the desired an effect on an undesired condition (e.g., a AD or LOAD). For example, a “therapeutically effective amount” refers to an amount that is sufficient to achieve the desired therapeutic result or to have an effect on undesired symptoms, but is generally insufficient to cause adverse side effects. In an aspect, “therapeutically effective amount” means an amount of a disclosed isolated nucleic acid molecule, a disclosed pharmaceutical formulation, a disclosed vector, or any combination thereof that (i) treats the particular disease, condition, or disorder (e.g., AD or LOAD), (ii) attenuates, ameliorates, or eliminates one or more symptoms of the particular disease, condition, or disorder (e.g., AD or LOAD), or (iii) delays the onset of one or more symptoms of the particular disease, condition, or disorder described herein (e.g., AD or LOAD). The specific therapeutically effective dose level for any particular patient will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof employed; the disclosed methods employed; the age, body weight, general health, sex and diet of the patient; the time of administration; the route of administration; the rate of excretion of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof employed; the duration of the treatment; drugs used in combination or coincidental with the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof employed, and other like factors well known in the medical arts. For example, it is well within the skill of the art to start doses of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof at levels lower than those required to achieve the desired therapeutic effect and to gradually increase the dosage until the desired effect is achieved. If desired, then the effective daily dose can be divided into multiple doses for purposes of administration. Consequently, a single dose of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof can contain such amounts or submultiples thereof to make up the daily dose. The dosage can be adjusted by the individual physician in the event of any contraindications. Dosage can vary, and can be administered in one or more dose administrations daily, for one or several days. Guidance can be found in the literature for appropriate dosages for given classes of pharmaceutical products. In further various aspects, a preparation can be administered in a “prophylactically effective amount”; that is, an amount effective for prevention of a disease or condition, such as, for example, AD or LOAD

As used herein, the term “pharmaceutically acceptable carrier” refers to sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, as well as sterile powders for reconstitution into sterile injectable solutions or dispersions just prior to use. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents, or vehicles include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol and the like), carboxymethylcellulose and suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate. In an aspect, a pharmaceutical carrier employed can be a solid, liquid, or gas. In an aspect, examples of solid carriers can include lactose, terra alba, sucrose, talc, gelatin, agar, pectin, acacia, magnesium stearate, and stearic acid. In an aspect, examples of liquid carriers can include sugar syrup, peanut oil, olive oil, and water. In an aspect, examples of gaseous carriers can include carbon dioxide and nitrogen. In preparing a disclosed composition for oral dosage form, any convenient pharmaceutical media can be employed. For example, water, glycols, oils, alcohols, flavoring agents, preservatives, coloring agents and the like can be used to form oral liquid preparations such as suspensions, elixirs and solutions; while carriers such as starches, sugars, microcrystalline cellulose, diluents, granulating agents, lubricants, binders, disintegrating agents, and the like can be used to form oral solid preparations such as powders, capsules and tablets. Because of their ease of administration, tablets and capsules are the preferred oral dosage units whereby solid pharmaceutical carriers are employed. Optionally, tablets can be coated by standard aqueous or nonaqueous techniques. Proper fluidity can be maintained, for example, by the use of coating materials such as lecithin, by the maintenance of the required particle size in the case of dispersions and by the use of surfactants. These compositions can also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the action of microorganisms can be ensured by the inclusion of various antibacterial and antifungal agents such as paraben, chlorobutanol, phenol, sorbic acid and the like. It can also be desirable to include isotonic agents such as sugars, sodium chloride and the like. Prolonged absorption of the injectable pharmaceutical form can be brought about by the inclusion of agents, such as aluminum monostearate and gelatin, which delay absorption. Injectable depot forms are made by forming microencapsule matrices of the drug in biodegradable polymers such as polylactide-polyglycolide, poly(orthoesters) and poly(anhydrides). Depending upon the ratio of drug to polymer and the nature of the particular polymer employed, the rate of drug release can be controlled. Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues. The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable media just prior to use. Suitable inert carriers can include sugars such as lactose. Desirably, at least 95% by weight of the particles of the active ingredient have an effective particle size in the range of 0.01 to 10 micrometers.

As used herein, the term “excipient” refers to an inert substance which is commonly used as a diluent, vehicle, preservative, binder, or stabilizing agent, and includes, but is not limited to, proteins (e.g., serum albumin, etc.), amino acids (e.g., aspartic acid, glutamic acid, lysine, arginine, glycine, histidine, etc.), fatty acids and phospholipids (e.g., alkyl sulfonates, caprylate, etc.), surfactants (e.g., SDS, polysorbate, nonionic surfactant, etc.), saccharides (e.g., sucrose, maltose, trehalose, etc.) and polyols (e.g., mannitol, sorbitol, etc.). See, also, for reference, Remington's Pharmaceutical Sciences, (1990) Mack Publishing Co., Easton, Pa., which is hereby incorporated by reference in its entirety.

As used herein, the term “package insert” is used to refer to instructions customarily included in commercial packages of therapeutic products, that contain information about the indications, usage, dosage, administration, contraindications and/or warnings concerning the use of such therapeutic products.

As used herein, the term “in combination” in the context of the administration of one or more of the disclosed agents, disclosed therapeutic agents, disclosed pharmaceutical formulations or a combination thereof includes the use of more than one therapy (e.g., additional therapeutic agents). Administration “in combination with” one or more additional therapeutic agents includes simultaneous (e.g., concurrent) and consecutive administration in any order. The use of the term “in combination” does not restrict the order in which therapies are administered to a subject. By way of non-limiting example, a first therapy (e.g., one or more of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof) may be administered prior to (e.g., 1 minute, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, or 12 weeks), concurrently, or after (e.g., 1 minute, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, or 12 weeks or longer) the administration of a second therapy (e.g., one or more of the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof or one or more additional therapeutic agents) to a subject having or diagnosed with AD or LOAD.

As used herein, “CRISPR or clustered regularly interspaced short palindromic repeat” is an ideal tool for correction of genetic abnormalities associated with diseases such as Alzheimer's disease or LOAD. The system can be designed to target genomic DNA directly. A CRISPR system involves two main components: a Cas9 enzyme and a guide (gRNA). The gRNA contains a targeting sequence for DNA binding (at, for example, the APOE promoter region) and a scaffold sequence for Cas9 binding. Cas9 nuclease is often used to “knockout” target genes such as for example, the APOE e4 allele. Also, multiple gRNAs can be employed to suppress or activate multiple genes simultaneously, hence increasing the treatment efficacy and reducing resistance potentially caused by new mutations in the target genes.

As used herein, “CRISPR-based endonucleases” include RNA-guided endonucleases that comprise at least one nuclease domain and at least one domain that interacts with a guide RNA. As known to the art, a guide RNA directs the CRISPR-based endonucleases to a targeted site in a nucleic acid at which site the CRISPR-based endonucleases cleaves at least one strand of the targeted nucleic acid sequence. As the guide RNA provides the specificity for the targeted cleavage, the CRISPR-based endonuclease is universal and can be used with different guide RNAs to cleave different target nucleic acid sequences. CRISPR-based endonucleases are RNA-guided endonucleases derived from CRISPR/Cas systems.

In an aspect, a disclosed CRISPR-based endonuclease can be derived from a CRISPR/Cas type I, type II, or type III system. Non-limiting examples of suitable CRISPR/Cas proteins include Cas3, Cas4, Cas5, Cas5e (or CasD), Cas6, Cas6e, Cas6f, Cas7, Cas8a1, Cas8a2, Cas8b, Cas8c, Cas9, Cas10, Cas10d, CasF, CasG, CasH, Csy1, Csy2, Csy3, Cse1 (or CasA), Cse2 (or CasB), Cse3 (or CasE), Cse4 (or CasC), Csc1, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10, Csx16, CsaX, Csx3, Csz1, Csx15, Csf1, Csf2, Csf3, Csf4, and Cu1966.

In an aspect, a disclosed CRISPR-based endonuclease can be derived from a type II CRISPR/Cas system. For example, in an aspect, a CRISPR-based endonuclease can be derived from a Cas9 protein. The Cas9 protein can be from Streptococcus pyogenes, Streptococcus thermophilus, Streptococcus sp, Nocardiopsis dassonvillei, Streptomyces pristinaespiralis, Streptomyces viridochromogenes, Streptomyces viridochromogenes, Streptosporangium roseum, Streptosporangium roseum, Alicyclobacillus acidocaldarius, Bacillus pseudomycoides, Bacillus selenitireducens, Exiguobacterium sibiricum, Lactobacillus delbrueckii, Lactobacillus salivarius, Microscilla marina, Burkholderiales bacterium, Polaromonas naphthalenivorans, Polaromonas sp., Crocosphaera watsonii, Cyanothece sp., Microcystis aeruginosa, Synechococcus sp., Acetohalobium arabaticum, Ammonifex degensii, Caldicelulosiruptor becscii, Candidatus Desulforudis, Clostridium botulinum, Clostridium difficile, Finegoldia magna, Natranaerobius thermophilus, Pelotomaculum thermopropionicum, Acidithiobacillus caldus, Acidithiobacillus ferrooxidans, Allochromatium vinosum, Marinobacter sp., Nitrosococcus halophilus, Nitrosococcus watsoni, Pseudoalteromonas haloplanktis, Ktedonobacter racemifer, Methanohalobium evestigatum, Anabaena variabilis, Nodularia spumigena, Nostoc sp., Arthrospira maxima, Arthrospira platensis, Arthrospira sp., Lyngbya sp., Microcoleus chthonoplastes, Oscillatoria sp., Petrotoga mobilis, Thermosipho africanus, or Acaryochloris marina. In an aspect, the CRISPR-based nuclease can be derived from a Cas9 protein from Streptococcus pyogenes. In an aspect, the CRISPR-based nuclease can comprise the sequence set forth in SEQ ID NO:64 or SEQ ID NO:65.

As used herein, “CRISPRa” refers to CRISPR Activation, which is using a dCas9 or dCas9-activator with a gRNA to increase transcription of a target gene.

As used herein, “CRISPRi” refers to CRISPR Interference, which is using a dCas9 or dCas9-repressor with a gRNA to repress/decrease transcription of a target gene.

As used herein, “dCas9” refers to enzymatically inactive form of Cas9, which can bind, but cannot cleave, DNA.

As used herein, “Protospacer Adjacent Motif” or “PAM” refers to a sequence adjacent to the target sequence that is necessary for Cas enzymes to bind target DNA.

Disclosed are the components to be used to prepare the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof as well the disclosed isolated nucleic acid molecules, disclosed pharmaceutical formulations, disclosed vectors, or any combination thereof used within the methods disclosed herein. These and other materials are disclosed herein, and it is understood that when combinations, subsets, interactions, groups, etc. of these materials are disclosed that while specific reference of each various individual and collective combinations and permutation of these compounds cannot be explicitly disclosed, each is specifically contemplated and described herein. For example, if a particular compound is disclosed and discussed and a number of modifications that can be made to a number of molecules including the compounds are discussed, specifically contemplated is each and every combination and permutation of the compound and the modifications that are possible unless specifically indicated to the contrary. Thus, if a class of molecules A, B, and C are disclosed as well as a class of molecules D, E, and F and an example of a combination molecule, A-D is disclosed, then even if each is not individually recited each is individually and collectively contemplated meaning combinations, A-E, A-F, B-D, B-E, B-F, C-D, C-E, and C-F are considered disclosed. Likewise, any subset or combination of these is also disclosed. Thus, for example, the sub-group of A-E, B-F, and C-E would be considered disclosed. This concept applies to all aspects of this application including, but not limited to, steps in methods of making and using the compositions of the invention. Thus, if there are a variety of additional steps that can be performed it is understood that each of these additional steps can be performed with any specific aspects or combination of aspects of the disclosed methods.

B. Compositions
1. Isolated Nucleic Acid Molecules

In an aspect, a disclosed Cas endonuclease can comprise Cas9, SpCas9, SaCas9, a variant Cas9, a dCas, or a dCas9. In an aspect, a disclosed Cas9 can comprise the sequence set forth in SEQ ID NO:64 or SEQ ID NO:65. In an aspect, a disclosed Cas9 can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:64 or SEQ ID NO:65 or a fragment thereof.

In an aspect, a disclosed variant Cas9 can comprise VQR, EQR, or VRER. In an aspect a disclosed VRER can comprise the sequence set forth in SEQ ID NO:15. In an aspect, a disclosed VRER can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:15 or a fragment thereof. In an aspect, a disclosed dCas can comprise dVQR, dEQR, or dVRER. In an aspect, a disclosed dCas can comprise the sequence set forth in SEQ ID NO:16. In an aspect, a disclosed dCas9 can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:16 or a fragment thereof. A SpCas9 (3′NGG-PAM sequence) can comprise SpCas9 VQR (3′NGAN or 3′NGNG), SpCas9 EQR (3′NGAG), or SpCas9 VRER (3′NGCG).

In an aspect, a disclosed VRER can have the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth below:

(SEQ ID NO: 15)

atggataaaaagtattctattggtttagacatcggcactaattccgttggatgggctgtcataaccgatgaatacaaagtaccttcaaagaaatt

taaggtgttggggaacacagaccgtcattcgattaaaaagaatcttatcggtgccctcctattcgatagtggcgaaacggcagaggcgactc

gcctgaaacgaaccgctcggagaaggtatacacgtcgcaagaaccgaatatgttacttacaagaaatttttagcaatgagatggccaaagtt

gacgattctttctttcaccgtttggaagagtccttccttgtcgaagaggacaagaaacatgaacggcaccccatctttggaaacatagtagatg

aggtggcatatcatgaaaagtacccaacgatttatcacctcagaaaaaagctagttgactcaactgataaagcggacctgaggttaatctact

tggctcttgcccatatgataaagttccgtgggcactttctcattgagggtgatctaaatccggacaactcggatgtcgacaaactgttcatcca

gttagtacaaacctataatcagttgtttgaagagaaccctataaatgcaagtggcgtggatgcgaaggctattcttagcgcccgcctctctaaa

tcccgacggctagaaaacctgatcgcacaattacccggagagaagaaaaatgggttgttcggtaaccttatagcgctctcactaggcctga

caccaaattttaagtcgaacttcgacttagctgaagatgccaaattgcagcttagtaaggacacgtacgatgacgatctcgacaatctactgg

cacaaattggagatcagtatgcggacttatttttggctgccaaaaaccttagcgatgcaatcctcctatctgacatactgagagttaatactgag

attaccaaggcgccgttatccgcttcaatgatcaaaaggtacgatgaacatcaccaagacttgacacttctcaaggccctagtccgtcagca

actgcctgagaaatataaggaaatattctttgatcagtcgaaaaacgggtacgcaggttatattgacggcggagcgagtcaagaggaattct

acaagtttatcaaacccatattagagaagatggatgggacggaagagttgcttgtaaaactcaatcgcgaagatctactgcgaaagcagcg

gactttcgacaacggtagcattccacatcaaatccacttaggcgaattgcatgctatacttagaaggcaggaggatttttatccgttcctcaaa

gacaatcgtgaaaagattgagaaaatcctaacctttcgcataccttactatgtgggacccctggcccgagggaactctcggttcgcatggat

gacaagaaagtccgaagaaacgattactccatggaattttgaggaagttgtcgataaaggtgcgtcagctcaatcgttcatcgagaggatga

ccaactttgacaagaatttaccgaacgaaaaagtattgcctaagcacagtttactttacgagtatttcacagtgtacaatgaactcacgaaagtt

aagtatgtcactgagggcatgcgtaaacccgcctttctaagcggagaacagaagaaagcaatagtagatctgttattcaagaccaaccgca

aagtgacagttaagcaattgaaagaggactactttaagaaaattgaatgcttcgattctgtcgagatctccggggtagaagatcgatttaatgc

gtcacttggtacgtatcatgacctcctaaagataattaaagataaggacttcctggataacgaagagaatgaagatatcttagaagatatagtg

ttgactcttaccctctttgaagatcgggaaatgattgaggaaagactaaaaacatacgctcacctgttcgacgataaggttatgaaacagttaa

agaggcgtcgctatacgggctggggacgattgtcgcggaaacttatcaacgggataagagacaagcaaagtggtaaaactattctcgattt

tctaaagagcgacggcttcgccaataggaactttatgcagctgatccatgatgactctttaaccttcaaagaggatatacaaaaggcacaggt

ttccggacaaggggactcattgcacgaacatattgcgaatcttgctggttcgccagccatcaaaaagggcatactccagacagtcaaagta

gtggatgagctagttaaggtcatgggacgtcacaaaccggaaaacattgtaatcgagatggcacgcgaaaatcaaacgactcagaaggg

gcaaaaaaacagtcgagagcggatgaagagaatagaagagggtattaaagaactgggcagccagatcttaaaggagcatcctgtggaa

aatacccaattgcagaacgagaaactttacctctattacctacaaaatggaagggacatgtatgttgatcaggaactggacataaaccgtttat

ctgattacgacgtcgatcacattgtaccccaatcctttttgaaggacgattcaatcgacaataaagtgcttacacgctcggataagaaccgag

ggaaaagtgacaatgttccaagcgaggaagtcgtaaagaaaatgaagaactattggcggcagctcctaaatgcgaaactgataacgcaaa

gaaagttcgataacttaactaaagctgagaggggggcttgtctgaacttgacaaggccggatttattaaacgtcagctcgtggaaacccgc

caaatcacaaagcatgttgcacagatactagattcccgaatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaagtaa

tcactttaaagtcaaaattggtgtcggacttcagaaaggattttcaattctataaagttagggagataaataactaccaccatgcgcacgacgc

ttatcttaatgccgtcgtagggaccgcactcattaagaaatacccgaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtcc

gtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagccaaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaatggggagacaggtgaaatcgtatgggataagggccgggactt

cgcgacggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaactgaggtgcagaccggagggttttcaaaggaatcgatt

cttccaaaaaggaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaagtacggtggcttcgtgagccctacagttgcct

attctgtcctagtagtggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaattattggggataacgattatggagcgc

tcgtcttttgaaaagaaccccatcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctcataattaaactaccaaagtatagt

ctgtttgagttagaaaatggccgaaaacggatgttggctagcgccagagagcttcaaaaggggaacgaactcgcactaccgtctaaatacg

tgaatttcctgtatttagcgtcccattacgagaagttgaaaggttcacctgaagataacgaacagaagcaactttttgttgagcagcacaaaca

ttatctcgacgaaatcatagagcaaatttcggaattcagtaagagagtcatcctagctgatgccaatctggacaaagtattaagcgcatacaa

caagcacagggataaacccatacgtgagcaggcggaaaatattatccatttgtttactcttaccaacctcggcgctccagccgcattcaagta

ttttgacacaacgatagatcgcaaagagtacagatctaccaaggaggtgctagacgcgacactgattcaccaatccatcacgggattatatg

aaactcggatagatttgtcacagcttgggggtgactga.

In an aspect, dCas9 can have the following sequence:

(SEQ ID NO: 16)

MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGA

LLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHR

LEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKAD

LRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENP

INASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTP

NFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAI

LLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEI

FFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR

KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPY

YVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDK

NLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVD

LLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKI

IKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQ

LKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDD

SLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKV

MGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHP

VENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDD

SIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNL

TKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLI

REVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKK

YPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEI

TLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEV

QTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVE

KGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPK

YSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSPE

DNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDK

PIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQ

SITGLYETRIDLSQLGGD.

In an aspect, a disclosed encoded polypeptide can comprise transcription activation activity, transcription repression activity, transcription release factor activity, histone modification activity, nucleic acid association activity, methyltransferase activity, demethylase activity, acetyltransferase activity, deacetylase activity, or any combination thereof. In an aspect, a disclosed encoded polypeptide can be histone deacetylase or heterochromatin protein 1. In an aspect, a disclosed encoded polypeptide can comprise transcription repression activity. In an aspect, a disclosed DNMT3A can have the amino acid sequence set forth in SEQ ID NO:17 or the nucleotide sequence set forth in SEQ ID NO:18. In an aspect, a disclosed DNMT3A can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:17 or SEQ ID NO:18 or a fragment thereof.

In an aspect, a disclosed DNMT3A can have the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 17)

PSRLQMFFANNHDQEFDPPKVYPPVPAEKRKPIRVLSLFDGIATGLLV

LKDLGIQVDRYIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKHIQE

WGPFDLVIGGSPCNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKE

GDDRPFFWLFENVVAMGVSDKRDISRFLESNPVMIDAKEVSAAHRARY

FWGNLPGMNRPLASTVNDKLELQECLEHGRIAKFSKVRTITTRSNSIK

QGKDQHFPVFMNEKEDILWCTEMERVFGFPVHYTDVSNMSRLARQRLL

GRSWSVPVIRHLFAPLKEYFACV.

In an aspect, a disclosed DNMT3A can have the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 18)

Ccccccggctccagatgttcttcgctaataaccacgaccaggaatttgac

cctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccg

ggtgctgtctctctttgatggaatcgctacagggctcctggtgctgaagg

acttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggac

tccatcacggtgggcatggtgcggcaccaggggaagatcatgtacgtcgg

ggacgtccgcagcgtcacacagaagcatatccaggagtggggcccattcg

atctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccct

gctcgcaagggcctctacgagggcactggccggctcttctttgagttcta

ccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttct

tctggctctttgagaatgtggtggccatgggcgttagtgacaagagggac

atctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagt

gtcagctgcacacagggcccgctacttctggggtaaccttcccggtatga

acaggccgttggcatccactgtgaatgataagctggagctgcaggagtgt

ctggagcatggcaggatagccaagttcagcaaagtgaggaccattactac

gaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttca

tgaatgagaaagaggacatcttatggtgcactgaaatggaaagggtattt

ggtttcccagtccactatactgacgtgtccaacatgagccgcttggcgag

gcagagactgctgggccggtcatggagcgtgccagtcatccgccacctct

tcgctccgctgaaggagtattttgcgtgtgtg.

In an aspect, at least one encoded polypeptide can comprise Kroppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2). In an aspect, a disclosed TRD of MeCP2 can comprise the nucleotide sequence set forth in SEQ ID NO:57 or the amino acid sequence set forth in SEQ ID NO:58. In an aspect, a disclosed TRD of MeCP2 can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:57 or SEQ ID NO:58 or a fragment thereof. In an aspect, a disclosed KRAB-MeCP2 can comprise the nucleotide sequence set forth in SEQ ID NO:62 or the amino acid sequence set forth in SEQ ID NO:63. In an aspect, a disclosed KRAB-MeCP2 can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:62 or SEQ ID NO:63 or a fragment thereof.

In an aspect, a disclosed MeCP2 TRD can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 57)

atgtcggagggtgtgcaggtgaaaagggtcctggagaaaagtcctgggaa

gctccttgtcaagatgccttttcaaacttcgccagggggcaaggctgagg

ggggtggggccaccacatccacccaggtcatggtgatcaaacgccccggc

aggaagcgaaaagctgaggccgaccctcaggccattcccaagaaacgggg

ccgaaagccggggagtgtggtggcagccgctgccgccgaggccaaaaaga

aagccgtgaaggagtcttctatccgatctgtgcaggagacagtactcccc

atcaagaagcgcaagacccgggagtaa.

In an aspect, a disclosed MeCP2 TRD can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 58)

MSEGVQVKRVLEKSPGKLLVKMPFQTSPGGKAEGGGATTSTQVMVIKRPG

RKRKAEADPQAIPKKRGRKPGSVVAAAAAEAKKKAVKESSIRSVQETVLP

IKKRKTRE*.

In an aspect, a disclosed KRAB-MeCP2 repressor can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 62)

atgcggacactggtgaccttcaaggatgtatttgtggacttcaccaggga

ggagtggaagctgctggacactgctcagcagatcgtgtacagaaatgtga

tgctggagaactataagaacctggtttccttgggttatcagcttactaag

ccagatgtgatcctccggttggagaagggagaagagccctcgggaggtgg

ttcgggaggtggttcggagggtgtgcaggtgaaaagggtcctggagaaaa

gtcctgggaagctccttgtcaagatgccttttcaaacttcgccagggggc

aaggctgaggggggtggggccaccacatccacccaggtcatggtgatcaa

acgccccggcaggaagcgaaaagctgaggccgaccctcaggccattccca

agaaacggggccgaaagccggggagtgtggtggcagccgctgccgccgag

gccaaaaagaaagccgtgaaggagtcttctatccgatctgtgcaggagac

Agtactccccatcaagaagcgcaagacccgggagtaa.

In an aspect, a disclosed KRAB-MeCP2 repressor can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 63)

MRTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGYQLTK

PDVILRLEKGEEPSGGGSGGGSEGVQVKRVLEKSPGKLLVKMPFQTSPGG

KAEGGGATTSTQVMVIKRPGRKRKAEADPQAIPKKRGRKPGSVVAAAAAE

AKKKAVKESSIRSVQETVLPIKKRKTRE*.

In an aspect, a disclosed gRNA can be designed to target exon 4 of the APOE gene or designed to target a protospacer-adjacent motif (PAM) created by a SNP rs429358 in exon 4 of the APOE gene. In an aspect, a disclosed gRNA can have the sequence set forth in any one of SEQ ID NO:05-SEQ ID NO:14, SEQ ID NO:25-SEQ ID NO:28, SEQ ID NO:39-SEQ ID NO:42, and SEQ ID NO:51-SEQ ID NO:52.

In an aspect, a disclosed Cas endonuclease can be fused to a disclosed polypeptide having an enzymatic activity. In an aspect, a disclosed Cas endonuclease can be dCas9 and a disclosed polypeptide can be DNMT3A. In an aspect, a disclosed dCas9-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO: 19. In an aspect, a dCas9-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:20. In an aspect, a disclosed dCas9-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:19 or SEQ ID NO:20 or a fragment thereof. In an aspect, a disclosed Cas endonuclease can be dVRER and the polypeptide can be DNMT3A. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:38. In an aspect, a disclosed dVRER-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:37. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:38 or SEQ ID NO:37 or a fragment thereof.

In an aspect, a disclosed Cas endonuclease can be dCas9 and the polypeptide can be Krüppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2). In an aspect, a disclosed Cas endonuclease can be dVRER and the polypeptide can be Krüppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2).

In an aspect, a disclosed dCas9-DNMT3A fusion protein can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 19)

DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGAL

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

ITGLYETRIDLSQLGGDKRPAATKKAGQAKKKKLEGGGGSGSPSRLQMFF

ANNHDQEFDPPKVYPPVPAEKRKPIRVLSLFDGIATGLLVLKDLGIQVDR

YIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKHIQEWGPFDLVIGGSP

CNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKEGDDRPFFWLFENVV

AMGVSDKRDISRFLESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTV

NDKLELQECLEHGRIAKFSKVRTITTRSNSIKQGKDQHFPVFMNEKEDIL

WCTEMERVFGFPVHYTDVSNMSRLARQRLLGRSWSVPVIRHLFAPLKEYF

AC.

In an aspect, a disclosed dCas9-DNMT3A fusion protein can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth:

(SEQ ID NO: 20)

gacaagaagtacagcatcggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaaga

aattcaaggtgctgggcaacaccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccga

ggccacccggctgaagagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgag

atggccaaggtggacgacagcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttc

ggcaacatcgtggacgaggtggcctaccacgagaagtaccccaccattaccacctgagaaagaaactggtggacagcaccgacaagg

ccgacctgcggctgatctatctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaaca

gcgacgtggacaagctgttcatccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgcc

aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgtt

cggcaacctgattgccctgagcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagca

aggacacctacgacgacgacctggacaacctgctggcccagateggcgaccagtacgccgacctgtttctggccgccaagaacctgtcc

gacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacga

gcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaa

cggctacgccggctacattgacggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccg

aggaactgctcgtgaagctgaacagagaggacctgctgcggaagcageggaccttegacaacggcagcatcccccaccagatccacct

gggagagctgcacgccattctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgacctt

ccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggatgaccagaaagagegaggaaaccatcaccccct

ggaacttcgaggaagtggtggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgag

aaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgaga

aagcccgccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctga

aagaggactacttcaagaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacatacca

cgatctgctgaaaattatcaaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgt

ttgaggacagagagatgatcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggag

atacaccggctggggcaggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagt

ccgacggcttcgccaacagaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccg

gccagggcgatagcctgcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggt

ggacgagctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagg

gacagaagaacagccgcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtg

gaaaacacccagctgcagaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaa

ccggctgtccgactacgatgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcga

caagaaccggggcaagagcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagaga

cagctggtggaaacccggcagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagc

tgatccgggaagtgaaagtgatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaac

aactaccaccacgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgt

gtacggcgactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttct

acagcaacatcatgaactttttcaagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaa

accggggagatcgtgtgggataagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagac

cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggac

cctaagaagtacggcggcttcgacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaact

gaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggcta

caaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgc

cggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagg

gctcccccgaggataatgagcagaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttct

ccaagagagtgatcctggccgacgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggc

cgagaatatcatccacctgtttaccctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtaca

ccagcaccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctggg

aggcgacaaaaggccggcggccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatcccc

ctcccggctccagatgttcttcgctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagc

ccatccgggtgctgtctctctttgatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcgg

aggtgtgtgaggactccatcacggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaa

gcatatccaggagtggggcccattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctct

acgagggcactggccggctcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctc

tttgagaatgtggtggccatgggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagt

gtcagctgcacacagggcccgctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctg

caggagtgtctggagcatggcaggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaaga

ccagcattttcctgtgttcatgaatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactg

acgtgtccaacatgagccgcttggcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaag

gagtattttgcgtgtgtg.

In an aspect, a disclosed SpCas9-dVRER-DNMT3A can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth below:

(SEQ ID NO: 37)

atggactataaggaccacgacggagactacaaggatcatgatattgattacaaagacgatgacgataagatggccccaaagaagaagcg

gaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatcggcctggccatcggcaccaactctgtgggctgggccgtg

atcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacagcatcaagaagaacctgatcggag

ccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagagaaccgccagaagaagatacaccagacggaagaaccgg

atctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgacagcttcttccacagactggaagagtccttcctggtggaag

aggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacctg

agaaagaaactggtggacagcaccgacaaggccgacctgcggctgatctatctggccctggcccacatgatcaagttccggggccactt

cctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcatccagctggtgcagacctacaaccagctgttcgagg

aaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgc

ccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctgagcctgggcctgacccccaacttcaagagcaacttcga

cctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacgacctggacaacctgctggcccagateggcgaccagtac

gccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccc

cctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattgacggcggagccagccaggaagagttctacaagttca

tcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctt

cgacaacggcagcatcccccaccagatccacctgggagagctgcacgccattctgcggcggcaggaagatttttacccattcctgaagga

caaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggat

gaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggtggacaagggcgcttccgcccagagcttcatcgagcgg

atgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataacgagctga

ccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaagaaaatcgagtgcttcgactccgtggaaatctccggcgtgg

aagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatcaaggacaaggacttcctggacaatgaggaaaacgagg

acattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaacggctgaaaacctatgcccacctgttcga

cgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaacggcatccgggaca

agcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatccacgacgacagcctga

cctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacattgccaatctggccggcagccccgc

cattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatc

gaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatgaagcggatcgaagagggcatcaaag

agctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaagctgtacctgtactacctgcagaatgg

gcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtggacgctatcgtgcctcagagctttctgaaggac

gactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagaggtcgtgaagaag

atgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaagttcgacaatctgaccaaggccgagagaggcggcct

gagcgaactggataaggccggcttcatcaagagacagctggtggaaacccggcagatcacaaagcacgtggcacagatactagattccc

gaatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaagtaatcactttaaagtcaaaattggtgtcggacttcagaaagg

attttcaattctataaagttagggagataaataactaccaccatgcgcacgacgcttatcttaatgccgtcgtagggaccgcactcattaaga

aatacccgaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtccgtaagatgatcgcgaaaagcgaacaggagataggc

aaggctacagccaaatacttcttttattctaacattatgaatttctttaagacggaaatcactctggcaaacggagagatacgcaaacgacct

ttaattgaaaccaatggggagacaggtgaaatcgtatgggataagggccgggacttcgcgacggtgagaaaagttttgtccatgccccaagt

caacatagtaaagaaaactgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaaggaatagtgataagctcatcgctcgta

aaaaggactgggacccgaaaaagtacggtggcttcgtgagccctacagttgcctattctgtcctagtagtggcaaaagttgagaagggaaa

atccaagaaactgaagtcagtcaaagaattattggggataacgattatggagcgctcgtcttttgaaaagaaccccatcgacttccttgaggc

gaaaggttacaaggaagtaaaaaaggatctcataattaaactaccaaagtatagtctgtttgagttagaaaatggccgaaaacggatgttgg

ctagcgccagagagcttcaaaaggggaacgaactcgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccattacgagaagttga

aaggttcacctgaagataacgaacagaagcaactttttgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcggaattca

gtaagagagtcatcctagctgatgccaatctggacaaagtattaagcgcatacaacaagcacagggataaacccatacgtgagcaggcgg

aaaatattatccatttgtttactcttaccaacctcggcgctccagccgcattcaagtattttgacacaacgatagatcgcaaagagtacagat

ctaccaaggaggtgctagacgcgacactgattcaccaatccatcacgggattatatgaaactcggatagatttgtcacagcttgggggtgacg

gatcccccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcgctaataaccac

gaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctctttgatggaatcgc

tacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatcacggtgggcatg

gtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcccattcgatctggt

gattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggctcttctttgagttct

accgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatgggcgttagtga

caagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggcccgctacttctgg

ggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatggcaggatagcc

aagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatgaatgagaaag

aggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttggcgaggcag

agactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtg.

In an aspect, a disclosed VRER-DNMT3A can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth below:

(SEQ ID NO: 38)

MAPKKKRKVGIHGVPAADKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKV

LGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQE

IFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPT

IYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKL

FIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKK

NGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGD

QYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTL

LKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMD

GTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLK

DNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDK

GASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGM

RKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVE

DRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEE

RLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLK

SDGFANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHANLAGSPAIKK

GILQTVKVVDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIE

EGIKELGSQILKEHPVENTQLQNEKLYLYYLONGRDMYVDQELDINRLSD

YDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQ

LLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQIL

DSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAH

DAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAK

YFFYSNIMNFFKTEITLAGEIRKRPLIETNGETGEIVWDKGRDFATVRKV

LSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFVS

PTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKG

YKEVKKDLIIKLPKYSLFELENGRKRMLASARELQKGNLKGSPEDNEQKQ

LFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQA

ENIIHLFTLTNLGAPAAFKYFDTTIDRKEYRSTKEVLDATLIHQSITGLY

ETRIDLSQLGGDGSPKKKVLEGGGGSGSPSRLQMFFANNHDQEFDPPKVY

PPVPAEKRKPIRVLSLFDGIATGLLVLKDLGIQVDRYIASEVCEDSITVG

MVRHQGKIMYVGDVRSVTQKHIQEWGPFDLVIGGSPCNDLSIVNPARKGL

YEGTGRLFFEFYRLLHDARPKEGDDRPFFWLFENVVAMGVSDKRDISRFL

ESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTVNDKLELQECLEHRI

AKFSKVRTITTRSNSIKQGKDQHFPVFMNEKEDILWCTEMERVFGFPVHY

TDVSNMSRLARQRLLGRSWSVPVIRHLFAPLKEYFACV.

Disclosed herein is an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

In an aspect, a disclosed fusion protein can encode a disclosed Cas endonuclease and a disclosed polypeptide.

In an aspect, at least one encoded polypeptide can comprise Krüppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2). In an aspect, a disclosed TRD of MeCP2 can comprise the nucleotide sequence set forth in SEQ ID NO:57 or the amino acid sequence set forth in SEQ ID NO:58. In an aspect, a disclosed TRD of MeCP2 can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:57 or SEQ ID NO:58 or a fragment thereof. In an aspect, a disclosed KRAB-MeCP2 can comprise the nucleotide sequence set forth in SEQ ID NO:62 or the amino acid sequence set forth in SEQ ID NO:63. In an aspect, a disclosed KRAB-MeCP2 can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:62 or SEQ ID NO:63 or a fragment thereof.

In an aspect, a disclosed fusion protein can comprise dCas9 and DNMT3A. In an aspect, a dCas9-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:19. In an aspect, a dCas9-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:20. In an aspect, a disclosed dCas9-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:19 or SEQ ID NO:20 or a fragment thereof. In an aspect, a disclosed Cas endonuclease can be dVRER and the polypeptide can be DNMT3A. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:38. In an aspect, a disclosed dVRER-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:37. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:38 or SEQ ID NO:37 or a fragment thereof.

In an aspect, a disclosed fusion protein can comprise dCas9 and Krüppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2). In an aspect, a disclosed fusion protein can comprise dVRER and Krüppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2).

2. Non-Viral Vectors

Disclosed herein is a non-viral vector comprising a disclosed isolated nucleic acid molecule.

Disclosed herein is a non-viral vector comprising a disclosed isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA.

Disclosed herein is a non-viral vector comprising a disclosed isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

In an aspect, a disclosed non-viral vector can be a polymer based vector, a peptide based vector, a lipid nanoparticle, a solid lipid nanoparticle, or a cationic lipid based vector.

In an aspect, a disclosed non-viral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In an aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect, a disclosed Cas endonuclease can be fused to a disclosed polypeptide having an enzymatic activity. In an aspect, a disclosed fusion protein can encode a disclosed Cas endonuclease and a disclosed polypeptide. In an aspect, a disclosed fusion protein can comprise dCas9 and DNMT3A. In an aspect, a dCas9-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:19. In an aspect, a dCas9-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:20. In an aspect, a disclosed dCas9-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:19 or SEQ ID NO:20 or a fragment thereof. In an aspect, a disclosed Cas endonuclease can be dVRER and the polypeptide can be DNMT3A. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:38. In an aspect, a disclosed dVRER-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:37. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:38 or SEQ ID NO:37 or a fragment thereof.

In an aspect, a disclosed non-viral vector can comprise one or more regulatory elements. Regulatory elements are known in the art and can comprise one or more of the following: a Sp1 responsive element, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR. In an aspect, a disclosed non-viral vector can comprise two Sp1 response elements, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR.

3. Viral Vectors

Disclosed herein is a viral vector comprising a disclosed isolated nucleic acid molecule.

Disclosed herein is a viral vector comprising a disclosed isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA.

In an aspect, a disclosed viral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In an aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect, a disclosed viral vector can comprise one or more regulatory elements. Regulatory elements are known in the art and can comprise one or more of the following: a Sp1 responsive element, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR. In an aspect, a disclosed viral vector can comprise two Sp1 response elements, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR.

In an aspect, a disclosed viral vector can be an adenovirus vector, an AAV vector, a herpes simplex virus vector, a retrovirus vector, a lentivirus vector, and alphavirus vector, a flavivirus vector, a rhabdovirus vector, a measles virus vector, a Newcastle disease viral vector, a poxvirus vector, or a picornavirus vector. In an aspect, a disclosed viral vector can be a lentiviral vector.

Disclosed herein is a viral vector comprising a disclosed isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

In an aspect, a disclosed viral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In a aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect, a disclosed fusion protein can encode a disclosed Cas endonuclease and a disclosed polypeptide. In an aspect, a disclosed fusion protein can comprise dCas9 and DNMT3A. In an aspect, a dCas9-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:19. In an aspect, a dCas9-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:20. In an aspect, a disclosed dCas9-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:19 or SEQ ID NO:20 or a fragment thereof. In an aspect, a disclosed Cas endonuclease can be dVRER and the polypeptide can be DNMT3A. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:38. In an aspect, a disclosed dVRER-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:37. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:38 or SEQ ID NO:37 or a fragment thereof.

In an aspect, a disclosed viral vector can comprise one or more regulatory elements. Regulatory elements are known in the art and can comprise one or more of the following: a Sp1 responsive element, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR. In an aspect, a disclosed vector can comprise two Sp1 response elements, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR.

In an aspect, a disclosed viral vector can be a lentiviral vector.

4. Lentiviral Vectors

In an aspect, a disclosed lentiviral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In an aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect, a disclosed encoded polypeptide can comprise transcription activation activity, transcription repression activity, transcription release factor activity, histone modification activity, nucleic acid association activity, methyltransferase activity, demethylase activity, acetyltransferase activity, deacetylase activity, or any combination thereof. In an aspect, a disclosed encoded polypeptide can be histone deacetylase or heterochromatin protein 1. In an aspect, a disclosed encoded polypeptide can comprise transcription repression activity. In an aspect, a disclosed DNMT3A can have the amino acid sequence set forth in SEQ ID NO: 17 or the nucleotide sequence set forth in SEQ ID NO:18. In an aspect, a disclosed DNMT3A can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:17 or SEQ ID NO:18 or a fragment thereof.

In an aspect, a disclosed fusion protein can comprise dCas9 and Kroppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2). In an aspect, a disclosed fusion protein can comprise dVRER and Krüppel-associated box (KRAB), the transcription repression domain (TRD) of Methyl-CpG Binding Protein 2 (MeCP2), or a fusion of KRAB-MeCP2 (KRAB-MeCP2).

In an aspect, a disclosed lentiviral vector can comprise one or more regulatory elements. Regulatory elements are known in the art and can comprise one or more of the following: a Sp1 responsive element, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR. In an aspect, a disclosed lentiviral vector can comprise two Sp1 response elements, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR.

Disclosed herein is a lentiviral vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

5. Pharmaceutical Formulations

In an aspect, a disclosed formulation can comprise (i) one or more active agents, (ii) biologically active agents, (iii) one or more pharmaceutically active agents, (iv) one or more immune-based therapeutic agents, (v) one or more clinically approved agents, or (vi) a combination thereof. In an aspect, a disclosed composition can comprise one or more proteasome inhibitors. In an aspect, a disclosed composition can comprise one or more immunosuppressives or immunosuppressive agents. In an aspect, an immunosuppressive agent can be anti-thymocyte globulin (ATG), cyclosporine (CSP), mycophenolate mofetil (MMF), or a combination thereof. In an aspect, a disclosed formulation can comprise a RNA therapeutic. A RNA therapeutic can comprise RNA-mediated interference (RNAi) and/or antisense oligonucleotides (ASO). In an aspect, a disclosed formulation can comprise a disclosed small molecule.

6. Host Cells

Disclosed herein is a host cell comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is a host cell comprising a lentiviral vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a fusion protein and (ii) at least one guide RNA.

Disclosed herein is a host cell comprising plasmid comprising the sequence set forth in any one of SEQ ID NO:21-24, SEQ ID NO:29-36, SEQ ID NO:43-50, SEQ ID NO:53-56, SEQ ID NO:59-61.

In an aspect, a disclosed viral vector or a disclosed lentiviral vector in a disclosed host cell can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In an aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect, a disclosed viral vector or a disclosed lentiviral vector in a disclosed host cell can comprise one or more regulatory elements. Regulatory elements are known in the art and can comprise one or more of the following: a Sp1 responsive element, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR. In an aspect, a disclosed viral vector or a disclosed lentiviral vector can comprise two Sp1 response elements, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR.

7. Guide RNAs (gRNAs)

Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:05-SEQ ID NO: 14. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:25-SEQ ID NO:28. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:39-SEQ ID NO:42. Disclosed herein is a guide RNA comprising the sequence set forth in any one of SEQ ID NO:51-SEQ ID NO:52. Disclosed gRNAs are listed below.

SEQ

ID NO.
gRNA Sequence
Description

5
gacagggggagccctataat
gRNA1 targeting promoter region of ApoE using

SpCas9

6
tcaggagagctactcggggt
gRNA2 targeting promoter region of ApoE using

SpCas9

7
actgggatgtaagccatagc
gRNA3 targeting promoter region of ApoE using

SpCas9

8
gttggagcttagaatgtgaa
gRNA4 targeting promoter region of ApoE using

SpCas9

9
gccctatccctgggggaggg
gRNA1 targeting promoter region of ApoE using VRER

Cas9

10
tcgggcttggggagaggagg
gRNA2 targeting promoter region of ApoE using VRER

Cas9

11
ctctccccaccccaccttct
gRNA3 targeting promoter region of ApoE using VRER

Cas9

12
tgtgaagggagaatgaggaa
gRNA4 targeting promoter region of ApoE using VRER

Cas9

13
ggcgaggagctgttcaccg
gRNA targeting GFP ORF using SpCas9

14
gccacaagttcagcgtgtcc
gRNA targeting GFP ORF using VRER-SpCas9

25
gacagggggagccctataat
gRNA1 cloned into pBK1026 and pBK1030 plasmids

26
tcaggagagctactcggggt
gRNA2 cloned into pBK1027 and pBK1031 plasmids

27
actgggatgtaagccatagc
gRNA3 cloned into pBK1028 and pBK1032 plasmids

28
gttggagcttagaatgtgaa
gRNA4 cloned into pBK1029 and pBK1033 plasmids

39
gccctatccctgggggaggg
gRNA1 cloned into pBK1105 and pBK1109 plasmids

40
tcgggcttggggagaggagg
gRNA2 cloned into pBK1106 and pBK1110 plasmids

41
ctctccccaccccaccttct
gRNA3 cloned into pBK1107 and pBK1111 plasmids

42
tgtgaagggagaatgaggaa
gRNA4 cloned into pBK1108 and pBK1112 plasmids

51
gggcgcggacatggaggacg
gRNA1 cloned into pBK1426 and pBK1428 plasmids

52
gggcgcggacatggaggacg
gRNA2 cloned into pBK1427 and pBK1429 plasmids

As known to the art, a gRNA provides the targeting of a CRISPR/Cas9-based epigenome modifying system. A guide RNA is a specific RNA sequence that recognizes the target DNA region of interest (such as, for example, APOE e4 allele) and directs the Cas endonuclease there for editing. The gRNA is made up of two parts: crispr RNA (crRNA), a 17-20 nucleotide sequence complementary to the target DNA, and a tracer RNA, which serves as a binding scaffold for the Cas nuclease. The CRISPR-associated (Cas) protein is a non-specific endonuclease, which can be directed to the specific DNA locus by a gRNA (where it makes a double-strand break).

In an aspect, a disclosed gRNA can serve to direct a disclosed endonucleases or a disclosed fusion product having an endonuclease to a target area of interest (such as, for example, the promoter of the APOE gene or the APOE e4 allele).

8. Plasmids

Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:21-SEQ ID NO:24. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:29-SEQ ID NO:36. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:43-SEQ ID NO:50. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:53-SEQ ID NO:56. Disclosed herein is a plasmid comprising the sequence set forth in any of SEQ ID NO:59-SEQ ID NO:61. Plasmids disclosed herein include but are not limited to those listed below.

SEQ ID NO
Plasmid Description

21
pBK546 carrying dCas9-DNMT3A fused transgene linked to puromycin

reporter via p2A cleavage signal

22
pBK539 carrying dCas9-DNMT3A fused transgene linked to GFP reporter via

p2A cleavage signal

23
pBK500 carrying all-in-one lentiviral vector containing fusion protein and

gRNA4

24
pBK744 carrying dCas9-DNMT3A fused transgene linked to GFP reporter via

p2A cleavage signal and carrying gRNA3 targeting rat/mouse intron Snca-

intron 1 sequences

29
pBK1026 carrying dCas9-(active) DNMT3A vector targeting promoter region

of ApoE gene and carrying gRNA1 and puromycin reporter separated via p2a

signal from DNMT3A and SpCas9 is present

30
pBK1027 carrying dCas9-(active) DNMT3A vector targeting promoter region

of ApoE gene and carrying gRNA2 and puromycin reporter separated via p2a

signal from DNMT3A and SpCas9 is present

31
pBK1028 carrying dCas9-(active) DNMT3A vector targeting promoter region

of ApoE gene and carrying gRNA3 and puromycin reporter separated via p2a

signal from DNMT3A and SpCas9 is present

32
pBK1029 carrying dCas9-(active) DNMT3A vector targeting promoter region

of ApoE gene and carrying gRNA4 and puromycin reporter separated via p2a

signal from DNMT3A and SpCas9 is present

33
pBK1030 carrying dCas9-(inactive) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA1 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

34
pBK1031 carrying dCas9-(inactive) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA2 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

35
pBK1032 carrying dCas9-(inactive) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA3 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

36
pBK1033 carrying dCas9-(inactive) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA4 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

43
pBK1105 carrying dCas9-VRER (active) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA1 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

44
pBK1106 carrying dCas9-VRER (active) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA2 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

45
pBK1107 carrying dCas9-VRER (active) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA3 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

46
pBK1108 carrying dCas9-VRER (active) DNMT3A vector targeting promoter

region of ApoE gene and carrying gRNA4 and puromycin reporter separated

via p2a signal from DNMT3A and SpCas9 is present

47
pBK1109 carrying dCas9-VRER (inactive) DNMT3A vector targeting

promoter region of ApoE gene and carrying gRNA1 and puromycin reporter

separated via p2a signal from DNMT3A and SpCas9 is present

48
pBK1110 carrying dCas9-VRER (inactive) DNMT3A vector targeting

promoter region of ApoE gene and carrying gRNA2 and puromycin reporter

separated via p2a signal from DNMT3A and SpCas9 is present

49
pBK1111 carrying dCas9-VRER (inactive) DNMT3A vector targeting

promoter region of ApoE gene and carrying gRNA3 and puromycin reporter

separated via p2a signal from DNMT3A and SpCas9 is present

50
pBK1112 carrying dCas9-VRER (inactive) DNMT3A vector targeting

promoter region of ApoE gene and carrying gRNA3 and puromycin reporter

separated via p2a signal from DNMT3A and SpCas9 is present

53
pBK1426 carrying dCas9-VRER (active) DNMT3A vector targeting SNP

region of ApoE4 and carrying gRNA1 and puromycin reporter separated via

p2a signal from DNMT3A and VRER-SpCas9 is present

54
pBK1427 carrying dCas9-VRER (active) DNMT3A vector targeting SNP

region of ApoE4 and carrying gRNA2 and puromycin reporter separated via

p2a signal from DNMT3A and VRER-SpCas9 is present

55
pBK1428 carrying dCas9-VRER (inactive) DNMT3A vector targeting SNP

region of ApoE4 and carrying gRNA1 and puromycin reporter separated via

p2a signal from DNMT3A and VRER-SpCas9 is present

56
pBK1429 carrying dCas9-VRER DNMT3A vector targeting SNP region of

ApoE4 and carrying gRNA2 and puromycin reporter separated via p2a signal

from DNMT3A and VRER-SpCas9 is present

59
pBK1531 carrying a Lentiviral vector with dCas9-VRER-MeCP2

Transcription Repression Domain (TRD) and gRNA1

60
pBK1532 carrying a Lentiviral vector with dCas9-VRER-MeCP2

Transcription Repression Domain (TRD) and gRNA2

61
pBK1536 carrying a Lentiviral vector with dCas9-VRER-MeCP2

Transcription Repression Domain (TRD) without gRNA

In an aspect, a disclosed pBK546 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 21)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataatt

tcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatatct

tgtggaaaggacgaaacaccggagacgtgtacacgtctctgttttagagctagaaatagcaagttaaaataaggctagtccgttatcaacttg

aaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcaga

gcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaactg

ggaaagtgatgtcgtgtactggctccgcctttttcccgagggtgggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttcg

caacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattaca

aagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatcg

gcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaac

accgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagaga

accgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgaca

gcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgagg

tggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatctat

ctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgaccgagtacaagcc

cacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccgegttcgccgactaccccgccacgcgcca

caccgtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccgagatcggccc

gcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgctccccggagtggaggcgg

ccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacctccccttctacgageggcteggcttcaccgtcaccgcc

gacgtcgaggtgcccgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctctggatt

acaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgc

ttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgtggtgt

gcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgctttccccctccctattgcc

acggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattccgtggtgttgtcggggaaa

tcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcct

ccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattc

actcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaa

cccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagac

ccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttg

cccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtagg

tgtcattctattctggggggtggggggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccccacgcgccctgtagcggcgcattaagcgcggcgggt

gtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttettcccttcctttctcgccacgttcgccg

gctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtga

tggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgttctttaatagtggactcttgttccaaactgg

aacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgatttaacaaaaat

ttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatc

tcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccat

agtcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcag

aggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccgggagc

ttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcatagtatatcggcatagtataatacgacaaggtgaggaacta

aaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctggaccgaccggctcgggt

tctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggaccaggtggtg

ccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtg

cacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttc

cgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaa

ataaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtc

tgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtc

gggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctc

actgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata

acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgc

ccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctgg

aagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagc

tcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagcc

agttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggt

catgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccag

ccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagta

gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggtt

cccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttgg

ccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaa

ccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaacttta

aaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgca

cccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataaggg

cgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaat

gtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK539 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 22)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataat

ttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatat

cttgtggaaaggacgaaacaccggagacgtgtacacgtctctgttttagagctagaaatagcaagttaaaataaggctagtccgttatcaacttg

aaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcaga

gcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaactg

ggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttcg

caacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattaca

aagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatcg

gcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaac

accgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagaga

accgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgaca

gcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgagg

tggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatctat

ctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcategagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggggccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgatggtgag

caagggcgaggagctgttcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagttcagcgtgtccggc

gagggcgagggcgatgccacctacggcaagctgaccctgaagttcatctgcaccaccggcaagctgcccgtgccctggcccaccctcgt

gaccaccctgacctacggcgtgcagtgcttcagccgctaccccgaccacatgaagcagcacgacttcttcaagtccgccatgcccgaagg

ctacgtccaggagcgcaccatcttcttcaaggacgacggcaactacaagacccgcgccgaggtgaagttcgagggcgacaccctggtga

accgcatcgagctgaagggcatcgacttcaaggaggacggcaacatcctggggcacaagctggagtacaactacaacagccacaacgt

ctatatcatggccgacaagcagaagaacggcatcaaggtgaacttcaagatccgccacaacatcgaggacggcagcgtgcagctcgcc

gaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctgagcacccagtccgccctgagca

aagaccccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatcactctcggcatggacgagctgtacaag

taaagcggccgcgtcgacaatcaacctctggattacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtgga

tacgctgctttaatgcctttgtatcatgctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgagga

gttgtggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctc

ctttccgggactttcgctttccccctccctattgccacggeggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttg

ggcactgacaattccgtggtgttgtcggggaagctgacgtcctttccatggctgctcgcctgtgttgccacctggattctgcgcgggacgtc

cttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctettccgcgtcttegccttcgc

cctcagacgagtcggatctccctttgggccgcctccccgcctggaattcgagctcggtacctttaagaccaatgacttacaaggcagctgta

gatcttagccactttttaaaagaaaaggggggactggaagggctaattcactcccaacgaagacaagatctgctttttgcttgtactgggtctc

tctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaag

tagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtagtagttcatgtc

atcttattattcagtatttataacttgcaaagaaatgaatatcagagagtgagaggaacttgtttattgcagcttataatggttacaaataaagcaa

tagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtctggctctagc

tatcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcagag

gccgaggccgcctcggcctctgagctattccagaagtagtgaggaggcttttttggaggcctagggacgtacccaattcgccctatagtga

gtcgtattacgcgcgctcactggccgtcgttttacaacgtcgtgactgggaaaaccctggcgttacccaacttaatcgccttgcagcacatcc

ccctttcgccagctggcgtaatagcgaagaggcccgcaccgatcgcccttcccaacagttgcgcagcctgaatggcgaatgggacgcgc

cctgtagcggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagegcccgctcctttcg

ctttcttcccttcctttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggca

cctcgaccccaaaaaacttgattagggtgatggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgt

tctttaatagtggactcttgttccaaactggaacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggccta

ttggttaaaaaatgagctgatttaacaaaaatttaacgcgaattttaacaaaatattaacgcttacaatttaggtgccggccatgaccgagatcg

gcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtgcacttcgtggccgaggagcaggactgacac

gtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttccgggacgccggctggatgatcctccagcg

cggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaataaagcaatagcatcacaaatttcacaa

ataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtctgtataccgtcgacctctagctagagct

tggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccggaagcataaagtgtaaagcctg

gggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcattaatg

aatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgttcggctg

cggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaag

gccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcga

cgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttccga

ccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgta

ggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactategtcttgagtccaacc

cggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacagagttcttga

agtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagccagttaccttcggaaaaagagttggtagct

cttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagat

cctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcaccta

gatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacct

atctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggcccca

gtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagccggaagggccgagcgcagaag

tggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagtagttcgccagttaatagtttgcgcaacgttgt

tgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcc

cccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatggttatggcag

cactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggc

gaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcg

gggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaactgatcttcagcatcttttactttcac

cagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataagggcgacacggaaatgttgaatactcatactctt

cctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK500 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 23)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggat

agcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaaga

attattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctccca

accccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttgc

atatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataa

tttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttata

tatcttgtggaaaggacgaaacaccgctgctcagggtagatagctggttttagagctagaaatagcaagttaaaataaggctagtccgttatcaa

cttgaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcag

agcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaact

gggaaagtgatgtcgtgtactggctccgcctttttcccgagggtgggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggttctagagcgctgccaccatggacaagaagtacagcatcggcctggacatcggcac

caactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacagc

atcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagagaaccgccagaagaaga

tacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgacagcttcttccacagactg

gaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgaggtggcctaccacgaga

agtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatctatctggccctggcccac

atgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcatccagctggtgcag

acctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaagagca

gacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggaaacctgattgccctgagcctgggcctgacc

cccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacgacctggacaacctgct

ggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcgacatcctgagagtga

acaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgaccctgctgaaagctctc

gtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattgacggcggagccag

ccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacc

tgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccattctgcggcggcagga

agatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgggccctctggccagg

ggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggtggacaagggcgctt

ccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgag

tacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagcggcgagcagaaaaa

ggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaagaaaatcgagtgcttcga

ctccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatcaaggacaaggacttc

ctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaacggctg

aaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaag

ctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaacagaaacttcatgcag

ctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacattgc

caatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggcac

aagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatgaagcg

gatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaagctgta

cctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtggaccatatcgtg

cctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaagagcgacaacgtgccct

ccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaagttcgacaatctgacc

aaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccggcagatcacaaagcac

gtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagtgatcaccctgaagtc

caagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacgacgcctacctgaac

gccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgtacgacgtgcggaa

gatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaactttttcaagaccgagatta

ccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtgggataagggccgggatt

ttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggcttcagcaaagagtct

atcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttcgacagccccaccg

tggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgctggggatcaccatca

tggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacctgatcatcaagctgc

ctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaaacgaactggccctg

ccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagcagaaacagctgtttgt

ggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccgacgctaatctggaca

aagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttaccctgaccaatctgg

gagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctggacgccaccctgatc

caccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaagcgacctgccgccacaaagaaggct

ggacaggctaagaagaagaaagattacaaagacgatgacgataagggatccggcgcaacaaacttctctctgctgaaacaagccggag

atgtcgaagagaatcctggaccgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccct

cgccgccgcgttcgccgactaccccgccacgcgccacacegtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaact

cttcctcacgcgcgtcgggctcgacatcggcaaggtgtgggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagag

cgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatgga

aggcctcctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtct

gggcagcgccgtcgtgctccccggagtggaggcggccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacct

ccccttctacgagcggctcggcttcaccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgcatgaccegcaagcccggt

gcctgaacgcgttaagtcgacaatcaacctctggattacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctat

gtggatacgctgctttaatgcctttgtatcatgctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctct

ttatgaggagttgtggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacc

tgtcagctcctttccgggactttcgctttccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcg

gctgttgggcactgacaattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcggga

cgtccttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttcegcgtcttcgcctt

cgccctcagacgagtcggatctccctttgggccgcctccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactt

tttaaaagaaaaggggggactggaagggctaattcactcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagacca

gatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccg

tctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagc

ctcgactgtgccttctagttgccagccatctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcct

aataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctattctggggggtggggggggcaggacagcaagggggaggattg

ggaagacaatagcaggcatgctggggatgcggtgggctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccc

cacgcgccctgtagcggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctc

ctttcgctttcttcccttcctttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgct

ttacggcacctcgaccccaaaaaacttgattagggtgatggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttg

gagtccacgttctttaatagtggactcttgttccaaactggaacaacactcaaccctatctcggtctattcttttgatttataagggattttgcc

gatttcggcctattggttaaaaaatgagctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtc

cccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggca

gaagtatgcaaagcatgcatctcaattagtcagcaaccatagtcccgcccctaactccgcccatcccgcccctaactccgcccagttcegcc

cattctccgccccatggctgactaattttttttatttatgcagaggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggctt

ttttggaggcctaggcttttgcaaaaagctcccgggagcttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcata

gtatatcggcatagtataatacgacaaggtgaggaactaaaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtc

gccggagcggtcgagttctggaccgaccggctcgggttctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgt

gaccctgttcatcagcgcggtccaggaccaggtggtgccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtac

gccgagtggtcggaggtcgtgtccacgaacttccgggacgcctccgggccggccatgaccgagatcggcgagcagccgtgggggcg

ggagttcgccctgcgcgacccggccggcaactgcgtgcacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattcca

ccgccgccttctatgaaaggttgggcttcggaatcgttttccgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttc

gcccaccccaacttgtttattgcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattc

tagttgtggtttgtccaaactcatcaatgtatcttatcatgtctgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgt

ttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaa

ctcacattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggag

aggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactc

aaaggcggtaatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccg

taaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaa

cccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgt

ccgcctttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctg

tgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccac

tggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctaca

ctagaagaacagtatttggtatctgcgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccacc

gctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagt

tttaaatcaatctaaagtatatatgagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcg

ttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacc

cacgctcaccggctccagatttatcagcaataaaccagccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctcca

tccagtctattaattgttgccgggaagctagagtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtg

tcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggtta

gctccttcggtcctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatg

ccatccgtaagatgcttttctgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtc

aatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttac

cgctgttgagatccagttcgatgtaacccactcgtgcacccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaaca

ggaaggcaaaatgccgcaaaaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatc

agggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacc

tgac.

In an aspect, a disclosed pBK744 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 24)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttgca

tatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataattt

cttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatatct

tgtggaaaggacgaaacaccgtttttcaagcggaaacgctagttttagagctagaaatagcaagttaaaataaggctagtccgttatcaactt

gaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcag

agcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaact

gggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatc

ggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa

caccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagag

aaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgac

agcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgag

gtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatcta

tctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggggccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgatggtgag

caagggcgaggagctgttcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagttcagcgtgtccggc

gagggcgagggcgatgccacctacggcaagctgaccctgaagttcatctgcaccaccggcaagctgcccgtgccctggcccaccctcgt

gaccaccctgacctacggcgtgcagtgcttcagccgctaccccgaccacatgaagcagcacgacttcttcaagtccgccatgcccgaagg

ctacgtccaggagcgcaccatcttcttcaaggacgacggcaactacaagacccgcgccgaggtgaagttcgagggcgacaccctggtga

accgcatcgagctgaagggcatcgacttcaaggaggacggcaacatcctggggcacaagctggagtacaactacaacagccacaacgt

ctatatcatggccgacaagcagaagaacggcatcaaggtgaacttcaagatccgccacaacatcgaggacggcagcgtgcagctcgcc

gaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctgagcacccagtccgccctgagca

aagaccccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatcactctcggcatggacgagctgtacaag

taaagcggccgcgtcgacaatcaacctctggattacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtgga

tacgctgctttaatgcctttgtatcatgctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgagg

agttgtggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctc

ctttccgggactttcgctttccccctccctattgccacggeggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttg

ggcactgacaattccgtggtgttgtcggggaagctgacgtcctttccatggctgctcgcctgtgttgccacctggattctgcgcgggacgtc

cttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgc

cctcagacgagtcggatctccctttgggccgcctccccgcctggaattcgagctcggtacctttaagaccaatgacttacaaggcagctgta

gatcttagccactttttaaaagaaaaggggggactggaagggctaattcactcccaacgaagacaagatctgctttttgcttgtactgggtctc

tctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaag

tagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtagtagttcatgtc

atcttattattcagtatttataacttgcaaagaaatgaatatcagagagtgagaggaacttgtttattgcagcttataatggttacaaataaagcaa

tagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtctggctctag

ctatcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcaga

ggccgaggccgcctcggcctctgagctattccagaagtagtgaggaggcttttttggaggcctagggacgtacccaattcgccctatagtga

gtcgtattacgcgcgctcactggccgtcgttttacaacgtcgtgactgggaaaaccctggcgttacccaacttaatcgccttgcagcacatcc

ccctttcgccagctggcgtaatagcgaagaggcccgcaccgatcgcccttcccaacagttgcgcagcctgaatggcgaatgggacgcgc

cctgtagcggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcg

ctttcttcccttcctttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggca

cctcgaccccaaaaaacttgattagggtgatggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgtt

ctttaatagtggactcttgttccaaactggaacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggccta

ttggttaaaaaatgagctgatttaacaaaaatttaacgcgaattttaacaaaatattaacgcttacaatttaggtgccggccatgaccgagatcg

gcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtgcacttcgtggccgaggagcaggactgacac

gtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttccgggacgccggctggatgatcctccagcg

cggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaat

aaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtctgtataccgtcgacctctagctagagcttgg

cgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccggaagcataaagtgtaaagcctg

gggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcattaatg

aatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgttcggctg

cggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaag

gccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcga

cgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttccga

ccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgta

ggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaacc

cggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacagagttcttga

agtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagccagttaccttcggaaaaagagttggtagct

cttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagat

cctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcaccta

gatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacct

atctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggcccca

gtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagccggaagggccgagcgcagaag

tggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagtagttcgccagttaatagtttgcgcaacgttgt

tgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcc

cccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatggttatggcag

cactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggc

gaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcg

gggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaactgatcttcagcatcttttactttcac

cagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataagggcgacacggaaatgttgaatactcatactctt

cctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggttcc

gcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1026 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 29)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattategtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataat

ttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatat

cttgtggaaaggacgaaacaccggacagggggagccctataatgttttagagctagaaatagcaagttaaaataaggctagtccgttatcaac

ttgaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggca

gagcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaac

tgggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatc

ggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa

caccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagag

aaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgac

agcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgag

gtggcctaccacgagaagtaccccaccattaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatcta

tctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgaccgagtacaagcc

cacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccgegttcgccgactaccccgccacgcgcca

caccgtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccgagatcggccc

gcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgctccccggagtggaggcgg

ccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacctccccttctacgageggcteggcttcaccgtcaccgcc

gacgtcgaggtgcccgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctctggattaca

aaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgc

ttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgtg

gtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgctttccccctcccta

ttgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattccgtggtgttgtcggggaaa

tcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcct

ccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattc

actcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaa

cccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagac

ccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttg

cccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtagg

tgtcattctattctggggggtggggggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccccacgcgccctgtagcggcgcattaagcgcggcgggt

gtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttcctttctcgccacgttcgccg

gctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtga

tggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgttctttaatagtggactcttgttccaaactgg

aacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgatttaacaaaa

atttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatc

tcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccat

agtcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcag

aggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccgggagc

ttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcatagtatatcggcatagtataatacgacaaggtgaggaacta

aaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctggaccgaccggctcgggt

tctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggaccaggtggtg

ccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtg

cacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttc

cgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaa

taaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtc

tgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtc

gggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctc

actgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata

acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgc

ccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctgg

aagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagc

tcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagcc

agttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggt

catgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccag

ccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagta

gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggtt

cccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttgg

ccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaa

ccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaacttta

aaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgca

cccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataaggg

cgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaat

gtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1027 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 30)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataat

ttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatat

cttgtggaaaggacgaaacaccgtcaggagagctactcggggtgttttagagctagaaatagcaagttaaaataaggctagtccgttatcaact

tgaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcag

agcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaact

gggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatc

ggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa

caccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagag

aaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgac

agcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgag

gtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatcta

tctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgaccgagtacaagcc

cacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccgegttegccgactaccccgccacgcgcca

caccgtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccgagatcggccc

gcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgctccccggagtggaggcgg

ccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacctccccttctacgageggctcggcttcaccgtcaccgcc

gacgtcgaggtgcccgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctctggatt

acaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgc

ttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgtggt

gtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgctttccccctccctattgcc

acggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattccgtggtgttgtcggggaaa

tcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcct

ccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattc

actcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaa

cccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagac

ccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttg

cccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtagg

tgtcattctattctggggggtggggggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccccacgcgccctgtagcggcgcattaagcgcggcgggt

gtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttcctttctcgccacgttcgccg

gctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtga

tggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgttctttaatagtggactcttgttccaaactgg

aacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgatttaacaaaa

atttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatc

tcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccat

agtcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcag

aggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccgggagc

ttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcatagtatatcggcatagtataatacgacaaggtgaggaacta

aaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctggaccgaccggctcgggt

tctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggaccaggtggtg

ccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtg

cacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttc

cgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaa

taaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtc

tgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtc

gggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctc

actgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata

acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgc

ccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctgg

aagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagc

tcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagcc

agttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggt

catgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccag

ccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagta

gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggtt

cccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttgg

ccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaa

ccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaacttta

aaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgca

cccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataaggg

cgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaat

gtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1028 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth below:

(SEQ ID NO: 31)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataat

ttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatat

cttgtggaaaggacgaaacaccgactgggatgtaagccatagcgttttagagctagaaatagcaagttaaaataaggctagtccgttatcaact

tgaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcag

agcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaact

gggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatc

ggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa

caccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagag

aaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgac

agcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgag

gtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatcta

tctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagateggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgaccgagtacaagcc

cacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccgegttcgccgactaccccgccacgcgcca

caccgtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccgagatcggccc

gcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgctccccggagtggaggcgg

ccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacctccccttctacgageggctcggcttcaccgtcaccgcc

gacgtcgaggtgcccgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctctggatt

acaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgc

ttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgtggtgt

gcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgctttccccctccctattgcc

acggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattccgtggtgttgtcggggaaa

tcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcct

ccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattc

actcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaa

cccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagac

ccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttg

cccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtagg

tgtcattctattctggggggtggggggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccccacgcgccctgtagcggcgcattaagcgcggcgggt

gtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttcctttctcgccacgttcgccg

gctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtga

tggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgttctttaatagtggactcttgttccaaactgg

aacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgatttaacaaaaat

ttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatc

tcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccat

agtcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcag

aggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccgggagc

ttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcatagtatatcggcatagtataatacgacaaggtgaggaacta

aaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctggaccgaccggctcgggt

tctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggaccaggtggtg

ccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtg

cacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttc

cgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaa

taaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtc

tgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtc

gggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctc

actgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata

acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgc

ccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctgg

aagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagc

tcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagcc

agttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggt

catgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccag

ccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagta

gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggtt

cccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttgg

ccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaa

ccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaacttta

aaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgca

cccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataaggg

cgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaa

tgtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1029 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth below:

(SEQ ID NO: 32)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatggggggacgttaacggggggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataat

ttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatat

cttgtggaaaggacgaaacaccggttggagcttagaatgtgaagttttagagctagaaatagcaagttaaaataaggctagtccgttatcaactt

gaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggcag

agcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaact

gggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatc

ggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa

caccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagag

aaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgac

gtggcctaccacgagaagtaccccaccattaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatcta

tctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgaccgagtacaagcc

cacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccgegttcgccgactaccccgccacgcgcca

caccgtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccgagatcggccc

gcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgctccccggagtggaggcgg

ccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacctccccttctacgageggcteggcttcaccgtcaccgcc

gacgtcgaggtgcccgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctctggatt

acaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgc

ttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgtggt

gcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgetttccccctccctattgcc

gtacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattccgtggtgttgtcggggaaa

tcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcct

ccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattc

actcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaa

cccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagac

ccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttg

cccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtagg

tgtcattctattctggggggtggggggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccccacgcgccctgtagcggcgcattaagcgcggcgggt

gtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttcctttctcgccacgttcgccg

gctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtga

tggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgttctttaatagtggactcttgttccaaactgg

aacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgatttaacaaaa

atttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatc

tcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccat

agtcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcag

aggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccgggagc

ttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcatagtatatcggcatagtataatacgacaaggtgaggaacta

aaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctggaccgaccggctcgggt

tctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggaccaggtggtg

ccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtg

cacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttc

cgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaa

taaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtc

tgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtc

gggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctc

actgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata

acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgc

ccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctgg

aagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagc

tcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagcc

agttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggt

catgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccag

ccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagta

gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggtt

cccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttgg

ccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaa

ccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaacttta

aaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgca

cccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataaggg

cgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaa

tgtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1030 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 33)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagtacaatctgctctgatgccgcatagttaagccagtatctgctccctg

cttgtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaacaaggcaaggcttgaccgacaattgcatgaagaatctgctt

agggttaggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggg

gtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgcccatt

gacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggtggagtatttacggtaaactgcccacttg

gcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgac

cttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggata

gcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatgt

cgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcagcgcgttttgcctgtactgggtct

ctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaa

gtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaa

cagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgagg

ggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcaggga

gctagaacgattcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacag

gatcagaagaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttag

acaagatagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagg

gacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgc

agagagaaaaaagagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacg

ctgacggtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgca

actcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttgggg

ttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctgg

atggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaag

aattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggag

gcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattcaccattatgtttcagacccacctcccaac

cccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatc

ggcactgcgtgcgccaattctgcagacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagtgcagggg

aaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacaggg

acagcagagatccagtttggttaattaatgggcgggacgttaacggggcggaacggtaccgagggcctatttcccatgattccttcatatttg

catatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataat

ttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatatct

tgtggaaaggacgaaacaccggacagggggagccctataatgttttagagctagaaatagcaagttaaaataaggctagtccgttatcaac

ttgaaaaagtggcaccgagtcggtgcttttttgaattcgctagctaggtcttgaaaggagtgggaattggctccggtgcccgtcagtgggca

gagcgcacatcgcccacagtccccgagaagttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgcggggtaaac

tgggaaagtgatgtcgtgtactggctccgcctttttcccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttctttttc

gcaacgggtttgccgccagaacacaggaccggtgccaccatggactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggtatccacggagtcccagcagccgacaagaagtacagcatc

ggcctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa

caccgaccggcacagcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaacagccgaggccacccggctgaagag

aaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtggacgac

agcttcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatcgtggacgag

gtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatcta

tctggccctggcccacatgatcaagttccggggccacttcctgatcgagggcgacctgaaccccgacaacagcgacgtggacaagctgtt

catccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgccaaggccatcctgtctgcca

gactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgattgccctg

agcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagctgagcaaggacacctacgacgacg

acctggacaacctgctggcccagateggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgacgccatcctgctgagcg

acatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgacc

ctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattttcttcgaccagagcaagaacggctacgccggctacattg

acggcggagccagccaggaagagttctacaagttcatcaagcccatcctggaaaagatggacggcaccgaggaactgctcgtgaagct

gaacagagaggacctgctgcggaagcagcggaccttcgacaacggcagcatcccccaccagatccacctgggagagctgcacgccatt

ctgcggcggcaggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaagatcctgaccttccgcatcccctactacgtgg

gccctctggccaggggaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcaccccctggaacttcgaggaagtggt

ggacaagggcgcttccgcccagagcttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcac

agcctgctgtacgagtacttcaccgtgtataacgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagc

ggcgagcagaaaaaggccatcgtggacctgctgttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaaga

aaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatc

aaggacaaggacttcctggacaatgaggaaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatg

atcgaggaacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggca

ggctgagccggaagctgatcaacggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaac

agaaacttcatgcagctgatccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcct

gcacgagcacattgccaatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaa

gtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgca

gaacgagaagctgtacctgtactacctgcagaatggggggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacga

tgtggacgctatcgtgcctcagagctttctgaaggacgactccatcgacaacaaggtgctgaccagaagcgacaagaaccggggcaaga

gcgacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagctgattacccagagaaa

gttcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacccgg

cagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaagt

gatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacg

acgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggcgactacaaggtgt

acgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacagcaacatcatgaacttttt

caagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaaccggggagatcgtgtggga

taagggccgggattttgccaccgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccgaggtgcagacaggcggct

tcagcaaagagtctatcctgcccaagaggaacagcgataagctgatcgccagaaagaaggactgggaccctaagaagtacggcggcttc

gacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagctgct

ggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagccaagggctacaaagaagtgaaaaaggacc

tgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaatgctggcctctgccggcgaactgcagaagggaa

acgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagca

gaaacagctgtttgtggaacagcacaagcactacctggacgagatcatcgagcagatcagcgagttctccaagagagtgatcctggccga

cgctaatctggacaaagtgctgtccgcctacaacaagcaccgggataagcccatcagagagcaggccgagaatatcatccacctgtttac

cctgaccaatctgggagcccctgccgccttcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctgg

acgccaccctgatccaccagagcatcaccggcctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggccggcgg

ccacgaaaaaggccggacaggccaaaaagaaaaagctcgagggcggaggcgggagcggatccccctcccggctccagatgttcttcg

ctaataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcccatccgggtgctgtctctcttt

gatggaatcgctacagggctcctggtgctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtgtgaggactccatca

cggtgggcatggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtggggcc

cattcgatctggtgattgggggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagggagatgatcgccccttcttctggctctttgcgaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcagctgcacacagggccc

gctacttctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaatgataagctggagctgcaggagtgtctggagcatgg

caggatagccaagttcagcaaagtgaggaccattactacgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgttcatg

aatgagaaagaggacatcttatggtgcactgaaatggaaagggtatttggtttcccagtccactatactgacgtgtccaacatgagccgcttg

gcgaggcagagactgctgggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccg

gccggcccggatccggcgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaatcctggaccgaccgagtacaagcc

cacggtgcgcctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgccacgcgcca

caccgtcgatccggaccgccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccgagatcggccc

gcgcatggccgagttgagcggttcccggctggccgcgcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgctccccggagtggaggcgg

ccgagcgcgccggggtgcccgccttcctggagacctccgcgccccgcaacctccccttctacgageggcteggcttcaccgtcaccgcc

gacgtcgaggtgcccgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctctggatt

acaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgc

ttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgtggtgt

gcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgctttccccctccctattgcc

acggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattccgtggtgttgtcggggaaa

tcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcct

ccccgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattc

actcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaa

cccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagac

ccttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttg

cccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtagg

tgtcattctattctggggggtggggggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggtatccccacgcgccctgtagcggcgcattaagcgcggcgggt

gtggtggttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttcctttctegccacgttcgccg

gctttccccgtcaagctctaaatcgggggctccctttagggttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtga

tggttcacgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgttctttaatagtggactcttgttccaaactgg

aacaacactcaaccctatctcggtctattcttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgatttaacaaaaat

ttaacgcgaattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatc

tcaattagtcagcaaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagcaaccat

agtcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgcag

aggccgaggccgcctctgcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccgggagc

ttgtatatccattttcggatctgatcagcacgtgttgacaattaatcatcggcatagtatatcggcatagtataatacgacaaggtgaggaacta

aaccatggccaagttgaccagtgccgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctggaccgaccggctcgggt

tctcccgggacttcgtggaggacgacttcgccggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggaccaggtggtg

ccggacaacaccctggcctgggtgtgggtgcgcggcctggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtggggggggagttcgccctgcgcgacccggccggcaactgcgtg

cacttcgtggccgaggagcaggactgacacgtgctacgagatttcgattccaccgccgccttctatgaaaggttgggcttcggaatcgttttc

cgggacgccggctggatgatcctccagcgcggggatctcatgctggagttcttcgcccaccccaacttgtttattgcagcttataatggttacaaa

taaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtc

tgtataccgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtc

gggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctc

actgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata

acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgc

ccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctgg

aagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagc

tcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagcc

agttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggt

catgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccag

ccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagta

gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccggtt

cccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttgg

ccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttctgtgactggtgagtactcaa

ccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagcagaacttta

aaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgca

cccaactgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataaggg

cgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaat

gtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1031 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 34)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cgtcaggagagctactcggggtgttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatcctggactcccg

gatgaacactaagtacgacgagaatgacaagctgatccgggaagtgaa

agtgatcaccctgaagtccaagctggtgtccgatttccggaaggattt

ccagttttacaaagtgcgcgagatcaacaactaccaccacgcccacga

cgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccc

taagctggaaagcgagttcgtgtacggcgactacaaggtgtacgacgt

gcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgc

caagtacttcttctacagcaacatcatgaactttttcaagaccgagat

taccctggccaacggcgagatccggaagcggcctctgatcgagacaaa

cggcgaaaccggggagatcgtgtgggataagggccgggattttgccac

cgtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagac

cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagag

gaacagcgataagctgatcgccagaaagaaggactgggaccctaagaa

gtacggcggcttcgacagccccaccgtggcctattctgtgctggtggt

ggccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaaga

gctgctggggatcaccatcatggaaagaagcagcttcgagaagaatcc

catcgactttctggaagccaagggctacaaagaagtgaaaaaggacct

gatcatcaagctgcctaagtactccctgttcgagctggaaaacggccg

gaagagaatgctggcctctgccggcgaactgcagaagggaaacgaact

ggccctgccctccaaatatgtgaacttcctgtacctggccagccacta

tgagaagctgaagggctcccccgaggataatgagcagaaacagctgtt

tgtggaacagcacaagcactacctggacgagatcatcgagcagatcag

cgagttctccaagagagtgatcctggccgacgctaatctggacaaagt

gctgtccgcctacaacaagcaccgggataagcccatcagagagcaggc

cgagaatatcatccacctgtttaccctgaccaatctgggagcccctgc

cgccttcaagtactttgacaccaccatcgaccggaagaggtacaccag

caccaaagaggtgctggacgccaccctgatccaccagagcatcaccgg

cctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaag

gccggcggccacgaaaaaggccggacaggccaaaaagaaaaagctcga

gggcggaggcgggagcggatccccctcccggctccagatgttcttcgc

taataaccacgaccaggaatttgaccctccaaaggtttacccacctgt

cccagctgagaagaggaagcccatccgggtgctgtctctctttgatgg

aatcgctacagggctcctggtgctgaaggacttgggcattcaggtgga

ccgctacattgcctcggaggtgtgtgaggactccatcacggtgggcat

ggtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgt

cacacagaagcatatccaggagtggggcccattcgatctggtgattgg

gggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaaggg

cctctacgagggcactggccggctcttctttgagttctaccgcctcct

gcatgatgcgcggcccaaggagggagatgatcgccccttcttctggct

ctttgcgaatgtggtggccatgggcgttagtgacaagagggacatctc

gcgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtc

agctgcacacagggcccgctacttctggggtaaccttcccggtatgaa

caggccgttggcatccactgtgaatgataagctggagctgcaggagtg

tctggagcatggcaggatagccaagttcagcaaagtgaggaccattac

tacgaggtcaaactccataaagcagggcaaagaccagcattttcctgt

gttcatgaatgagaaagaggacatcttatggtgcactgaaatggaaag

ggtatttggtttcccagtccactatactgacgtgtccaacatgagccg

cttggcgaggcagagactgctgggccggtcatggagcgtgccagtcat

ccgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccgg

ccggcccggatccggcgcaacaaacttctctctgctgaaacaagccgg

agatgtcgaagagaatcctggaccgaccgagtacaagcccacggtgcg

cctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgc

cgcgttcgccgactaccccgccacgcgccacaccgtcgatccggaccg

ccacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgt

cgggctcgacatcggcaaggtgtgggtcgcggacgacggcgccgcggt

ggcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgc

cgagatcggcccgcgcatggccgagttgagcggttcccggctggccgc

gcagcaacagatggaaggcctcctggcgccgcaccggcccaaggagcc

cgcgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaa

gggtctgggcagcgccgtcgtgctccccggagtggaggcggccgagcg

cgccggggtgcccgccttcctggagacctccgcgccccgcaacctccc

cttctacgagcggctcggcttcaccgtcaccgccgacgtcgaggtgcc

cgaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaac

gcgttaagtcgacaatcaacctctggattacaaaatttgtgaaagatt

gactggtattcttaactatgttgctccttttacgctatgtggatacgc

tgctttaatgcctttgtatcatgctattgcttcccgtatggctttcat

tttctcctccttgtataaatcctggttgctgtctctttatgaggagtt

gtggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctga

cgcaacccccactggttggggcattgccaccacctgtcagctcctttc

cgggactttcgctttccccctccctattgccacggcggaactcatcgc

cgcctgccttgcccgctgctggacaggggctcggctgttgggcactga

caattccgtggtgttgtcggggaaatcatcgtcctttccttggctgct

cgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgt

cccttcggccctcaatccagcggaccttccttcccgcggcctgctgcc

ggctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcg

gatctccctttgggccgcctccccgcgtcgactttaagaccaatgact

tacaaggcagctgtagatcttagccactttttaaaagaaaagggggga

ctggaagggctaattcactcccaacgaagacaagatctgctttttgct

tgtactgggtctctctggttagaccagatctgagcctgggagctctct

ggctaactagggaacccactgcttaagcctcaataaagcttgccttga

gtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactag

agatccctcagacccttttagtcagtgtggaaaatctctagcagggcc

cgtttaaacccgctgatcagcctcgactgtgccttctagttgccagcc

atctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgc

cactcccactgtcctttcctaataaaatgaggaaattgcatcgcattg

tctgagtaggtgtcattctattctggggggtggggggggcaggacagc

aagggggaggattgggaagacaatagcaggcatgctggggatgcggtg

ggctctatggcttctgaggcggaaagaaccagctggggctctaggggg

tatccccacgcgccctgtagcggcgcattaagcgcggcgggtgtggtg

gttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgct

cctttcgctttcttcccttcctttctcgccacgttcgccggctttccc

cgtcaagctctaaatcgggggctccctttagggttccgatttagtgct

ttacggcacctcgaccccaaaaaacttgattagggtgatggttcacgt

agtgggccatcgccctgatagacggtttttcgccctttgacgttggag

tccacgttctttaatagtggactcttgttccaaactggaacaacactc

aaccctatctcggtctattcttttgatttataagggattttgccgatt

tcggcctattggttaaaaaatgagctgatttaacaaaaatttaacgcg

aattaattctgtggaatgtgtgtcagttagggtgtggaaagtccccag

gctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcag

caaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgc

aaagcatgcatctcaattagtcagcaaccatagtcccgcccctaactc

cgcccatcccgcccctaactccgcccagttccgcccattctccgcccc

atggctgactaattttttttatttatgcagaggccgaggccgcctctg

cctctgagctattccagaagtagtgaggaggcttttttggaggcctag

gcttttgcaaaaagctcccgggagcttgtatatccattttcggatctg

atcagcacgtgttgacaattaatcatcggcatagtatatcggcatagt

ataatacgacaaggtgaggaactaaaccatggccaagttgaccagtgc

cgttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctg

gaccgaccggctcgggttctcccgggacttcgtggaggacgacttcgc

cggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccagga

ccaggtggtgccggacaacaccctggcctgggtgtgggtgcgcggcct

ggacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccg

ggacgcctccgggccggccatgaccgagatcggcgagcagccgtgggg

ggggagttcgccctgcgcgacccggccggcaactgcgtgcacttcgtg

gccgaggagcaggactgacacgtgctacgagatttcgattccaccgcc

gccttctatgaaaggttgggcttcggaatcgttttccgggacgccggc

tggatgatcctccagcgcggggatctcatgctggagttcttcgcccac

cccaacttgtttattgcagcttataatggttacaaataaagcaatagc

atcacaaatttcacaaataaagcatttttttcactgcattctagttgt

ggtttgtccaaactcatcaatgtatcttatcatgtctgtataccgtcg

acctctagctagagcttggcgtaatcatggtcatagctgtttcctgtg

tgaaattgttatccgctcacaattccacacaacatacgagccggaagc

ataaagtgtaaagcctggggtgcctaatgagtgagctaactcacatta

attgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgc

cagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgt

attgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtc

gttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacgg

ttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaa

ggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgttt

ttccataggctccgcccccctgacgagcatcacaaaaatcgacgctca

agtcagaggtggcgaaacccgacaggactataaagataccaggcgttt

ccccctggaagctccctcgtgcgctctcctgttccgaccctgccgctt

accggatacctgtccgcctttctcccttcgggaagcgtggcgctttct

catagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctcc

aagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcc

ttatccggtaactatcgtcttgagtccaacccggtaagacacgactta

tcgccactggcagcagccactggtaacaggattagcagagcgaggtat

gtaggcggtgctacagagttcttgaagtggtggcctaactacggctac

actagaagaacagtatttggtatctgcgctctgctgaagccagttacc

ttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgct

ggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaa

aaaggatctcaagaagatcctttgatcttttctacggggtctgacgct

cagtggaacgaaaactcacgttaagggattttggtcatgagattatca

aaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaa

tcaatctaaagtatatatgagtaaacttggtctgacagttaccaatgc

ttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatcc

atagttgcctgactccccgtcgtgtagataactacgatacgggagggc

ttaccatctggccccagtgctgcaatgataccgcgagacccacgctca

ccggctccagatttatcagcaataaaccagccagccggaagggccgag

cgcagaagtggtcctgcaactttatccgcctccatccagtctattaat

tgttgccgggaagctagagtaagtagttcgccagttaatagtttgcgc

aacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgttt

ggtatggcttcattcagctccggttcccaacgatcaaggcgagttaca

tgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccg

atcgttgtcagaagtaagttggccgcagtgttatcactcatggttatg

gcagcactgcataattctcttactgtcatgccatccgtaagatgcttt

tctgtgactggtgagtactcaaccaagtcattctgagaatagtgtatg

cggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcg

ccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcg

gggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatg

taacccactcgtgcacccaactgatcttcagcatcttttactttcacc

agcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaag

ggaataagggcgacacggaaatgttgaatactcatactcttccttttt

caatattattgaagcatttatcagggttattgtctcatgagcggatac

atatttgaatgtatttagaaaaataaacaaataggggttccgcgcaca

tttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1032 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 35)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggggtaggcgtgtacggtgggaggtctatataagcag

cgcgttttgcctgtactgggtctctctggttagaccagatctgagcct

gggagctctctggctaactagggaacccactgcttaagcctcaataaa

gcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgact

ctggtaactagagatccctcagacccttttagtcagtgtggaaaatct

ctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccaga

ggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaag

aggcgaggggcggcgactggtgagtacgccaaaaattttgactagcgg

aggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaa

gaaaaaatataaattaaaacatatagtatgggcaagcagggagctaga

acgattcgcagttaatcctggcctgttagaaacatcagaaggctgtag

acaaatactgggacagctacaaccatcccttcagacaggatcagaaga

acttagatcattatataatacagtagcaaccctctattgtgtgcatca

aaggatagagataaaagacaccaaggaagctttagacaagatagagga

agagcaaaaaaaagtaagaccaccgcacagcaagcggccgctgatctt

cagacctggaggaggagatatgagggacaattggagaagtgaattata

taaatataaagtagtaaaaattgaaccattaggagtagcacccaccaa

ggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatagg

agctttgttccttgggttcttgggagcagcaggaagcactatgggcgc

agcgtcaatgacgctgacggtacaggccagacaattattgtctggtat

agtgcagcagcagaacaatttgctgagggctattgaggcgcaacagca

tctgttgcaactcacagtctggggcatcaagcagctccaggcaagaat

cctggctgtggaaagatacctaaaggatcaacagctcctggggatttg

gggttgctctggaaaactcatttgcaccactgctgtgccttggaatgc

tagttggagtaataaatctctggaacagatttggaatcacacgacctg

gatggagtgggacagagaaattaacaattacacaagcttaatacactc

cttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaatt

attggaattagataaatgggcaagtttgtggaattggtttaacataac

aaattggctgtggtatataaaattattcataatgatagtaggaggctt

ggtaggtttaagaatagtttttgctgtactttctatagtgaatagagt

taggcagggatattcaccattatcgtttcagacccacctcccaacccc

gaggggacccgacaggcccgaaggaatagaagaagaaggtggagagag

agacagagacagatccattcgattagtgaacggatcggcactgcgtgc

gccaattctgcagacaaatggcagtattcatccacaattttaaaagaa

aaggggggattggggggtacagtgcaggggaaagaatagtagacataa

tagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaa

ttcaaaattttcgggtttattacagggacagcagagatccagtttggt

taattaatggggggacgttaacggggcggaacggtaccgagggcctat

ttcccatgattccttcatatttgcatatacgatacaaggctgttagag

agataattagaattaatttgactgtaaacacaaagatattagtacaaa

atacgtgacgtagaaagtaataatttcttgggtagtttgcagttttaa

aattatgttttaaaatggactatcatatgcttaccgtaacttgaaagt

atttcgatttcttggctttatatatcttgtggaaaggacgaaacaccg

actgggatgtaagccatagcgttttagagctagaaatagcaagttaaa

ataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtgc

ttttttgaattcgctagctaggtcttgaaaggagtgggaattggctcc

ggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaag

ttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgc

ggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttccc

gaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttc

tttttcgcaacgggtttgccgccagaacacaggaccggtgccaccatg

gactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggt

atccacggagtcccagcagccgacaagaagtacagcatcggcctggcc

atcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaag

gtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacagc

atcaagaagaacctgatcggagccctgctgttcgacagcggcgaaaca

gccgaggccacccggctgaagagaaccgccagaagaagatacaccaga

cggaagaaccggatctgctatctgcaagagatcttcagcaacgagatg

gccaaggtggacgacagcttcttccacagactggaagagtccttcctg

gtggaagaggataagaagcacgagcggcaccccatcttcggcaacatc

gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctg

agaaagaaactggtggacagcaccgacaaggccgacctgcggctgatc

tatctggccctggcccacatgatcaagttccggggccacttcctgatc

gagggcgacctgaaccccgacaacagcgacgtggacaagctgttcatc

cagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaac

gccagcggcgtggacgccaaggccatcctgtctgccagactgagcaag

agcagacggctggaaaatctgatcgcccagctgcccggcgagaagaag

aatggcctgttcggcaacctgattgccctgagcctgggcctgaccccc

aacttcaagagcaacttcgacctggccgaggatgccaaactgcagctg

agcaaggacacctacgacgacgacctggacaacctgctggcccagatc

ggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgac

gccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaag

gcccccctgagcgcctctatgatcaagagatacgacgagcaccaccag

gacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaag

tacaaagagattttcttcgaccagagcaagaacggctacgccggctac

attgacggcggagccagccaggaagagttctacaagttcatcaagccc

atcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaac

agagaggacctgctgcggaagcagcggaccttcgacaacggcagcatc

ccccaccagatccacctgggagagctgcacgccattctgcggcggcag

gaagatttttacccattcctgaaggacaaccgggaaaagatcgagaag

atcctgaccttccgcatcccctactacgtgggccctctggccagggga

aacagcagattcgcctggatgaccagaaagagcgaggaaaccatcacc

ccctggaacttcgaggaagtggtggacaagggcgcttccgcccagagc

ttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaag

gtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataac

gagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgcc

ttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaag

accaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaag

aaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcgg

ttcaacgcctccctgggcacataccacgatctgctgaaaattatcaag

gacaaggacttcctggacaatgaggaaaacgaggacattctggaagat

atcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaa

cggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcag

ctgaagcggcggagatacaccggctggggcaggctgagccggaagctg

atcaacggcatccgggacaagcagtccggcaagacaatcctggatttc

ctgaagtccgacggcttcgccaacagaaacttcatgcagctgatccac

gacgacagcctgacctttaaagaggacatccagaaagcccaggtgtcc

ggccagggcgatagcctgcacgagcacattgccaatctggccggcagc

cccgccattaagaagggcatcctgcagacagtgaaggtggtggacgag

ctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcgaa

atggccagagagaaccagaccacccagaagggacagaagaacagccgc

gagagaatgaagcggatcgaagagggcatcaaagagctgggcagccag

atcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaag

ctgtacctgtactacctgcagaatggggggatatgtacgtggaccagg

aactggacatcaaccggctgtccgactacgatgtggacgctatcgtgc

ctcagagctttctgaaggacgactccatcgacaacaaggtgctgacca

gaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagagg

tcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagc

tgattacccagagaaagttcgacaatctgaccaaggccgagagaggcg

gcctgagcgaactggataaggccggcttcatcaagagacagctggtgg

aaacccggcagatcacaaagcacgtggcacagatcctggactcccgga

tgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaag

tgatcaccctgaagtccaagctggtgtccgatttccggaaggatttcc

agttttacaaagtgcgcgagatcaacaactaccaccacgcccacgacg

cctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtacccta

agctggaaagcgagttcgtgtacggcgactacaaggtgtacgacgtgc

ggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgcca

agtacttcttctacagcaacatcatgaactttttcaagaccgagatta

ccctggccaacggcgagatccggaagcggcctctgatcgagacaaacg

gcgaaaccggggagatcgtgtgggataagggccgggattttgccaccg

tgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagaccg

aggtgcagacaggcggcttcagcaaagagtctatcctgcccaagagga

acagcgataagctgatcgccagaaagaaggactgggaccctaagaagt

acggcggcttcgacagccccaccgtggcctattctgtgctggtggtgg

ccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagagc

tgctggggatcaccatcatggaaagaagcagcttcgagaagaatccca

tcgactttctggaagccaagggctacaaagaagtgaaaaaggacctga

tcatcaagctgcctaagtactccctgttcgagctggaaaacggccgga

agagaatgctggcctctgccggcgaactgcagaagggaaacgaactgg

ccctgccctccaaatatgtgaacttcctgtacctggccagccactatg

agaagctgaagggctcccccgaggataatgagcagaaacagctgtttg

tggaacagcacaagcactacctggacgagatcatcgagcagatcagcg

agttctccaagagagtgatcctggccgacgctaatctggacaaagtgc

tgtccgcctacaacaagcaccgggataagcccatcagagagcaggccg

agaatatcatccacctgtttaccctgaccaatctgggagcccctgccg

ccttcaagtactttgacaccaccatcgaccggaagaggtacaccagca

ccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcc

tgtacgagacacggatcgacctgtctcagctgggaggcgacaaaaggc

cggcggccacgaaaaaggccggacaggccaaaaagaaaaagctcgagg

gcggaggcgggagcggatccccctcccggctccagatgttcttcgcta

ataaccacgaccaggaatttgaccctccaaaggtttacccacctgtcc

cagctgagaagaggaagcccatccgggtgctgtctctctttgatggaa

tcgctacagggctcctggtgctgaaggacttgggcattcaggtggacc

gctacattgcctcggaggtgtgtgaggactccatcacggtgggcatgg

tgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtca

cacagaagcatatccaggagtggggcccattcgatctggtgattgggg

gcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggcc

tctacgagggcactggccggctcttctttgagttctaccgcctcctgc

atgatgcgcggcccaaggagggagatgatcgccccttcttctggctct

ttgcgaatgtggtggccatgggcgttagtgacaagagggacatctcgc

gatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtcag

ctgcacacagggcccgctacttctggggtaaccttcccggtatgaaca

ggccgttggcatccactgtgaatgataagctggagctgcaggagtgtc

tggagcatggcaggatagccaagttcagcaaagtgaggaccattacta

cgaggtcaaactccataaagcagggcaaagaccagcattttcctgtgt

tcatgaatgagaaagaggacatcttatggtgcactgaaatggaaaggg

tatttggtttcccagtccactatactgacgtgtccaacatgagccgct

tggcgaggcagagactgctgggccggtcatggagcgtgccagtcatcc

gccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccggcc

ggcccggatccggcgcaacaaacttctctctgctgaaacaagccggag

atgtcgaagagaatcctggaccgaccgagtacaagcccacggtgcgcc

tcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgccg

cgttcgccgactaccccgccacgcgccacaccgtcgatccggaccgcc

acatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtcg

ggctcgacatcggcaaggtgtgggtcgcggacgacggcgccgcggtgg

cggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgccg

agatcggcccgcgcatggccgagttgagcggttcccggctggccgcgc

agcaacagatggaaggcctcctggcgccgcaccggcccaaggagcccg

cgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaagg

gtctgggcagcgccgtcgtgctccccggagtggaggcggccgagcgcg

ccggggtgcccgccttcctggagacctccgcgccccgcaacctcccct

tctacgagcggctcggcttcaccgtcaccgccgacgtcgaggtgcccg

aaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacgc

gttaagtcgacaatcaacctctggattacaaaatttgtgaaagattga

ctggtattcttaactatgttgctccttttacgctatgtggatacgctg

ctttaatgcctttgtatcatgctattgcttcccgtatggctttcattt

tctcctccttgtataaatcctggttgctgtctctttatgaggagttgt

ggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacg

caacccccactggttggggcattgccaccacctgtcagctcctttccg

ggactttcgctttccccctccctattgccacggcggaactcatcgccg

cctgccttgcccgctgctggacaggggctcggctgttgggcactgaca

attccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcg

cctgtgttgccacctggattctgcgcgggacgtccttctgctacgtcc

cttcggccctcaatccagcggaccttccttcccgcggcctgctgccgg

ctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcgga

tctccctttgggccgcctccccgcgtcgactttaagaccaatgactta

caaggcagctgtagatcttagccactttttaaaagaaaaggggggact

ggaagggctaattcactcccaacgaagacaagatctgctttttgcttg

tactgggtctctctggttagaccagatctgagcctgggagctctctgg

ctaactagggaacccactgcttaagcctcaataaagcttgccttgagt

gcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagag

atccctcagacccttttagtcagtgtggaaaatctctagcagggcccg

tttaaacccgctgatcagcctcgactgtgccttctagttgccagccat

ctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgcca

ctcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtc

tgagtaggtgtcattctattctggggggtggggggggcaggacagcaa

gggggaggattgggaagacaatagcaggcatgctggggatgcggtggg

ctctatggcttctgaggcggaaagaaccagctggggctctagggggta

tccccacgcgccctgtagcggcgcattaagcgcggcgggtgtggtggt

tacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctcc

tttcgctttcttcccttcctttctcgccacgttcgccggctttccccg

tcaagctctaaatcgggggctccctttagggttccgatttagtgcttt

acggcacctcgaccccaaaaaacttgattagggtgatggttcacgtag

tgggccatcgccctgatagacggtttttcgccctttgacgttggagtc

cacgttctttaatagtggactcttgttccaaactggaacaacactcaa

ccctatctcggtctattcttttgatttataagggattttgccgatttc

ggcctattggttaaaaaatgagctgatttaacaaaaatttaacgcgaa

ttaattctgtggaatgtgtgtcagttagggtgtggaaagtccccaggc

tccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagca

accaggtgtggaaagtccccaggctccccagcaggcagaagtatgcaa

agcatgcatctcaattagtcagcaaccatagtcccgcccctaactccg

cccatcccgcccctaactccgcccagttccgcccattctccgccccat

ggctgactaattttttttatttatgcagaggccgaggccgcctctgcc

tctgagctattccagaagtagtgaggaggcttttttggaggcctaggc

ttttgcaaaaagctcccgggagcttgtatatccattttcggatctgat

cagcacgtgttgacaattaatcatcggcatagtatatcggcatagtat

aatacgacaaggtgaggaactaaaccatggccaagttgaccagtgccg

ttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctgga

ccgaccggctcgggttctcccgggacttcgtggaggacgacttcgccg

gtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggacc

aggtggtgccggacaacaccctggcctgggtgtgggtgcgcggcctgg

acgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccggg

acgcctccgggccggccatgaccgagatcggcgagcagccgtgggggg

ggagttcgccctgcgcgacccggccggcaactgcgtgcacttcgtggc

cgaggagcaggactgacacgtgctacgagatttcgattccaccgccgc

cttctatgaaaggttgggcttcggaatcgttttccgggacgccggctg

gatgatcctccagcgcggggatctcatgctggagttcttcgcccaccc

caacttgtttattgcagcttataatggttacaaataaagcaatagcat

cacaaatttcacaaataaagcatttttttcactgcattctagttgtgg

tttgtccaaactcatcaatgtatcttatcatgtctgtataccgtcgac

ctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgtg

aaattgttatccgctcacaattccacacaacatacgagccggaagcat

aaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaat

tgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgcca

gctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtat

tgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgt

tcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggtt

atccacagaatcaggggataacgcaggaaagaacatgtgagcaaaagg

ccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgttttt

ccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaag

tcagaggtggcgaaacccgacaggactataaagataccaggcgtttcc

ccctggaagctccctcgtgcgctctcctgttccgaccctgccgcttac

cggatacctgtccgcctttctcccttcgggaagcgtggcgctttctca

tagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaa

gctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcctt

atccggtaactatcgtcttgagtccaacccggtaagacacgacttatc

gccactggcagcagccactggtaacaggattagcagagcgaggtatgt

aggcggtgctacagagttcttgaagtggtggcctaactacggctacac

tagaagaacagtatttggtatctgcgctctgctgaagccagttacctt

cggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctgg

tagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaa

aggatctcaagaagatcctttgatcttttctacggggtctgacgctca

gtggaacgaaaactcacgttaagggattttggtcatgagattatcaaa

aaggatcttcacctagatccttttaaattaaaaatgaagttttaaatc

aatctaaagtatatatgagtaaacttggtctgacagttaccaatgctt

aatcagtgaggcacctatctcagcgatctgtctatttcgttcatccat

agttgcctgactccccgtcgtgtagataactacgatacgggagggctt

accatctggccccagtgctgcaatgataccgcgagacccacgctcacc

ggctccagatttatcagcaataaaccagccagccggaagggccgagcg

cagaagtggtcctgcaactttatccgcctccatccagtctattaattg

ttgccgggaagctagagtaagtagttcgccagttaatagtttgcgcaa

cgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttgg

tatggcttcattcagctccggttcccaacgatcaaggcgagttacatg

atcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgat

cgttgtcagaagtaagttggccgcagtgttatcactcatggttatggc

agcactgcataattctcttactgtcatgccatccgtaagatgcttttc

tgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcg

gcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgcc

acatagcagaactttaaaagtgctcatcattggaaaacgttcttcggg

gcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgta

acccactcgtgcacccaactgatcttcagcatcttttactttcaccag

cgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaaggg

aataagggcgacacggaaatgttgaatactcatactcttcctttttca

atattattgaagcatttatcagggttattgtctcatgagcggatacat

atttgaatgtatttagaaaaataaacaaataggggttccgcgcacatt

tccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1033 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 36)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggggagtatttacggtaaactgcccacttggcagtacatcaagt

gtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatg

gcccgcctggcattatgcccagtacatgaccttatgggactttcctac

ttggcagtacatctacgtattagtcatcgctattaccatggtgatgcg

gttttggcagtacatcaatgggcgtggatagcggtttgactcacgggg

atttccaagtctccaccccattgacgtcaatgggagtttgttttggca

ccaaaatcaacgggactttccaaaatgtcgtaacaactccgccccatt

gacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcag

cgcgttttgcctgtactgggtctctctggttagaccagatctgagcct

gggagctctctggctaactagggaacccactgcttaagcctcaataaa

gcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgact

ctggtaactagagatccctcagacccttttagtcagtgtggaaaatct

ctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccaga

ggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaag

aggcgaggggcggcgactggtgagtacgccaaaaattttgactagcgg

aggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaa

gaaaaaatataaattaaaacatatagtatgggcaagcagggagctaga

acgattcgcagttaatcctggcctgttagaaacatcagaaggctgtag

acaaatactgggacagctacaaccatcccttcagacaggatcagaaga

acttagatcattatataatacagtagcaaccctctattgtgtgcatca

aaggatagagataaaagacaccaaggaagctttagacaagatagagga

agagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggcggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

ggttggagcttagaatgtgaagttttagagctagaaatagcaagttaa

aataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtg

cttttttgaattcgctagctaggtcttgaaaggagtgggaattggctc

cggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaa

gttggggggaggggtcggcaattgatccggtgcctagagaaggtggcg

cggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttcc

cgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgtt

ctttttcgcaacgggtttgccgccagaacacaggaccggtgccaccat

ggactataaggaccacgacggagactacaaggatcatgatattgatta

caaagacgatgacgataagatggccccaaagaagaagcggaaggtcgg

tatccacggagtcccagcagccgacaagaagtacagcatcggcctggc

catcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaa

ggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacag

catcaagaagaacctgatcggagccctgctgttcgacagcggcgaaac

agccgaggccacccggctgaagagaaccgccagaagaagatacaccag

acggaagaaccggatctgctatctgcaagagatcttcagcaacgagat

ggccaaggtggacgacagcttcttccacagactggaagagtccttcct

ggtggaagaggataagaagcacgagcggcaccccatcttcggcaacat

cgtggacgaggtggcctaccacgagaagtaccccaccatctaccacct

gagaaagaaactggtggacagcaccgacaaggccgacctgcggctgat

ctatctggccctggcccacatgatcaagttccggggccacttcctgat

cgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcat

ccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaa

cgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaa

gagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaa

gaatggcctgttcggcaacctgattgccctgagcctgggcctgacccc

caacttcaagagcaacttcgacctggccgaggatgccaaactgcagct

gagcaaggacacctacgacgacgacctggacaacctgctggcccagat

cggcgaccagtacgccgacctgtttctggccgccaagaacctgtccga

cgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaa

ggcccccctgagcgcctctatgatcaagagatacgacgagcaccacca

ggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaa

gtacaaagagattttcttcgaccagagcaagaacggctacgccggcta

cattgacggcggagccagccaggaagagttctacaagttcatcaagcc

catcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaa

cagagaggacctgctgcggaagcagcggaccttcgacaacggcagcat

cccccaccagatccacctgggagagctgcacgccattctgcggcggca

ggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaa

gatcctgaccttccgcatcccctactacgtgggccctctggccagggg

aaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcac

cccctggaacttcgaggaagtggtggacaagggcgcttccgcccagag

cttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaa

ggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataa

cgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgc

cttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaa

gaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcg

gttcaacgcctccctgggcacataccacgatctgctgaaaattatcaa

ggacaaggacttcctggacaatgaggaaaacgaggacattctggaaga

tatcgtgctgaccctgacactgtttgaggacagagagatgatcgagga

acggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagca

gctgaagcggcggagatacaccggctggggcaggctgagccggaagct

gatcaacggcatccgggacaagcagtccggcaagacaatcctggattt

cctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcca

cgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtc

cggccagggcgatagcctgcacgagcacattgccaatctggccggcag

ccccgccattaagaagggcatcctgcagacagtgaaggtggtggacga

gctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcga

aatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcca

gatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaa

gctgtacctgtactacctgcagaatggggggatatgtacgtggaccag

gaactggacatcaaccggctgtccgactacgatgtggacgctatcgtg

cctcagagctttctgaaggacgactccatcgacaacaaggtgctgacc

agaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagag

gtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaag

ctgattacccagagaaagttcgacaatctgaccaaggccgagagaggc

ggcctgagcgaactggataaggccggcttcatcaagagacagctggtg

gaaacccggcagatcacaaagcacgtggcacagatcctggactcccgg

atgaacactaagtacgacgagaatgacaagctgatccgggaagtgaaa

gtgatcaccctgaagtccaagctggtgtccgatttccggaaggatttc

cagttttacaaagtgcgcgagatcaacaactaccaccacgcccacgac

gcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccct

aagctggaaagcgagttcgtgtacggcgactacaaggtgtacgacgtg

cggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgcc

aagtacttcttctacagcaacatcatgaactttttcaagaccgagatt

accctggccaacggcgagatccggaagcggcctctgatcgagacaaac

ggcgaaaccggggagatcgtgtgggataagggccgggattttgccacc

gtgcggaaagtgctgagcatgccccaagtgaatatcgtgaaaaagacc

gaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagagg

aacagcgataagctgatcgccagaaagaaggactgggaccctaagaag

tacggcggcttcgacagccccaccgtggcctattctgtgctggtggtg

gccaaagtggaaaagggcaagtccaagaaactgaagagtgtgaaagag

ctgctggggatcaccatcatggaaagaagcagcttcgagaagaatccc

atcgactttctggaagccaagggctacaaagaagtgaaaaaggacctg

atcatcaagctgcctaagtactccctgttcgagctggaaaacggccgg

aagagaatgctggcctctgccggcgaactgcagaagggaaacgaactg

gccctgccctccaaatatgtgaacttcctgtacctggccagccactat

gagaagctgaagggctcccccgaggataatgagcagaaacagctgttt

gtggaacagcacaagcactacctggacgagatcatcgagcagatcagc

gagttctccaagagagtgatcctggccgacgctaatctggacaaagtg

ctgtccgcctacaacaagcaccgggataagcccatcagagagcaggcc

gagaatatcatccacctgtttaccctgaccaatctgggagcccctgcc

gccttcaagtactttgacaccaccatcgaccggaagaggtacaccagc

accaaagaggtgctggacgccaccctgatccaccagagcatcaccggc

ctgtacgagacacggatcgacctgtctcagctgggaggcgacaaaagg

ccggcggccacgaaaaaggccggacaggccaaaaagaaaaagctcgag

ggcggaggcgggagcggatccccctcccggctccagatgttcttcgct

aataaccacgaccaggaatttgaccctccaaaggtttacccacctgtc

ccagctgagaagaggaagcccatccgggtgctgtctctctttgatgga

atcgctacagggctcctggtgctgaaggacttgggcattcaggtggac

cgctacattgcctcggaggtgtgtgaggactccatcacggtgggcatg

gtgcggcaccaggggaagatcatgtacgtcggggacgtccgcagcgtc

acacagaagcatatccaggagtggggcccattcgatctggtgattggg

ggcagtccctgcaatgacctctccatcgtcaaccctgctcgcaagggc

ctctacgagggcactggccggctcttctttgagttctaccgcctcctg

catgatgcgcggcccaaggagggagatgatcgccccttcttctggctc

tttgcgaatgtggtggccatgggcgttagtgacaagagggacatctcg

cgatttctcgagtccaaccctgtgatgattgatgccaaagaagtgtca

gctgcacacagggcccgctacttctggggtaaccttcccggtatgaac

aggccgttggcatccactgtgaatgataagctggagctgcaggagtgt

ctggagcatggcaggatagccaagttcagcaaagtgaggaccattact

acgaggtcaaactccataaagcagggcaaagaccagcattttcctgtg

ttcatgaatgagaaagaggacatcttatggtgcactgaaatggaaagg

gtatttggtttcccagtccactatactgacgtgtccaacatgagccgc

ttggcgaggcagagactgctgggccggtcatggagcgtgccagtcatc

cgccacctcttcgctccgctgaaggagtattttgcgtgtgtgtccggc

cggcccggatccggcgcaacaaacttctctctgctgaaacaagccgga

gatgtcgaagagaatcctggaccgaccgagtacaagcccacggtgcgc

ctcgccacccgcgacgacgtccccagggccgtacgcaccctcgccgcc

gcgttcgccgactaccccgccacgcgccacaccgtcgatccggaccgc

cacatcgagcgggtcaccgagctgcaagaactcttcctcacgcgcgtc

gggctcgacatcggcaaggtgtgggtcgcggacgacggcgccgcggtg

gcggtctggaccacgccggagagcgtcgaagcgggggcggtgttcgcc

gagatcggcccgcgcatggccgagttgagcggttcccggctggccgcg

cagcaacagatggaaggcctcctggcgccgcaccggcccaaggagccc

gcgtggttcctggccaccgtcggagtctcgcccgaccaccagggcaag

ggtctgggcagcgccgtcgtgctccccggagtggaggcggccgagcgc

gccggggtgcccgccttcctggagacctccgcgccccgcaacctcccc

ttctacgagcggctcggcttcaccgtcaccgccgacgtcgaggtgccc

gaaggaccgcgcacctggtgcatgacccgcaagcccggtgcctgaacg

cgttaagtcgacaatcaacctctggattacaaaatttgtgaaagattg

actggtattcttaactatgttgctccttttacgctatgtggatacgct

gctttaatgcctttgtatcatgctattgcttcccgtatggctttcatt

ttctcctccttgtataaatcctggttgctgtctctttatgaggagttg

tggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgac

gcaacccccactggttggggcattgccaccacctgtcagctcctttcc

gggactttcgctttccccctccctattgccacggcggaactcatcgcc

gcctgccttgcccgctgctggacaggggctcggctgttgggcactgac

aattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctc

gcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtc

ccttcggccctcaatccagcggaccttccttcccgcggcctgctgccg

gctctgcggcctcttccgcgtcttcgccttcgccctcagacgagtcgg

atctccctttgggccgcctccccgcgtcgactttaagaccaatgactt

acaaggcagctgtagatcttagccactttttaaaagaaaaggggggac

tggaagggctaattcactcccaacgaagacaagatctgctttttgctt

gtactgggtctctctggttagaccagatctgagcctgggagctctctg

gctaactagggaacccactgcttaagcctcaataaagcttgccttgag

tgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactaga

gatccctcagacccttttagtcagtgtggaaaatctctagcagggccc

gtttaaacccgctgatcagcctcgactgtgccttctagttgccagcca

tctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgcc

actcccactgtcctttcctaataaaatgaggaaattgcatcgcattgt

ctgagtaggtgtcattctattctggggggtggggggggcaggacagca

agggggaggattgggaagacaatagcaggcatgctggggatgcggtgg

gctctatggcttctgaggcggaaagaaccagctggggctctagggggt

atccccacgcgccctgtagcggcgcattaagcgcggcgggtgtggtgg

ttacgcgcagcgtgaccgctacacttgccagcgccctagcgcccgctc

ctttcgctttcttcccttcctttctcgccacgttcgccggctttcccc

gtcaagctctaaatcgggggctccctttagggttccgatttagtgctt

tacggcacctcgaccccaaaaaacttgattagggtgatggttcacgta

gtgggccatcgccctgatagacggtttttcgccctttgacgttggagt

ccacgttctttaatagtggactcttgttccaaactggaacaacactca

accctatctcggtctattcttttgatttataagggattttgccgattt

cggcctattggttaaaaaatgagctgatttaacaaaaatttaacgcga

attaattctgtggaatgtgtgtcagttagggtgtggaaagtccccagg

ctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagc

aaccaggtgtggaaagtccccaggctccccagcaggcagaagtatgca

aagcatgcatctcaattagtcagcaaccatagtcccgcccctaactcc

gcccatcccgcccctaactccgcccagttccgcccattctccgcccca

tggctgactaattttttttatttatgcagaggccgaggccgcctctgc

ctctgagctattccagaagtagtgaggaggcttttttggaggcctagg

cttttgcaaaaagctcccgggagcttgtatatccattttcggatctga

tcagcacgtgttgacaattaatcatcggcatagtatatcggcatagta

taatacgacaaggtgaggaactaaaccatggccaagttgaccagtgcc

gttccggtgctcaccgcgcgcgacgtcgccggagcggtcgagttctgg

accgaccggctcgggttctcccgggacttcgtggaggacgacttcgcc

ggtgtggtccgggacgacgtgaccctgttcatcagcgcggtccaggac

caggtggtgccggacaacaccctggcctgggtgtgggtgcgcggcctg

gacgagctgtacgccgagtggtcggaggtcgtgtccacgaacttccgg

gacgcctccgggccggccatgaccgagatcggcgagcagccgtggggg

gggagttcgccctgcgcgacccggccggcaactgcgtgcacttcgtgg

ccgaggagcaggactgacacgtgctacgagatttcgattccaccgccg

ccttctatgaaaggttgggcttcggaatcgttttccgggacgccggct

ggatgatcctccagcgcggggatctcatgctggagttcttcgcccacc

ccaacttgtttattgcagcttataatggttacaaataaagcaatagca

tcacaaatttcacaaataaagcatttttttcactgcattctagttgtg

gtttgtccaaactcatcaatgtatcttatcatgtctgtataccgtcga

cctctagctagagcttggcgtaatcatggtcatagctgtttcctgtgt

gaaattgttatccgctcacaattccacacaacatacgagccggaagca

taaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaa

ttgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgcc

agctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgta

ttgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcg

ttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggt

tatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaag

gccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttt

tccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaa

gtcagaggtggcgaaacccgacaggactataaagataccaggcgtttc

cccctggaagctccctcgtgcgctctcctgttccgaccctgccgctta

ccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctc

atagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctcca

agctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgcct

tatccggtaactatcgtcttgagtccaacccggtaagacacgacttat

cgccactggcagcagccactggtaacaggattagcagagcgaggtatg

taggcggtgctacagagttcttgaagtggtggcctaactacggctaca

ctagaagaacagtatttggtatctgcgctctgctgaagccagttacct

tcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctg

gtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaa

aaggatctcaagaagatcctttgatcttttctacggggtctgacgctc

agtggaacgaaaactcacgttaagggattttggtcatgagattatcaa

aaaggatcttcacctagatccttttaaattaaaaatgaagttttaaat

caatctaaagtatatatgagtaaacttggtctgacagttaccaatgct

taatcagtgaggcacctatctcagcgatctgtctatttcgttcatcca

tagttgcctgactccccgtcgtgtagataactacgatacgggagggct

taccatctggccccagtgctgcaatgataccgcgagacccacgctcac

cggctccagatttatcagcaataaaccagccagccggaagggccgagc

gcagaagtggtcctgcaactttatccgcctccatccagtctattaatt

gttgccgggaagctagagtaagtagttcgccagttaatagtttgcgca

acgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttg

gtatggcttcattcagctccggttcccaacgatcaaggcgagttacat

gatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccga

tcgttgtcagaagtaagttggccgcagtgttatcactcatggttatgg

cagcactgcataattctcttactgtcatgccatccgtaagatgctttt

ctgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgc

ggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgc

cacatagcagaactttaaaagtgctcatcattggaaaacgttcttcgg

ggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgt

aacccactcgtgcacccaactgatcttcagcatcttttactttcacca

gcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagg

gaataagggcgacacggaaatgttgaatactcatactcttcctttttc

aatattattgaagcatttatcagggttattgtctcatgagcggataca

tatttgaatgtatttagaaaaataaacaaataggggttccgcgcacat

ttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1105 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 43)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cggccctatccctgggggaggggttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

ccccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatc

cccctcccggctccagatgttcttcgctaataaccacgaccaggaatt

tgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcc

catccgggtgctgtctctctttgatggaatcgctacagggctcctggt

gctgaaggacttgggcattcaggtggaccgctacattgcctcggaggt

gtgtgaggactccatcacggtgggcatggtgcggcaccaggggaagat

catgtacgtcggggacgtccgcagcgtcacacagaagcatatccagga

gtggggcccattcgatctggtgattgggggcagtccctgcaatgacct

ctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccg

gctcttctttgagttctaccgcctcctgcatgatgcgcggcccaagga

gggagatgatcgccccttcttctggctctttgagaatgtggtggccat

gggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccc

tgtgatgattgatgccaaagaagtgtcagctgcacacagggcccgcta

cttctggggtaaccttcccggtatgaacaggccgttggcatccactgt

gaatgataagctggagctgcaggagtgtctggagcatggcaggatagc

caagttcagcaaagtgaggaccattactacgaggtcaaactccataaa

gcagggcaaagaccagcattttcctgtgttcatgaatgagaaagagga

catcttatggtgcactgaaatggaaagggtatttggtttcccagtcca

ctatactgacgtgtccaacatgagccgcttggcgaggcagagactgct

gggccggtcatggagcgtgccagtcatccgccacctcttcgctccgct

gaaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaac

aaacttctctctgctgaaacaagccggagatgtcgaagagaatcctgg

accgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgt

ccccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgc

cacgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccga

gctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggt

gtgggtcgcggacgacggcgccgcggtggcggtctggaccacgccgga

gagcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggc

cgagttgagcggttcccggctggccgcgcagcaacagatggaaggcct

cctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgt

cggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgt

gctccccggagtggaggcggccgagcgcgccggggtgcccgccttcct

ggagacctccgcgccccgcaacctccccttctacgagcggctcggctt

caccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtg

catgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacc

tctggattacaaaatttgtgaaagattgactggtattcttaactatgt

tgctccttttacgctatgtggatacgctgctttaatgcctttgtatca

tgctattgcttcccgtatggctttcattttctcctccttgtataaatc

ctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg

tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttgggg

cattgccaccacctgtcagctcctttccgggactttcgctttccccct

ccctattgccacggggaactcatcgccgcctgccttgcccgctgctgg

acaggggctcggctgttgggcactgacaattccgtggtgttgtcgggg

aaatcatcgtcctttccttggctgctcgcctgtgttgccacctggatt

ctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcg

gaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgt

cttcgccttcgccctcagacgagtcggatctccctttgggccgcctcc

ccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctta

gccactttttaaaagaaaaggggggactggaagggctaattcactccc

aacgaagacaagatctgctttttgcttgtactgggtctctctggttag

accagatctgagcctgggagctctctggctaactagggaacccactgc

ttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgccc

gtctgttgtgtgactctggtaactagagatccctcagacccttttagt

cagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcc

tcgactgtgccttctagttgccagccatctgttgtttgcccctccccc

gtgccttccttgaccctggaaggtgccactcccactgtcctttcctaa

taaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctatt

ctggggggtggggggggcaggacagcaagggggaggattgggaagaca

atagcaggcatgctggggatgcggtgggctctatggcttctgaggcgg

aaagaaccagctggggctctagggggtatccccacgcgccctgtagcg

gcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgcta

cacttgccagcgccctagcgcccgctcctttcgctttcttcccttcct

ttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggc

tccctttagggttccgatttagtgctttacggcacctcgaccccaaaa

aacttgattagggtgatggttcacgtagtgggccatcgccctgataga

cggtttttcgccctttgacgttggagtccacgttctttaatagtggac

tcttgttccaaactggaacaacactcaaccctatctcggtctattctt

ttgatttataagggattttgccgatttcggcctattggttaaaaaatg

agctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtg

tcagttagggtgtggaaagtccccaggctccccagcaggcagaagtat

gcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccc

aggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtc

agcaaccatagtcccgcccctaactccgcccatcccgcccctaactcc

gcccagttccgcccattctccgccccatggctgactaattttttttat

ttatgcagaggccgaggccgcctctgcctctgagctattccagaagta

gtgaggaggcttttttggaggcctaggcttttgcaaaaagctccctac

cgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttc

ctgtgtgaaattgttatccgctcacaattccacacaacatacgagccg

gaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactca

cattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgt

cgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtt

tgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgct

cggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaa

tacggttatccacagaatcaggggataacgcaggaaagaacatgtgag

caaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgac

gctcaagtcagaggtggcgaaacccgacaggactataaagataccagg

cgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgc

cgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgc

tttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttc

gctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgct

gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcga

ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacg

gctacactagaagaacagtatttggtatctgcgctctgctgaagccag

ttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgca

gaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagat

tatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagtt

ttaaatcaatctaaagtatatatgagtaaacttggtctgacagttacc

aatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt

catccatagttgcctgactccccgtcgtgtagataactacgatacggg

agggcttaccatctggccccagtgctgcaatgataccgcgagacccac

gctcaccggctccagatttatcagcaataaaccagccagccggaaggg

ccgagcgcagaagtggtcctgcaactttatccgcctccatccagtcta

ttaattgttgccgggaagctagagtaagtagttcgccagttaatagtt

tgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgt

cgtttggtatggcttcattcagctccggttcccaacgatcaaggcgag

ttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtc

ctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatgg

ttatggcagcactgcataattctcttactgtcatgccatccgtaagat

gcttttctgtgactggtgagtactcaaccaagtcattctgagaatagt

gtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataata

ccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgtt

cttcggggcgaaaactctcaaggatcttaccgctgttgagatccagtt

cgatgtaacccactcgtgcacccaactgatcttcagcatcttttactt

tcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaa

aaaagggaataagggcgacacggaaatgttgaatactcatactcttcc

tttttcaatattattgaagcatttatcagggttattgtctcatgagcg

gatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1106 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 44)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggggtaggcgtgtacggtgggaggtctatataagcag

cgcgttttgcctgtactgggtctctctggttagaccagatctgagcct

gggagctctctggctaactagggaacccactgcttaagcctcaataaa

gcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgact

ctggtaactagagatccctcagacccttttagtcagtgtggaaaatct

ctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccaga

ggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaag

aggcgaggggcggcgactggtgagtacgccaaaaattttgactagcgg

aggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaa

gaaaaaatataaattaaaacatatagtatgggcaagcagggagctaga

acgattcgcagttaatcctggcctgttagaaacatcagaaggctgtag

acaaatactgggacagctacaaccatcccttcagacaggatcagaaga

acttagatcattatataatacagtagcaaccctctattgtgtgcatca

aaggatagagataaaagacaccaaggaagctttagacaagatagagga

agagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggggaacggtaccgagggcctat

ttcccatgattccttcatatttgcatatacgatacaaggctgttagag

agataattagaattaatttgactgtaaacacaaagatattagtacaaa

atacgtgacgtagaaagtaataatttcttgggtagtttgcagttttaa

aattatgttttaaaatggactatcatatgcttaccgtaacttgaaagt

atttcgatttcttggctttatatatcttgtggaaaggacgaaacaccg

tcgggcttggggagaggagggttttagagctagaaatagcaagttaaa

ataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtgc

ttttttgaattcgctagctaggtcttgaaaggagtgggaattggctcc

ggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaag

ttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgc

ggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttccc

gaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttc

tttttcgcaacgggtttgccgccagaacacaggaccggtgccaccatg

gactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggt

atccacggagtcccagcagccgacaagaagtacagcatcggcctggcc

atcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaag

gtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacagc

atcaagaagaacctgatcggagccctgctgttcgacagcggcgaaaca

gccgaggccacccggctgaagagaaccgccagaagaagatacaccaga

cggaagaaccggatctgctatctgcaagagatcttcagcaacgagatg

gccaaggtggacgacagcttcttccacagactggaagagtccttcctg

gtggaagaggataagaagcacgagcggcaccccatcttcggcaacatc

gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctg

agaaagaaactggtggacagcaccgacaaggccgacctgcggctgatc

tatctggccctggcccacatgatcaagttccggggccacttcctgatc

gagggcgacctgaaccccgacaacagcgacgtggacaagctgttcatc

cagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaac

gccagcggcgtggacgccaaggccatcctgtctgccagactgagcaag

agcagacggctggaaaatctgatcgcccagctgcccggcgagaagaag

aatggcctgttcggcaacctgattgccctgagcctgggcctgaccccc

aacttcaagagcaacttcgacctggccgaggatgccaaactgcagctg

agcaaggacacctacgacgacgacctggacaacctgctggcccagatc

ggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgac

gccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaag

gcccccctgagcgcctctatgatcaagagatacgacgagcaccaccag

gacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaag

tacaaagagattttcttcgaccagagcaagaacggctacgccggctac

attgacggcggagccagccaggaagagttctacaagttcatcaagccc

atcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaac

agagaggacctgctgcggaagcagcggaccttcgacaacggcagcatc

ccccaccagatccacctgggagagctgcacgccattctgcggcggcag

gaagatttttacccattcctgaaggacaaccgggaaaagatcgagaag

atcctgaccttccgcatcccctactacgtgggccctctggccagggga

aacagcagattcgcctggatgaccagaaagagcgaggaaaccatcacc

ccctggaacttcgaggaagtggtggacaagggcgcttccgcccagagc

ttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaag

gtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataac

gagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgcc

ttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaag

accaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaag

aaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcgg

ttcaacgcctccctgggcacataccacgatctgctgaaaattatcaag

gacaaggacttcctggacaatgaggaaaacgaggacattctggaagat

atcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaa

cggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcag

ctgaagcggcggagatacaccggctggggcaggctgagccggaagctg

atcaacggcatccgggacaagcagtccggcaagacaatcctggatttc

ctgaagtccgacggcttcgccaacagaaacttcatgcagctgatccac

gacgacagcctgacctttaaagaggacatccagaaagcccaggtgtcc

ggccagggcgatagcctgcacgagcacattgccaatctggccggcagc

cccgccattaagaagggcatcctgcagacagtgaaggtggtggacgag

ctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcgaa

atggccagagagaaccagaccacccagaagggacagaagaacagccgc

gagagaatgaagcggatcgaagagggcatcaaagagctgggcagccag

atcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaag

ctgtacctgtactacctgcagaatggggggatatgtacgtggaccagg

aactggacatcaaccggctgtccgactacgatgtggacgctatcgtgc

ctcagagctttctgaaggacgactccatcgacaacaaggtgctgacca

gaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagagg

tcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagc

tgattacccagagaaagttcgacaatctgaccaaggccgagagaggcg

gcctgagcgaactggataaggccggcttcatcaagagacagctggtgg

aaacccggcagatcacaaagcacgtggcacagatactagattcccgaa

tgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaag

taatcactttaaagtcaaaattggtgtcggacttcagaaaggattttc

aattctataaagttagggagataaataactaccaccatgcgcacgacg

cttatcttaatgccgtcgtagggaccgcactcattaagaaatacccga

agctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtcc

gtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcca

aatacttcttttattctaacattatgaatttctttaagacggaaatca

ctctggcaaacggagagatacgcaaacgacctttaattgaaaccaatg

gggagacaggtgaaatcgtatgggataagggccgggacttcgcgacgg

tgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaactg

aggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagga

atagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaagt

acggtggcttcgtgagccctacagttgcctattctgtcctagtagtgg

caaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaat

tattggggataacgattatggagcgctcgtcttttgaaaagaacccca

tcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctca

taattaaactaccaaagtatagtctgtttgagttagaaaatggccgaa

aacggatgttggctagcgccagagagcttcaaaaggggaacgaactcg

cactaccgtctaaatacgtgaatttcctgtatttagcgtcccattacg

agaagttgaaaggttcacctgaagataacgaacagaagcaactttttg

ttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcgg

aattcagtaagagagtcatcctagctgatgccaatctggacaaagtat

taagcgcatacaacaagcacagggataaacccatacgtgagcaggcgg

aaaatattatccatttgtttactcttaccaacctcggcgctccagccg

cattcaagtattttgacacaacgatagatcgcaaagagtacagatcta

ccaaggaggtgctagacgcgacactgattcaccaatccatcacgggat

tatatgaaactcggatagatttgtcacagcttgggggtgacggatccc

ccaagaagaagaggaaagtcctcgagggcggagggggagcggatcccc

ctcccggctccagatgttcttcgctaataaccacgaccaggaatttga

ccctccaaaggtttacccacctgtcccagctgagaagaggaagcccat

ccgggtgctgtctctctttgatggaatcgctacagggctcctggtgct

gaaggacttgggcattcaggtggaccgctacattgcctcggaggtgtg

tgaggactccatcacggtgggcatggtgcggcaccaggggaagatcat

gtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagtg

gggcccattcgatctggtgattgggggcagtccctgcaatgacctctc

catcgtcaaccctgctcgcaagggcctctacgagggcactggccggct

cttctttgagttctaccgcctcctgcatgatgcgcggcccaaggaggg

agatgatcgccccttcttctggctctttgagaatgtggtggccatggg

cgttagtgacaagagggacatctcgcgatttctcgagtccaaccctgt

gatgattgatgccaaagaagtgtcagctgcacacagggcccgctactt

ctggggtaaccttcccggtatgaacaggccgttggcatccactgtgaa

tgataagctggagctgcaggagtgtctggagcatggcaggatagccaa

gttcagcaaagtgaggaccattactacgaggtcaaactccataaagca

gggcaaagaccagcattttcctgtgttcatgaatgagaaagaggacat

cttatggtgcactgaaatggaaagggtatttggtttcccagtccacta

tactgacgtgtccaacatgagccgcttggcgaggcagagactgctggg

ccggtcatggagcgtgccagtcatccgccacctcttcgctccgctgaa

ggagtattttgcgtgtgtgtccggccggcccggatccggcgcaacaaa

cttctctctgctgaaacaagccggagatgtcgaagagaatcctggacc

gaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtccc

cagggccgtacgcaccctcgccgccgcgttcgccgactaccccgccac

gcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgagct

gcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgtg

ggtcgcggacgacggcgccgcggtggcggtctggaccacgccggagag

cgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggccga

gttgagcggttcccggctggccgcgcagcaacagatggaaggcctcct

ggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtcgg

agtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgct

ccccggagtggaggcggccgagcgcgccggggtgcccgccttcctgga

gacctccgcgccccgcaacctccccttctacgagcggctcggcttcac

cgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgcat

gacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctct

ggattacaaaatttgtgaaagattgactggtattcttaactatgttgc

tccttttacgctatgtggatacgctgctttaatgcctttgtatcatgc

tattgcttcccgtatggctttcattttctcctccttgtataaatcctg

gttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtgg

cgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcat

tgccaccacctgtcagctcctttccgggactttcgctttccccctccc

tattgccacggcggaactcatcgccgcctgccttgcccgctgctggac

aggggctcggctgttgggcactgacaattccgtggtgttgtcggggaa

atcatcgtcctttccttggctgctcgcctgtgttgccacctggattct

gcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgga

ccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtct

tcgccttcgccctcagacgagtcggatctccctttgggccgcctcccc

gcgtcgactttaagaccaatgacttacaaggcagctgtagatcttagc

cactttttaaaagaaaagggggactggaagggctaattcactcccaac

gaagacaagatctgctttttgcttgtactgggtctctctggttagacc

agatctgagcctgggagctctctggctaactagggaacccactgctta

agcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtc

tgttgtgtgactctggtaactagagatccctcagacccttttagtcag

tgtggaaaatctctagcagggcccgtttaaacccgctgatcagcctcg

actgtgccttctagttgccagccatctgttgtttgcccctcccccgtg

ccttccttgaccctggaaggtgccactcccactgtcctttcctaataa

aatgaggaaattgcatcgcattgtctgagtaggtgtcattctattctg

gggggtggggggggcaggacagcaagggggaggattgggaagacaata

gcaggcatgctggggatgcggtgggctctatggcttctgaggcggaaa

gaaccagctggggctctagggggtatccccacgcgccctgtagcggcg

cattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgctacac

ttgccagcgccctagcgcccgctcctttcgctttcttcccttcctttc

tcgccacgttcgccggctttccccgtcaagctctaaatcgggggctcc

ctttagggttccgatttagtgctttacggcacctcgaccccaaaaaac

ttgattagggtgatggttcacgtagtgggccatcgccctgatagacgg

tttttcgccctttgacgttggagtccacgttctttaatagtggactct

tgttccaaactggaacaacactcaaccctatctcggtctattcttttg

atttataagggattttgccgatttcggcctattggttaaaaaatgagc

tgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtgtca

gttagggtgtggaaagtccccaggctccccagcaggcagaagtatgca

aagcatgcatctcaattagtcagcaaccaggtgtggaaagtccccagg

ctccccagcaggcagaagtatgcaaagcatgcatctcaattagtcagc

aaccatagtcccgcccctaactccgcccatcccgcccctaactccgcc

cagttccgcccattctccgccccatggctgactaattttttttattta

tgcagaggccgaggccgcctctgcctctgagctattccagaagtagtg

aggaggcttttttggaggcctaggcttttgcaaaaagctccctaccgt

cgacctctagctagagcttggcgtaatcatggtcatagctgtttcctg

tgtgaaattgttatccgctcacaattccacacaacatacgagccggaa

gcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacat

taattgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgt

gccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgc

gtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcgg

tcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatac

ggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaa

aaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgt

ttttccataggctccgcccccctgacgagcatcacaaaaatcgacgct

caagtcagaggtggcgaaacccgacaggactataaagataccaggcgt

ttccccctggaagctccctcgtgcgctctcctgttccgaccctgccgc

ttaccggatacctgtccgcctttctcccttcgggaagcgtggcgcttt

ctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcgct

ccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcg

ccttatccggtaactatcgtcttgagtccaacccggtaagacacgact

tatcgccactggcagcagccactggtaacaggattagcagagcgaggt

atgtaggcggtgctacagagttcttgaagtggtggcctaactacggct

acactagaagaacagtatttggtatctgcgctctgctgaagccagtta

ccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccg

ctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaa

aaaaaggatctcaagaagatcctttgatcttttctacggggtctgacg

ctcagtggaacgaaaactcacgttaagggattttggtcatgagattat

caaaaaggatcttcacctagatccttttaaattaaaaatgaagtttta

aatcaatctaaagtatatatgagtaaacttggtctgacagttaccaat

gcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcat

ccatagttgcctgactccccgtcgtgtagataactacgatacgggagg

gcttaccatctggccccagtgctgcaatgataccgcgagacccacgct

caccggctccagatttatcagcaataaaccagccagccggaagggccg

agcgcagaagtggtcctgcaactttatccgcctccatccagtctatta

attgttgccgggaagctagagtaagtagttcgccagttaatagtttgc

gcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgt

ttggtatggcttcattcagctccggttcccaacgatcaaggcgagtta

catgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctc

cgatcgttgtcagaagtaagttggccgcagtgttatcactcatggtta

tggcagcactgcataattctcttactgtcatgccatccgtaagatgct

tttctgtgactggtgagtactcaaccaagtcattctgagaatagtgta

tgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccg

cgccacatagcagaactttaaaagtgctcatcattggaaaacgttctt

cggggcgaaaactctcaaggatcttaccgctgttgagatccagttcga

tgtaacccactcgtgcacccaactgatcttcagcatcttttactttca

ccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaa

agggaataagggcgacacggaaatgttgaatactcatactcttccttt

ttcaatattattgaagcatttatcagggttattgtctcatgagcggat

acatatttgaatgtatttagaaaaataaacaaataggggttccgcgca

catttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1107 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 45)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggggtaggcgtgtacggtgggaggtctatataagcag

cgcgttttgcctgtactgggtctctctggttagaccagatctgagcct

gggagctctctggctaactagggaacccactgcttaagcctcaataaa

gcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgact

ctggtaactagagatccctcagacccttttagtcagtgtggaaaatct

ctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccaga

ggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaag

aggcgaggggcggcgactggtgagtacgccaaaaattttgactagcgg

aggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaa

gaaaaaatataaattaaaacatatagtatgggcaagcagggagctaga

acgattcgcagttaatcctggcctgttagaaacatcagaaggctgtag

acaaatactgggacagctacaaccatcccttcagacaggatcagaaga

acttagatcattatataatacagtagcaaccctctattgtgtgcatca

aaggatagagataaaagacaccaaggaagctttagacaagatagagga

agagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggcggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

gctctccccaccccaccttctgttttagagctagaaatagcaagttaa

aataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtg

cttttttgaattcgctagctaggtcttgaaaggagtgggaattggctc

cggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaa

gttggggggaggggtcggcaattgatccggtgcctagagaaggtggcg

cggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttcc

cgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgtt

ctttttcgcaacgggtttgccgccagaacacaggaccggtgccaccat

ggactataaggaccacgacggagactacaaggatcatgatattgatta

caaagacgatgacgataagatggccccaaagaagaagcggaaggtcgg

tatccacggagtcccagcagccgacaagaagtacagcatcggcctggc

catcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaa

ggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacag

catcaagaagaacctgatcggagccctgctgttcgacagcggcgaaac

agccgaggccacccggctgaagagaaccgccagaagaagatacaccag

acggaagaaccggatctgctatctgcaagagatcttcagcaacgagat

ggccaaggtggacgacagcttcttccacagactggaagagtccttcct

ggtggaagaggataagaagcacgagcggcaccccatcttcggcaacat

cgtggacgaggtggcctaccacgagaagtaccccaccatctaccacct

gagaaagaaactggtggacagcaccgacaaggccgacctgcggctgat

ctatctggccctggcccacatgatcaagttccggggccacttcctgat

cgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcat

ccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaa

cgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaa

gagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaa

gaatggcctgttcggcaacctgattgccctgagcctgggcctgacccc

caacttcaagagcaacttcgacctggccgaggatgccaaactgcagct

gagcaaggacacctacgacgacgacctggacaacctgctggcccagat

cggcgaccagtacgccgacctgtttctggccgccaagaacctgtccga

cgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaa

ggcccccctgagcgcctctatgatcaagagatacgacgagcaccacca

ggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaa

gtacaaagagattttcttcgaccagagcaagaacggctacgccggcta

cattgacggcggagccagccaggaagagttctacaagttcatcaagcc

catcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaa

cagagaggacctgctgcggaagcagcggaccttcgacaacggcagcat

cccccaccagatccacctgggagagctgcacgccattctgcggcggca

ggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaa

gatcctgaccttccgcatcccctactacgtgggccctctggccagggg

aaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcac

cccctggaacttcgaggaagtggtggacaagggcgcttccgcccagag

cttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaa

ggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataa

cgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgc

cttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaa

gaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcg

gttcaacgcctccctgggcacataccacgatctgctgaaaattatcaa

ggacaaggacttcctggacaatgaggaaaacgaggacattctggaaga

tatcgtgctgaccctgacactgtttgaggacagagagatgatcgagga

acggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagca

gctgaagcggcggagatacaccggctggggcaggctgagccggaagct

gatcaacggcatccgggacaagcagtccggcaagacaatcctggattt

cctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcca

cgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtc

cggccagggcgatagcctgcacgagcacattgccaatctggccggcag

ccccgccattaagaagggcatcctgcagacagtgaaggtggtggacga

gctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcga

aatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcca

gatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaa

gctgtacctgtactacctgcagaatggggggatatgtacgtggaccag

gaactggacatcaaccggctgtccgactacgatgtggacgctatcgtg

cctcagagctttctgaaggacgactccatcgacaacaaggtgctgacc

agaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagag

gtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaag

ctgattacccagagaaagttcgacaatctgaccaaggccgagagaggc

ggcctgagcgaactggataaggccggcttcatcaagagacagctggtg

gaaacccggcagatcacaaagcacgtggcacagatactagattcccga

atgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaa

gtaatcactttaaagtcaaaattggtgtcggacttcagaaaggatttt

caattctataaagttagggagataaataactaccaccatgcgcacgac

gcttatcttaatgccgtcgtagggaccgcactcattaagaaatacccg

aagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtc

cgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcc

aaatacttcttttattctaacattatgaatttctttaagacggaaatc

actctggcaaacggagagatacgcaaacgacctttaattgaaaccaat

ggggagacaggtgaaatcgtatgggataagggccgggacttcgcgacg

gtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaact

gaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagg

aatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaag

tacggtggcttcgtgagccctacagttgcctattctgtcctagtagtg

gcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaa

ttattggggataacgattatggagcgctcgtcttttgaaaagaacccc

atcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctc

ataattaaactaccaaagtatagtctgtttgagttagaaaatggccga

aaacggatgttggctagcgccagagagcttcaaaaggggaacgaactc

gcactaccgtctaaatacgtgaatttcctgtatttagcgtcccattac

gagaagttgaaaggttcacctgaagataacgaacagaagcaacttttt

gttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcg

gaattcagtaagagagtcatcctagctgatgccaatctggacaaagta

ttaagcgcatacaacaagcacagggataaacccatacgtgagcaggcg

gaaaatattatccatttgtttactcttaccaacctcggcgctccagcc

gcattcaagtattttgacacaacgatagatcgcaaagagtacagatct

accaaggaggtgctagacgcgacactgattcaccaatccatcacggga

ttatatgaaactcggatagatttgtcacagcttgggggtgacggatcc

cccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatcc

ccctcccggctccagatgttcttcgctaataaccacgaccaggaattt

gaccctccaaaggtttacccacctgtcccagctgagaagaggaagccc

atccgggtgctgtctctctttgatggaatcgctacagggctcctggtg

ctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtg

tgtgaggactccatcacggtgggcatggtgcggcaccaggggaagatc

atgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggag

tggggcccattcgatctggtgattgggggcagtccctgcaatgacctc

tccatcgtcaaccctgctcgcaagggcctctacgagggcactggccgg

ctcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggag

ggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccct

gtgatgattgatgccaaagaagtgtcagctgcacacagggcccgctac

ttctggggtaaccttcccggtatgaacaggccgttggcatccactgtg

aatgataagctggagctgcaggagtgtctggagcatggcaggatagcc

aagttcagcaaagtgaggaccattactacgaggtcaaactccataaag

cagggcaaagaccagcattttcctgtgttcatgaatgagaaagaggac

atcttatggtgcactgaaatggaaagggtatttggtttcccagtccac

tatactgacgtgtccaacatgagccgcttggcgaggcagagactgctg

ggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctg

aaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaaca

aacttctctctgctgaaacaagccggagatgtcgaagagaatcctgga

ccgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtc

cccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgcc

acgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgag

ctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtg

tgggtcgcggacgacggcgccgcggtggcggtctggaccacgccggag

agcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggcc

gagttgagcggttcccggctggccgcgcagcaacagatggaaggcctc

ctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtc

ggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtg

ctccccggagtggaggcggccgagcgcgccggggtgcccgccttcctg

gagacctccgcgccccgcaacctccccttctacgagcggctcggcttc

accgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgc

atgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacct

ctggattacaaaatttgtgaaagattgactggtattcttaactatgtt

gctccttttacgctatgtggatacgctgctttaatgcctttgtatcat

gctattgcttcccgtatggctttcattttctcctccttgtataaatcc

tggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgt

ggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggc

attgccaccacctgtcagctcctttccgggactttcgctttccccctc

cctattgccacggcggaactcatcgccgcctgccttgcccgctgctgg

acaggggctcggctgttgggcactgacaattccgtggtgttgtcgggg

aaatcatcgtcctttccttggctgctcgcctgtgttgccacctggatt

ctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcg

gaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgt

cttcgccttcgccctcagacgagtcggatctccctttgggccgcctcc

ccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctta

gccactttttaaaagaaaaggggggactggaagggctaattcactccc

aacgaagacaagatctgctttttgcttgtactgggtctctctggttag

accagatctgagcctgggagctctctggctaactagggaacccactgc

ttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgccc

gtctgttgtgtgactctggtaactagagatccctcagacccttttagt

cagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcc

tcgactgtgccttctagttgccagccatctgttgtttgcccctccccc

gtgccttccttgaccctggaaggtgccactcccactgtcctttcctaa

taaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctatt

ctggggggggggtggggcaggacagcaagggggaggattgggaagaca

atagcaggcatgctggggatgcggtgggctctatggcttctgaggcgg

aaagaaccagctggggctctagggggtatccccacgcgccctgtagcg

gcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgcta

cacttgccagcgccctagcgcccgctcctttcgctttcttcccttcct

ttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggc

tccctttagggttccgatttagtgctttacggcacctcgaccccaaaa

aacttgattagggtgatggttcacgtagtgggccatcgccctgataga

cggtttttcgccctttgacgttggagtccacgttctttaatagtggac

tcttgttccaaactggaacaacactcaaccctatctcggtctattctt

ttgatttataagggattttgccgatttcggcctattggttaaaaaatg

agctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtg

tcagttagggtgtggaaagtccccaggctccccagcaggcagaagtat

gcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccc

aggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtc

agcaaccatagtcccgcccctaactccgcccatcccgcccctaactcc

gcccagttccgcccattctccgccccatggctgactaattttttttat

ttatgcagaggccgaggccgcctctgcctctgagctattccagaagta

gtgaggaggcttttttggaggcctaggcttttgcaaaaagctccctac

cgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttc

ctgtgtgaaattgttatccgctcacaattccacacaacatacgagccg

gaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactca

cattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgt

cgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtt

tgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgct

cggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaa

tacggttatccacagaatcaggggataacgcaggaaagaacatgtgag

caaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgac

gctcaagtcagaggtggcgaaacccgacaggactataaagataccagg

cgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgc

cgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgc

tttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttc

gctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgct

gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcga

ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacg

gctacactagaagaacagtatttggtatctgcgctctgctgaagccag

ttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgca

gaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagat

tatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagtt

ttaaatcaatctaaagtatatatgagtaaacttggtctgacagttacc

aatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt

catccatagttgcctgactccccgtcgtgtagataactacgatacggg

agggcttaccatctggccccagtgctgcaatgataccgcgagacccac

gctcaccggctccagatttatcagcaataaaccagccagccggaaggg

ccgagcgcagaagtggtcctgcaactttatccgcctccatccagtcta

ttaattgttgccgggaagctagagtaagtagttcgccagttaatagtt

tgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgt

cgtttggtatggcttcattcagctccggttcccaacgatcaaggcgag

ttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtc

ctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatgg

ttatggcagcactgcataattctcttactgtcatgccatccgtaagat

gcttttctgtgactggtgagtactcaaccaagtcattctgagaatagt

gtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataata

ccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgtt

cttcggggcgaaaactctcaaggatcttaccgctgttgagatccagtt

cgatgtaacccactcgtgcacccaactgatcttcagcatcttttactt

tcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaa

aaaagggaataagggcgacacggaaatgttgaatactcatactcttcc

tttttcaatattattgaagcatttatcagggttattgtctcatgagcg

gatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1108 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 46)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cgtgtgaagggagaatgaggaagttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

ccccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatc

cccctcccggctccagatgttcttcgctaataaccacgaccaggaatt

tgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcc

catccgggtgctgtctctctttgatggaatcgctacagggctcctggt

gctgaaggacttgggcattcaggtggaccgctacattgcctcggaggt

gtgtgaggactccatcacggtgggcatggtgcggcaccaggggaagat

catgtacgtcggggacgtccgcagcgtcacacagaagcatatccagga

gtggggcccattcgatctggtgattgggggcagtccctgcaatgacct

ctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccg

gctcttctttgagttctaccgcctcctgcatgatgcgcggcccaagga

gggagatgatcgccccttcttctggctctttgagaatgtggtggccat

gggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccc

tgtgatgattgatgccaaagaagtgtcagctgcacacagggcccgcta

cttctggggtaaccttcccggtatgaacaggccgttggcatccactgt

gaatgataagctggagctgcaggagtgtctggagcatggcaggatagc

caagttcagcaaagtgaggaccattactacgaggtcaaactccataaa

gcagggcaaagaccagcattttcctgtgttcatgaatgagaaagagga

catcttatggtgcactgaaatggaaagggtatttggtttcccagtcca

ctatactgacgtgtccaacatgagccgcttggcgaggcagagactgct

gggccggtcatggagcgtgccagtcatccgccacctcttcgctccgct

gaaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaac

aaacttctctctgctgaaacaagccggagatgtcgaagagaatcctgg

accgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgt

ccccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgc

cacgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccga

gctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggt

gtgggtcgcggacgacggcgccgcggtggcggtctggaccacgccgga

gagcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggc

cgagttgagcggttcccggctggccgcgcagcaacagatggaaggcct

cctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgt

cggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgt

gctccccggagtggaggcggccgagcgcgccggggtgcccgccttcct

ggagacctccgcgccccgcaacctccccttctacgagcggctcggctt

caccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtg

catgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacc

tctggattacaaaatttgtgaaagattgactggtattcttaactatgt

tgctccttttacgctatgtggatacgctgctttaatgcctttgtatca

tgctattgcttcccgtatggctttcattttctcctccttgtataaatc

ctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg

tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttgggg

cattgccaccacctgtcagctcctttccgggactttcgctttccccct

ccctattgccacggcggaactcatcgccgcctgccttgcccgctgctg

gacaggggctcggctgttgggcactgacaattccgtggtgttgtcggg

gaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggat

tctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagc

ggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcg

tcttcgccttcgccctcagacgagtcggatctccctttgggccgcctc

cccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctt

agccactttttaaaagaaaaggggggactggaagggctaattcactcc

caacgaagacaagatctgctttttgcttgtactgggtctctctggtta

gaccagatctgagcctgggagctctctggctaactagggaacccactg

cttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcc

cgtctgttgtgtgactctggtaactagagatccctcagacccttttag

tcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagc

ctcgactgtgccttctagttgccagccatctgttgtttgcccctcccc

cgtgccttccttgaccctggaaggtgccactcccactgtcctttccta

ataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctat

tctggggggtggggggggcaggacagcaagggggaggattgggaagac

aatagcaggcatgctggggatgcggtgggctctatggcttctgaggcg

gaaagaaccagctggggctctagggggtatccccacgcgccctgtagc

ggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgct

acacttgccagcgccctagcgcccgctcctttcgctttcttcccttcc

tttctcgccacgttcgccggctttccccgtcaagctctaaatcggggg

ctccctttagggttccgatttagtgctttacggcacctcgaccccaaa

aaacttgattagggtgatggttcacgtagtgggccatcgccctgatag

acggtttttcgccctttgacgttggagtccacgttctttaatagtgga

ctcttgttccaaactggaacaacactcaaccctatctcggtctattct

tttgatttataagggattttgccgatttcggcctattggttaaaaaat

gagctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgt

gtcagttagggtgtggaaagtccccaggctccccagcaggcagaagta

tgcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtccc

caggctccccagcaggcagaagtatgcaaagcatgcatctcaattagt

cagcaaccatagtcccgcccctaactccgcccatcccgcccctaactc

cgcccagttccgcccattctccgccccatggctgactaatttttttta

tttatgcagaggccgaggccgcctctgcctctgagctattccagaagt

agtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccta

ccgtcgacctctagctagagcttggcgtaatcatggtcatagctgttt

cctgtgtgaaattgttatccgctcacaattccacacaacatacgagcc

ggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactc

acattaattgcgttgcgctcactgcccgctttccagtcgggaaacctg

tcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggt

ttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgc

tcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta

atacggttatccacagaatcaggggataacgcaggaaagaacatgtga

gcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctg

gcgtttttccataggctccgcccccctgacgagcatcacaaaaatcga

cgctcaagtcagaggtggcgaaacccgacaggactataaagataccag

gcgtttccccctggaagctccctcgtgcgctctcctgttccgaccctg

ccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcg

ctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgtt

cgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgc

tgcgccttatccggtaactatcgtcttgagtccaacccggtaagacac

gacttatcgccactggcagcagccactggtaacaggattagcagagcg

aggtatgtaggcggtgctacagagttcttgaagtggtggcctaactac

ggctacactagaagaacagtatttggtatctgcgctctgctgaagcca

gttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacc

accgctggtagcggtggtttttttgtttgcaagcagcagattacgcgc

agaaaaaaaggatctcaagaagatcctttgatcttttctacggggtct

gacgctcagtggaacgaaaactcacgttaagggattttggtcatgaga

ttatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagt

tttaaatcaatctaaagtatatatgagtaaacttggtctgacagttac

caatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgt

tcatccatagttgcctgactccccgtcgtgtagataactacgatacgg

gagggcttaccatctggccccagtgctgcaatgataccgcgagaccca

cgctcaccggctccagatttatcagcaataaaccagccagccggaagg

gccgagcgcagaagtggtcctgcaactttatccgcctccatccagtct

attaattgttgccgggaagctagagtaagtagttcgccagttaatagt

ttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcg

tcgtttggtatggcttcattcagctccggttcccaacgatcaaggcga

gttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggt

cctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatg

gttatggcagcactgcataattctcttactgtcatgccatccgtaaga

tgcttttctgtgactggtgagtactcaaccaagtcattctgagaatag

tgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataat

accgcgccacatagcagaactttaaaagtgctcatcattggaaaacgt

tcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagt

tcgatgtaacccactcgtgcacccaactgatcttcagcatcttttact

ttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca

aaaaagggaataagggcgacacggaaatgttgaatactcatactcttc

ctttttcaatattattgaagcatttatcagggttattgtctcatgagc

ggatacatatttgaatgtatttagaaaaataaacaaataggggttccg

cgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1109 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 47)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacgggggaggtctatataagcag

cgcgttttgcctgtactgggtctctctggttagaccagatctgagcct

gggagctctctggctaactagggaacccactgcttaagcctcaataaa

gcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgact

ctggtaactagagatccctcagacccttttagtcagtgtggaaaatct

ctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccaga

ggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaag

aggcgaggggcggcgactggtgagtacgccaaaaattttgactagcgg

aggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaa

gaaaaaatataaattaaaacatatagtatgggcaagcagggagctaga

acgattcgcagttaatcctggcctgttagaaacatcagaaggctgtag

acaaatactgggacagctacaaccatcccttcagacaggatcagaaga

acttagatcattatataatacagtagcaaccctctattgtgtgcatca

aaggatagagataaaagacaccaaggaagctttagacaagatagagga

agagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggcggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

ggccctatccctgggggaggggttttagagctagaaatagcaagttaa

aataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtg

cttttttgaattcgctagctaggtcttgaaaggagtgggaattggctc

cggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaa

gttggggggaggggtcggcaattgatccggtgcctagagaaggtggcg

cggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttcc

cgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgtt

ctttttcgcaacgggtttgccgccagaacacaggaccggtgccaccat

ggactataaggaccacgacggagactacaaggatcatgatattgatta

caaagacgatgacgataagatggccccaaagaagaagcggaaggtcgg

tatccacggagtcccagcagccgacaagaagtacagcatcggcctggc

catcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaa

ggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacag

catcaagaagaacctgatcggagccctgctgttcgacagcggcgaaac

agccgaggccacccggctgaagagaaccgccagaagaagatacaccag

acggaagaaccggatctgctatctgcaagagatcttcagcaacgagat

ggccaaggtggacgacagcttcttccacagactggaagagtccttcct

ggtggaagaggataagaagcacgagcggcaccccatcttcggcaacat

cgtggacgaggtggcctaccacgagaagtaccccaccatctaccacct

gagaaagaaactggtggacagcaccgacaaggccgacctgcggctgat

ctatctggccctggcccacatgatcaagttccggggccacttcctgat

cgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcat

ccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaa

cgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaa

gagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaa

gaatggcctgttcggcaacctgattgccctgagcctgggcctgacccc

caacttcaagagcaacttcgacctggccgaggatgccaaactgcagct

gagcaaggacacctacgacgacgacctggacaacctgctggcccagat

cggcgaccagtacgccgacctgtttctggccgccaagaacctgtccga

cgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaa

ggcccccctgagcgcctctatgatcaagagatacgacgagcaccacca

ggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaa

gtacaaagagattttcttcgaccagagcaagaacggctacgccggcta

cattgacggcggagccagccaggaagagttctacaagttcatcaagcc

catcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaa

cagagaggacctgctgcggaagcagcggaccttcgacaacggcagcat

cccccaccagatccacctgggagagctgcacgccattctgcggcggca

ggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaa

gatcctgaccttccgcatcccctactacgtgggccctctggccagggg

aaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcac

cccctggaacttcgaggaagtggtggacaagggcgcttccgcccagag

cttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaa

ggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataa

cgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgc

cttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaa

gaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcg

gttcaacgcctccctgggcacataccacgatctgctgaaaattatcaa

ggacaaggacttcctggacaatgaggaaaacgaggacattctggaaga

tatcgtgctgaccctgacactgtttgaggacagagagatgatcgagga

acggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagca

gctgaagcggcggagatacaccggctggggcaggctgagccggaagct

gatcaacggcatccgggacaagcagtccggcaagacaatcctggattt

cctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcca

cgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtc

cggccagggcgatagcctgcacgagcacattgccaatctggccggcag

ccccgccattaagaagggcatcctgcagacagtgaaggtggtggacga

gctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcga

aatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcca

gatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaa

gctgtacctgtactacctgcagaatggggggatatgtacgtggaccag

gaactggacatcaaccggctgtccgactacgatgtggacgctatcgtg

cctcagagctttctgaaggacgactccatcgacaacaaggtgctgacc

agaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagag

gtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaag

ctgattacccagagaaagttcgacaatctgaccaaggccgagagaggc

ggcctgagcgaactggataaggccggcttcatcaagagacagctggtg

gaaacccggcagatcacaaagcacgtggcacagatactagattcccga

atgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaa

gtaatcactttaaagtcaaaattggtgtcggacttcagaaaggatttt

caattctataaagttagggagataaataactaccaccatgcgcacgac

gcttatcttaatgccgtcgtagggaccgcactcattaagaaatacccg

aagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtc

cgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcc

aaatacttcttttattctaacattatgaatttctttaagacggaaatc

actctggcaaacggagagatacgcaaacgacctttaattgaaaccaat

ggggagacaggtgaaatcgtatgggataagggccgggacttcgcgacg

gtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaact

gaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagg

aatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaag

tacggtggcttcgtgagccctacagttgcctattctgtcctagtagtg

gcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaa

ttattggggataacgattatggagcgctcgtcttttgaaaagaacccc

atcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctc

ataattaaactaccaaagtatagtctgtttgagttagaaaatggccga

aaacggatgttggctagcgccagagagcttcaaaaggggaacgaactc

gcactaccgtctaaatacgtgaatttcctgtatttagcgtcccattac

gagaagttgaaaggttcacctgaagataacgaacagaagcaacttttt

gttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcg

gaattcagtaagagagtcatcctagctgatgccaatctggacaaagta

ttaagcgcatacaacaagcacagggataaacccatacgtgagcaggcg

gaaaatattatccatttgtttactcttaccaacctcggcgctccagcc

gcattcaagtattttgacacaacgatagatcgcaaagagtacagatct

accaaggaggtgctagacgcgacactgattcaccaatccatcacggga

ttatatgaaactcggatagatttgtcacagcttgggggtgacggatcc

cccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatcc

ccctcccggctccagatgttcttcgctaataaccacgaccaggaattt

gaccctccaaaggtttacccacctgtcccagctgagaagaggaagccc

atccgggtgctgtctctctttgatggaatcgctacagggctcctggtg

ctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtg

tgtgaggactccatcacggtgggcatggtgcggcaccaggggaagatc

atgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggag

tggggcccattcgatctggtgattgggggcagtccctgcaatgacctc

tccatcgtcaaccctgctcgcaagggcctctacgagggcactggccgg

ctcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggag

ggagatgatcgccccttcttctggctctttgcgaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccct

gtgatgattgatgccaaagaagtgtcagctgcacacagggcccgctac

ttctggggtaaccttcccggtatgaacaggccgttggcatccactgtg

aatgataagctggagctgcaggagtgtctggagcatggcaggatagcc

aagttcagcaaagtgaggaccattactacgaggtcaaactccataaag

cagggcaaagaccagcattttcctgtgttcatgaatgagaaagaggac

atcttatggtgcactgaaatggaaagggtatttggtttcccagtccac

tatactgacgtgtccaacatgagccgcttggcgaggcagagactgctg

ggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctg

aaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaaca

aacttctctctgctgaaacaagccggagatgtcgaagagaatcctgga

ccgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtc

cccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgcc

acgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgag

ctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtg

tgggtcgcggacgacggcgccgcggtggcggtctggaccacgccggag

agcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggcc

gagttgagcggttcccggctggccgcgcagcaacagatggaaggcctc

ctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtc

ggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtg

ctccccggagtggaggcggccgagcgcgccggggtgcccgccttcctg

gagacctccgcgccccgcaacctccccttctacgagcggctcggcttc

accgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgc

atgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacct

ctggattacaaaatttgtgaaagattgactggtattcttaactatgtt

gctccttttacgctatgtggatacgctgctttaatgcctttgtatcat

gctattgcttcccgtatggctttcattttctcctccttgtataaatcc

tggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgt

ggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggc

attgccaccacctgtcagctcctttccgggactttcgctttccccctc

cctattgccacggcggaactcatcgccgcctgccttgcccgctgctgg

acaggggctcggctgttgggcactgacaattccgtggtgttgtcgggg

aaatcatcgtcctttccttggctgctcgcctgtgttgccacctggatt

ctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcg

gaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgt

cttcgccttcgccctcagacgagtcggatctccctttgggccgcctcc

ccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctta

gccactttttaaaagaaaaggggggactggaagggctaattcactccc

aacgaagacaagatctgctttttgcttgtactgggtctctctggttag

accagatctgagcctgggagctctctggctaactagggaacccactgc

ttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgccc

gtctgttgtgtgactctggtaactagagatccctcagacccttttagt

cagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcc

tcgactgtgccttctagttgccagccatctgttgtttgcccctccccc

gtgccttccttgaccctggaaggtgccactcccactgtcctttcctaa

taaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctatt

ctggggggtggggggggcaggacagcaagggggaggattgggaagaca

atagcaggcatgctggggatgcggtgggctctatggcttctgaggcgg

aaagaaccagctggggctctagggggtatccccacgcgccctgtagcg

gcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgcta

cacttgccagcgccctagcgcccgctcctttcgctttcttcccttcct

ttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggc

tccctttagggttccgatttagtgctttacggcacctcgaccccaaaa

aacttgattagggtgatggttcacgtagtgggccatcgccctgataga

cggtttttcgccctttgacgttggagtccacgttctttaatagtggac

tcttgttccaaactggaacaacactcaaccctatctcggtctattctt

ttgatttataagggattttgccgatttcggcctattggttaaaaaatg

agctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtg

tcagttagggtgtggaaagtccccaggctccccagcaggcagaagtat

gcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccc

aggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtc

agcaaccatagtcccgcccctaactccgcccatcccgcccctaactcc

gcccagttccgcccattctccgccccatggctgactaattttttttat

ttatgcagaggccgaggccgcctctgcctctgagctattccagaagta

gtgaggaggcttttttggaggcctaggcttttgcaaaaagctccctac

cgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttc

ctgtgtgaaattgttatccgctcacaattccacacaacatacgagccg

gaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactca

cattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgt

cgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtt

tgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgct

cggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaa

tacggttatccacagaatcaggggataacgcaggaaagaacatgtgag

caaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgac

gctcaagtcagaggtggcgaaacccgacaggactataaagataccagg

cgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgc

cgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgc

tttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttc

gctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgct

gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcga

ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacg

gctacactagaagaacagtatttggtatctgcgctctgctgaagccag

ttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgca

gaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagat

tatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagtt

ttaaatcaatctaaagtatatatgagtaaacttggtctgacagttacc

aatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt

catccatagttgcctgactccccgtcgtgtagataactacgatacggg

agggcttaccatctggccccagtgctgcaatgataccgcgagacccac

gctcaccggctccagatttatcagcaataaaccagccagccggaaggg

ccgagcgcagaagtggtcctgcaactttatccgcctccatccagtcta

ttaattgttgccgggaagctagagtaagtagttcgccagttaatagtt

tgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgt

cgtttggtatggcttcattcagctccggttcccaacgatcaaggcgag

ttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtc

ctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatgg

ttatggcagcactgcataattctcttactgtcatgccatccgtaagat

gcttttctgtgactggtgagtactcaaccaagtcattctgagaatagt

gtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataata

ccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgtt

cttcggggcgaaaactctcaaggatcttaccgctgttgagatccagtt

cgatgtaacccactcgtgcacccaactgatcttcagcatcttttactt

tcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaa

aaaagggaataagggcgacacggaaatgttgaatactcatactcttcc

tttttcaatattattgaagcatttatcagggttattgtctcatgagcg

gatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1110 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 48)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cgtcgggcttggggagaggagggttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

ccccaagaagaagaggaaagtcctcgagggggaggcgggagcggatcc

ccctcccggctccagatgttcttcgctaataaccacgaccaggaattt

gaccctccaaaggtttacccacctgtcccagctgagaagaggaagccc

atccgggtgctgtctctctttgatggaatcgctacagggctcctggtg

ctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtg

tgtgaggactccatcacggtgggcatggtgcggcaccaggggaagatc

atgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggag

tggggcccattcgatctggtgattgggggcagtccctgcaatgacctc

tccatcgtcaaccctgctcgcaagggcctctacgagggcactggccgg

ctcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggag

ggagatgatcgccccttcttctggctctttgcgaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccct

gtgatgattgatgccaaagaagtgtcagctgcacacagggcccgctac

ttctggggtaaccttcccggtatgaacaggccgttggcatccactgtg

aatgataagctggagctgcaggagtgtctggagcatggcaggatagcc

aagttcagcaaagtgaggaccattactacgaggtcaaactccataaag

cagggcaaagaccagcattttcctgtgttcatgaatgagaaagaggac

atcttatggtgcactgaaatggaaagggtatttggtttcccagtccac

tatactgacgtgtccaacatgagccgcttggcgaggcagagactgctg

ggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctg

aaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaaca

aacttctctctgctgaaacaagccggagatgtcgaagagaatcctgga

ccgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtc

cccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgcc

acgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgag

ctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtg

tgggtcgcggacgacggcgccgcggtggcggtctggaccacgccggag

agcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggcc

gagttgagcggttcccggctggccgcgcagcaacagatggaaggcctc

ctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtc

ggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtg

ctccccggagtggaggcggccgagcgcgccggggtgcccgccttcctg

gagacctccgcgccccgcaacctccccttctacgagcggctcggcttc

accgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgc

atgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacct

ctggattacaaaatttgtgaaagattgactggtattcttaactatgtt

gctccttttacgctatgtggatacgctgctttaatgcctttgtatcat

gctattgcttcccgtatggctttcattttctcctccttgtataaatcc

tggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgt

ggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggc

attgccaccacctgtcagctcctttccgggactttcgctttccccctc

cctattgccacggcggaactcatcgccgcctgccttgcccgctgctgg

acaggggctcggctgttgggcactgacaattccgtggtgttgtcgggg

aaatcatcgtcctttccttggctgctcgcctgtgttgccacctggatt

ctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcg

gaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgt

cttcgccttcgccctcagacgagtcggatctccctttgggccgcctcc

ccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctta

gccactttttaaaagaaaaggggggactggaagggctaattcactccc

aacgaagacaagatctgctttttgcttgtactgggtctctctggttag

accagatctgagcctgggagctctctggctaactagggaacccactgc

ttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgccc

gtctgttgtgtgactctggtaactagagatccctcagacccttttagt

cagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcc

tcgactgtgccttctagttgccagccatctgttgtttgcccctccccc

gtgccttccttgaccctggaaggtgccactcccactgtcctttcctaa

taaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctatt

ctggggggtggggggggcaggacagcaagggggaggattgggaagaca

atagcaggcatgctggggatgcggtgggctctatggcttctgaggcgg

aaagaaccagctggggctctagggggtatccccacgcgccctgtagcg

gcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgcta

cacttgccagcgccctagcgcccgctcctttcgctttcttcccttcct

ttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggc

tccctttagggttccgatttagtgctttacggcacctcgaccccaaaa

aacttgattagggtgatggttcacgtagtgggccatcgccctgataga

cggtttttcgccctttgacgttggagtccacgttctttaatagtggac

tcttgttccaaactggaacaacactcaaccctatctcggtctattctt

ttgatttataagggattttgccgatttcggcctattggttaaaaaatg

agctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtg

tcagttagggtgtggaaagtccccaggctccccagcaggcagaagtat

gcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccc

aggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtc

agcaaccatagtcccgcccctaactccgcccatcccgcccctaactcc

gcccagttccgcccattctccgccccatggctgactaattttttttat

ttatgcagaggccgaggccgcctctgcctctgagctattccagaagta

gtgaggaggcttttttggaggcctaggcttttgcaaaaagctccctac

cgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttc

ctgtgtgaaattgttatccgctcacaattccacacaacatacgagccg

gaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactca

cattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgt

cgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtt

tgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgct

cggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaa

tacggttatccacagaatcaggggataacgcaggaaagaacatgtgag

caaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgac

gctcaagtcagaggtggcgaaacccgacaggactataaagataccagg

cgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgc

cgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgc

tttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttc

gctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgct

gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcga

ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacg

gctacactagaagaacagtatttggtatctgcgctctgctgaagccag

ttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgca

gaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagat

tatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagtt

ttaaatcaatctaaagtatatatgagtaaacttggtctgacagttacc

aatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt

catccatagttgcctgactccccgtcgtgtagataactacgatacggg

agggcttaccatctggccccagtgctgcaatgataccgcgagacccac

gctcaccggctccagatttatcagcaataaaccagccagccggaaggg

ccgagcgcagaagtggtcctgcaactttatccgcctccatccagtcta

ttaattgttgccgggaagctagagtaagtagttcgccagttaatagtt

tgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgt

cgtttggtatggcttcattcagctccggttcccaacgatcaaggcgag

ttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtc

ctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatgg

ttatggcagcactgcataattctcttactgtcatgccatccgtaagat

gcttttctgtgactggtgagtactcaaccaagtcattctgagaatagt

gtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataata

ccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgtt

cttcggggcgaaaactctcaaggatcttaccgctgttgagatccagtt

cgatgtaacccactcgtgcacccaactgatcttcagcatcttttactt

tcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaa

aaaagggaataagggcgacacggaaatgttgaatactcatactcttcc

tttttcaatattattgaagcatttatcagggttattgtctcatgagcg

gatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1111 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 49)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

gctctccccaccccaccttctgttttagagctagaaatagcaagttaa

aataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtg

cttttttgaattcgctagctaggtcttgaaaggagtgggaattggctc

cggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaa

gttggggggaggggtcggcaattgatccggtgcctagagaaggtggcg

cggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttcc

cgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgtt

ctttttcgcaacgggtttgccgccagaacacaggaccggtgccaccat

ggactataaggaccacgacggagactacaaggatcatgatattgatta

caaagacgatgacgataagatggccccaaagaagaagcggaaggtcgg

tatccacggagtcccagcagccgacaagaagtacagcatcggcctggc

catcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaa

ggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacag

catcaagaagaacctgatcggagccctgctgttcgacagcggcgaaac

agccgaggccacccggctgaagagaaccgccagaagaagatacaccag

acggaagaaccggatctgctatctgcaagagatcttcagcaacgagat

ggccaaggtggacgacagcttcttccacagactggaagagtccttcct

ggtggaagaggataagaagcacgagcggcaccccatcttcggcaacat

cgtggacgaggtggcctaccacgagaagtaccccaccatctaccacct

gagaaagaaactggtggacagcaccgacaaggccgacctgcggctgat

ctatctggccctggcccacatgatcaagttccggggccacttcctgat

cgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcat

ccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaa

cgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaa

gagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaa

gaatggcctgttcggcaacctgattgccctgagcctgggcctgacccc

caacttcaagagcaacttcgacctggccgaggatgccaaactgcagct

gagcaaggacacctacgacgacgacctggacaacctgctggcccagat

cggcgaccagtacgccgacctgtttctggccgccaagaacctgtccga

cgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaa

ggcccccctgagcgcctctatgatcaagagatacgacgagcaccacca

ggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaa

gtacaaagagattttcttcgaccagagcaagaacggctacgccggcta

cattgacggcggagccagccaggaagagttctacaagttcatcaagcc

catcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaa

cagagaggacctgctgcggaagcagcggaccttcgacaacggcagcat

cccccaccagatccacctgggagagctgcacgccattctgcggcggca

ggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaa

gatcctgaccttccgcatcccctactacgtgggccctctggccagggg

aaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcac

cccctggaacttcgaggaagtggtggacaagggcgcttccgcccagag

cttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaa

ggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataa

cgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgc

cttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaa

gaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcg

gttcaacgcctccctgggcacataccacgatctgctgaaaattatcaa

ggacaaggacttcctggacaatgaggaaaacgaggacattctggaaga

tatcgtgctgaccctgacactgtttgaggacagagagatgatcgagga

acggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagca

gctgaagcggcggagatacaccggctggggcaggctgagccggaagct

gatcaacggcatccgggacaagcagtccggcaagacaatcctggattt

cctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcca

cgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtc

cggccagggcgatagcctgcacgagcacattgccaatctggccggcag

ccccgccattaagaagggcatcctgcagacagtgaaggtggtggacga

gctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcga

aatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcca

gatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaa

gctgtacctgtactacctgcagaatgggcgggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

ccccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatc

cccctcccggctccagatgttcttcgctaataaccacgaccaggaatt

tgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcc

catccgggtgctgtctctctttgatggaatcgctacagggctcctggt

gctgaaggacttgggcattcaggtggaccgctacattgcctcggaggt

gtgtgaggactccatcacggtgggcatggtgcggcaccaggggaagat

catgtacgtcggggacgtccgcagcgtcacacagaagcatatccagga

gtggggcccattcgatctggtgattgggggcagtccctgcaatgacct

ctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccg

gctcttctttgagttctaccgcctcctgcatgatgcgcggcccaagga

gggagatgatcgccccttcttctggctctttgcgaatgtggtggccat

gggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccc

tgtgatgattgatgccaaagaagtgtcagctgcacacagggcccgcta

cttctggggtaaccttcccggtatgaacaggccgttggcatccactgt

gaatgataagctggagctgcaggagtgtctggagcatggcaggatagc

caagttcagcaaagtgaggaccattactacgaggtcaaactccataaa

gcagggcaaagaccagcattttcctgtgttcatgaatgagaaagagga

catcttatggtgcactgaaatggaaagggtatttggtttcccagtcca

ctatactgacgtgtccaacatgagccgcttggcgaggcagagactgct

gggccggtcatggagcgtgccagtcatccgccacctcttcgctccgct

gaaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaac

aaacttctctctgctgaaacaagccggagatgtcgaagagaatcctgg

accgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgt

ccccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgc

cacgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccga

gctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggt

gtgggtcgcggacgacggcgccgcggtggcggtctggaccacgccgga

gagcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggc

cgagttgagcggttcccggctggccgcgcagcaacagatggaaggcct

cctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgt

cggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgt

gctccccggagtggaggcggccgagcgcgccggggtgcccgccttcct

ggagacctccgcgccccgcaacctccccttctacgagcggctcggctt

caccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtg

catgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacc

tctggattacaaaatttgtgaaagattgactggtattcttaactatgt

tgctccttttacgctatgtggatacgctgctttaatgcctttgtatca

tgctattgcttcccgtatggctttcattttctcctccttgtataaatc

ctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg

tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttgggg

cattgccaccacctgtcagctcctttccgggactttcgctttccccct

ccctattgccacggcggaactcatcgccgcctgccttgcccgctgctg

gacaggggctcggctgttgggcactgacaattccgtggtgttgtcggg

gaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggat

tctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagc

ggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcg

tcttcgccttcgccctcagacgagtcggatctccctttgggccgcctc

cccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctt

agccactttttaaaagaaaaggggggactggaagggctaattcactcc

caacgaagacaagatctgctttttgcttgtactgggtctctctggtta

gaccagatctgagcctgggagctctctggctaactagggaacccactg

cttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcc

cgtctgttgtgtgactctggtaactagagatccctcagacccttttag

tcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagc

ctcgactgtgccttctagttgccagccatctgttgtttgcccctcccc

cgtgccttccttgaccctggaaggtgccactcccactgtcctttccta

ataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctat

tctggggggtggggggggcaggacagcaagggggaggattgggaagac

aatagcaggcatgctggggatgcggtgggctctatggcttctgaggcg

gaaagaaccagctggggctctagggggtatccccacgcgccctgtagc

ggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgct

acacttgccagcgccctagcgcccgctcctttcgctttcttcccttcc

tttctcgccacgttcgccggctttccccgtcaagctctaaatcggggg

ctccctttagggttccgatttagtgctttacggcacctcgaccccaaa

aaacttgattagggtgatggttcacgtagtgggccatcgccctgatag

acggtttttcgccctttgacgttggagtccacgttctttaatagtgga

ctcttgttccaaactggaacaacactcaaccctatctcggtctattct

tttgatttataagggattttgccgatttcggcctattggttaaaaaat

gagctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgt

gtcagttagggtgtggaaagtccccaggctccccagcaggcagaagta

tgcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtccc

caggctccccagcaggcagaagtatgcaaagcatgcatctcaattagt

cagcaaccatagtcccgcccctaactccgcccatcccgcccctaactc

cgcccagttccgcccattctccgccccatggctgactaatttttttta

tttatgcagaggccgaggccgcctctgcctctgagctattccagaagt

agtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccta

ccgtcgacctctagctagagcttggcgtaatcatggtcatagctgttt

cctgtgtgaaattgttatccgctcacaattccacacaacatacgagcc

ggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactc

acattaattgcgttgcgctcactgcccgctttccagtcgggaaacctg

tcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggt

ttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgc

tcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta

atacggttatccacagaatcaggggataacgcaggaaagaacatgtga

gcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctg

gcgtttttccataggctccgcccccctgacgagcatcacaaaaatcga

cgctcaagtcagaggtggcgaaacccgacaggactataaagataccag

gcgtttccccctggaagctccctcgtgcgctctcctgttccgaccctg

ccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcg

ctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgtt

cgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgc

tgcgccttatccggtaactatcgtcttgagtccaacccggtaagacac

gacttatcgccactggcagcagccactggtaacaggattagcagagcg

aggtatgtaggcggtgctacagagttcttgaagtggtggcctaactac

ggctacactagaagaacagtatttggtatctgcgctctgctgaagcca

gttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacc

accgctggtagcggtggtttttttgtttgcaagcagcagattacgcgc

agaaaaaaaggatctcaagaagatcctttgatcttttctacggggtct

gacgctcagtggaacgaaaactcacgttaagggattttggtcatgaga

ttatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagt

tttaaatcaatctaaagtatatatgagtaaacttggtctgacagttac

caatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgt

tcatccatagttgcctgactccccgtcgtgtagataactacgatacgg

gagggcttaccatctggccccagtgctgcaatgataccgcgagaccca

cgctcaccggctccagatttatcagcaataaaccagccagccggaagg

gccgagcgcagaagtggtcctgcaactttatccgcctccatccagtct

attaattgttgccgggaagctagagtaagtagttcgccagttaatagt

ttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcg

tcgtttggtatggcttcattcagctccggttcccaacgatcaaggcga

gttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggt

cctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatg

gttatggcagcactgcataattctcttactgtcatgccatccgtaaga

tgcttttctgtgactggtgagtactcaaccaagtcattctgagaatag

tgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataat

accgcgccacatagcagaactttaaaagtgctcatcattggaaaacgt

tcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagt

tcgatgtaacccactcgtgcacccaactgatcttcagcatcttttact

ttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca

aaaaagggaataagggcgacacggaaatgttgaatactcatactcttc

ctttttcaatattattgaagcatttatcagggttattgtctcatgagc

ggatacatatttgaatgtatttagaaaaataaacaaataggggttccg

cgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1112 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 50)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cgtgtgaagggagaatgaggaagttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

ccccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatc

cccctcccggctccagatgttcttcgctaataaccacgaccaggaatt

tgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcc

catccgggtgctgtctctctttgatggaatcgctacagggctcctggt

gctgaaggacttgggcattcaggtggaccgctacattgcctcggaggt

gtgtgaggactccatcacggtgggcatggtgcggcaccaggggaagat

catgtacgtcggggacgtccgcagcgtcacacagaagcatatccagga

gtggggcccattcgatctggtgattgggggcagtccctgcaatgacct

ctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccg

gctcttctttgagttctaccgcctcctgcatgatgcgcggcccaagga

gggagatgatcgccccttcttctggctctttgcgaatgtggtggccat

gggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccc

tgtgatgattgatgccaaagaagtgtcagctgcacacagggcccgcta

cttctggggtaaccttcccggtatgaacaggccgttggcatccactgt

gaatgataagctggagctgcaggagtgtctggagcatggcaggatagc

caagttcagcaaagtgaggaccattactacgaggtcaaactccataaa

gcagggcaaagaccagcattttcctgtgttcatgaatgagaaagagga

catcttatggtgcactgaaatggaaagggtatttggtttcccagtcca

ctatactgacgtgtccaacatgagccgcttggcgaggcagagactgct

gggccggtcatggagcgtgccagtcatccgccacctcttcgctccgct

gaaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaac

aaacttctctctgctgaaacaagccggagatgtcgaagagaatcctgg

accgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgt

ccccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgc

cacgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccga

gctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggt

gtgggtcgcggacgacggcgccgcggtggcggtctggaccacgccgga

gagcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggc

cgagttgagcggttcccggctggccgcgcagcaacagatggaaggcct

cctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgt

cggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgt

gctccccggagtggaggcggccgagcgcgccggggtgcccgccttcct

ggagacctccgcgccccgcaacctccccttctacgagcggctcggctt

caccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtg

catgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacc

tctggattacaaaatttgtgaaagattgactggtattcttaactatgt

tgctccttttacgctatgtggatacgctgctttaatgcctttgtatca

tgctattgcttcccgtatggctttcattttctcctccttgtataaatc

ctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg

tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttgggg

cattgccaccacctgtcagctcctttccgggactttcgctttccccct

ccctattgccacggcggaactcatcgccgcctgccttgcccgctgctg

gacaggggctcggctgttgggcactgacaattccgtggtgttgtcggg

gaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggat

tctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagc

ggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcg

tcttcgccttcgccctcagacgagtcggatctccctttgggccgcctc

cccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctt

agccactttttaaaagaaaaggggggactggaagggctaattcactcc

caacgaagacaagatctgctttttgcttgtactgggtctctctggtta

gaccagatctgagcctgggagctctctggctaactagggaacccactg

cttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcc

cgtctgttgtgtgactctggtaactagagatccctcagacccttttag

tcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagc

ctcgactgtgccttctagttgccagccatctgttgtttgcccctcccc

cgtgccttccttgaccctggaaggtgccactcccactgtcctttccta

ataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctat

tctggggggtggggggggcaggacagcaagggggaggattgggaagac

aatagcaggcatgctggggatgcggtgggctctatggcttctgaggcg

gaaagaaccagctggggctctagggggtatccccacgcgccctgtagc

ggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgct

acacttgccagcgccctagcgcccgctcctttcgctttcttcccttcc

tttctcgccacgttcgccggctttccccgtcaagctctaaatcggggg

ctccctttagggttccgatttagtgctttacggcacctcgaccccaaa

aaacttgattagggtgatggttcacgtagtgggccatcgccctgatag

acggtttttcgccctttgacgttggagtccacgttctttaatagtgga

ctcttgttccaaactggaacaacactcaaccctatctcggtctattct

tttgatttataagggattttgccgatttcggcctattggttaaaaaat

gagctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgt

gtcagttagggtgtggaaagtccccaggctccccagcaggcagaagta

tgcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtccc

caggctccccagcaggcagaagtatgcaaagcatgcatctcaattagt

cagcaaccatagtcccgcccctaactccgcccatcccgcccctaactc

cgcccagttccgcccattctccgccccatggctgactaatttttttta

tttatgcagaggccgaggccgcctctgcctctgagctattccagaagt

agtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccta

ccgtcgacctctagctagagcttggcgtaatcatggtcatagctgttt

cctgtgtgaaattgttatccgctcacaattccacacaacatacgagcc

ggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactc

acattaattgcgttgcgctcactgcccgctttccagtcgggaaacctg

tcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggt

ttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgc

tcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta

atacggttatccacagaatcaggggataacgcaggaaagaacatgtga

gcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctg

gcgtttttccataggctccgcccccctgacgagcatcacaaaaatcga

cgctcaagtcagaggtggcgaaacccgacaggactataaagataccag

gcgtttccccctggaagctccctcgtgcgctctcctgttccgaccctg

ccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcg

ctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgtt

cgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgc

tgcgccttatccggtaactatcgtcttgagtccaacccggtaagacac

gacttatcgccactggcagcagccactggtaacaggattagcagagcg

aggtatgtaggcggtgctacagagttcttgaagtggtggcctaactac

ggctacactagaagaacagtatttggtatctgcgctctgctgaagcca

gttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacc

accgctggtagcggtggtttttttgtttgcaagcagcagattacgcgc

agaaaaaaaggatctcaagaagatcctttgatcttttctacggggtct

gacgctcagtggaacgaaaactcacgttaagggattttggtcatgaga

ttatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagt

tttaaatcaatctaaagtatatatgagtaaacttggtctgacagttac

caatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgt

tcatccatagttgcctgactccccgtcgtgtagataactacgatacgg

gagggcttaccatctggccccagtgctgcaatgataccgcgagaccca

cgctcaccggctccagatttatcagcaataaaccagccagccggaagg

gccgagcgcagaagtggtcctgcaactttatccgcctccatccagtct

attaattgttgccgggaagctagagtaagtagttcgccagttaatagt

ttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcg

tcgtttggtatggcttcattcagctccggttcccaacgatcaaggcga

gttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggt

cctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatg

gttatggcagcactgcataattctcttactgtcatgccatccgtaaga

tgcttttctgtgactggtgagtactcaaccaagtcattctgagaatag

tgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataat

accgcgccacatagcagaactttaaaagtgctcatcattggaaaacgt

tcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagt

tcgatgtaacccactcgtgcacccaactgatcttcagcatcttttact

ttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca

aaaaagggaataagggcgacacggaaatgttgaatactcatactcttc

ctttttcaatattattgaagcatttatcagggttattgtctcatgagc

ggatacatatttgaatgtatttagaaaaataaacaaataggggttccg

cgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1426 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 53)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cggcggtactgcaccaggcggcgttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

ccccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatc

cccctcccggctccagatgttcttcgctaataaccacgaccaggaatt

tgaccctccaaaggtttacccacctgtcccagctgagaagaggaagcc

catccgggtgctgtctctctttgatggaatcgctacagggctcctggt

gctgaaggacttgggcattcaggtggaccgctacattgcctcggaggt

gtgtgaggactccatcacggtgggcatggtgcggcaccaggggaagat

catgtacgtcggggacgtccgcagcgtcacacagaagcatatccagga

gtggggcccattcgatctggtgattgggggcagtccctgcaatgacct

ctccatcgtcaaccctgctcgcaagggcctctacgagggcactggccg

gctcttctttgagttctaccgcctcctgcatgatgcgcggcccaagga

gggagatgatcgccccttcttctggctctttgagaatgtggtggccat

gggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccc

tgtgatgattgatgccaaagaagtgtcagctgcacacagggcccgcta

cttctggggtaaccttcccggtatgaacaggccgttggcatccactgt

gaatgataagctggagctgcaggagtgtctggagcatggcaggatagc

caagttcagcaaagtgaggaccattactacgaggtcaaactccataaa

gcagggcaaagaccagcattttcctgtgttcatgaatgagaaagagga

catcttatggtgcactgaaatggaaagggtatttggtttcccagtcca

ctatactgacgtgtccaacatgagccgcttggcgaggcagagactgct

gggccggtcatggagcgtgccagtcatccgccacctcttcgctccgct

gaaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaac

aaacttctctctgctgaaacaagccggagatgtcgaagagaatcctgg

accgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgt

ccccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgc

cacgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccga

gctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggt

gtgggtcgcggacgacggcgccgcggtggcggtctggaccacgccgga

gagcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggc

cgagttgagcggttcccggctggccgcgcagcaacagatggaaggcct

cctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgt

cggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgt

gctccccggagtggaggcggccgagcgcgccggggtgcccgccttcct

ggagacctccgcgccccgcaacctccccttctacgagcggctcggctt

caccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtg

catgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacc

tctggattacaaaatttgtgaaagattgactggtattcttaactatgt

tgctccttttacgctatgtggatacgctgctttaatgcctttgtatca

tgctattgcttcccgtatggctttcattttctcctccttgtataaatc

ctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg

tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttgggg

cattgccaccacctgtcagctcctttccgggactttcgctttccccct

ccctattgccacggcggaactcatcgccgcctgccttgcccgctgctg

gacaggggctcggctgttgggcactgacaattccgtggtgttgtcggg

gaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggat

tctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagc

ggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcg

tcttcgccttcgccctcagacgagtcggatctccctttgggccgcctc

cccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctt

agccactttttaaaagaaaaggggggactggaagggctaattcactcc

caacgaagacaagatctgctttttgcttgtactgggtctctctggtta

gaccagatctgagcctgggagctctctggctaactagggaacccactg

cttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcc

cgtctgttgtgtgactctggtaactagagatccctcagacccttttag

tcagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagc

ctcgactgtgccttctagttgccagccatctgttgtttgcccctcccc

cgtgccttccttgaccctggaaggtgccactcccactgtcctttccta

ataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctat

tctggggggtggggggggcaggacagcaagggggaggattgggaagac

aatagcaggcatgctggggatgcggtgggctctatggcttctgaggcg

gaaagaaccagctggggctctagggggtatccccacgcgccctgtagc

ggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgct

acacttgccagcgccctagcgcccgctcctttcgctttcttcccttcc

tttctcgccacgttcgccggctttccccgtcaagctctaaatcggggg

ctccctttagggttccgatttagtgctttacggcacctcgaccccaaa

aaacttgattagggtgatggttcacgtagtgggccatcgccctgatag

acggtttttcgccctttgacgttggagtccacgttctttaatagtgga

ctcttgttccaaactggaacaacactcaaccctatctcggtctattct

tttgatttataagggattttgccgatttcggcctattggttaaaaaat

gagctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgt

gtcagttagggtgtggaaagtccccaggctccccagcaggcagaagta

tgcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtccc

caggctccccagcaggcagaagtatgcaaagcatgcatctcaattagt

cagcaaccatagtcccgcccctaactccgcccatcccgcccctaactc

cgcccagttccgcccattctccgccccatggctgactaatttttttta

tttatgcagaggccgaggccgcctctgcctctgagctattccagaagt

agtgaggaggcttttttggaggcctaggcttttgcaaaaagctcccta

ccgtcgacctctagctagagcttggcgtaatcatggtcatagctgttt

cctgtgtgaaattgttatccgctcacaattccacacaacatacgagcc

ggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactc

acattaattgcgttgcgctcactgcccgctttccagtcgggaaacctg

tcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggt

ttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgc

tcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta

atacggttatccacagaatcaggggataacgcaggaaagaacatgtga

gcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctg

gcgtttttccataggctccgcccccctgacgagcatcacaaaaatcga

cgctcaagtcagaggtggcgaaacccgacaggactataaagataccag

gcgtttccccctggaagctccctcgtgcgctctcctgttccgaccctg

ccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcg

ctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgtt

cgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgc

tgcgccttatccggtaactatcgtcttgagtccaacccggtaagacac

gacttatcgccactggcagcagccactggtaacaggattagcagagcg

aggtatgtaggcggtgctacagagttcttgaagtggtggcctaactac

ggctacactagaagaacagtatttggtatctgcgctctgctgaagcca

gttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacc

accgctggtagcggtggtttttttgtttgcaagcagcagattacgcgc

agaaaaaaaggatctcaagaagatcctttgatcttttctacggggtct

gacgctcagtggaacgaaaactcacgttaagggattttggtcatgaga

ttatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagt

tttaaatcaatctaaagtatatatgagtaaacttggtctgacagttac

caatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgt

tcatccatagttgcctgactccccgtcgtgtagataactacgatacgg

gagggcttaccatctggccccagtgctgcaatgataccgcgagaccca

cgctcaccggctccagatttatcagcaataaaccagccagccggaagg

gccgagcgcagaagtggtcctgcaactttatccgcctccatccagtct

attaattgttgccgggaagctagagtaagtagttcgccagttaatagt

ttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcg

tcgtttggtatggcttcattcagctccggttcccaacgatcaaggcga

gttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggt

cctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatg

gttatggcagcactgcataattctcttactgtcatgccatccgtaaga

tgcttttctgtgactggtgagtactcaaccaagtcattctgagaatag

tgtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataat

accgcgccacatagcagaactttaaaagtgctcatcattggaaaacgt

tcttcggggcgaaaactctcaaggatcttaccgctgttgagatccagt

tcgatgtaacccactcgtgcacccaactgatcttcagcatcttttact

ttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca

aaaaagggaataagggcgacacggaaatgttgaatactcatactcttc

ctttttcaatattattgaagcatttatcagggttattgtctcatgagc

ggatacatatttgaatgtatttagaaaaataaacaaataggggttccg

cgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1427 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 54)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaaaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggcggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

ggggcgcggacatggaggacggttttagagctagaaatagcaagttaa

aataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtg

cttttttgaattcgctagctaggtcttgaaaggagtgggaattggctc

cggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaa

gttggggggaggggtcggcaattgatccggtgcctagagaaggtggcg

cggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttcc

cgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgtt

ctttttcgcaacgggtttgccgccagaacacaggaccggtgccaccat

ggactataaggaccacgacggagactacaaggatcatgatattgatta

caaagacgatgacgataagatggccccaaagaagaagcggaaggtcgg

tatccacggagtcccagcagccgacaagaagtacagcatcggcctggc

catcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaa

ggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacag

catcaagaagaacctgatcggagccctgctgttcgacagcggcgaaac

agccgaggccacccggctgaagagaaccgccagaagaagatacaccag

acggaagaaccggatctgctatctgcaagagatcttcagcaacgagat

ggccaaggtggacgacagcttcttccacagactggaagagtccttcct

ggtggaagaggataagaagcacgagcggcaccccatcttcggcaacat

cgtggacgaggtggcctaccacgagaagtaccccaccatctaccacct

gagaaagaaactggtggacagcaccgacaaggccgacctgcggctgat

ctatctggccctggcccacatgatcaagttccggggccacttcctgat

cgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcat

ccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaa

cgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaa

gagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaa

gaatggcctgttcggcaacctgattgccctgagcctgggcctgacccc

caacttcaagagcaacttcgacctggccgaggatgccaaactgcagct

gagcaaggacacctacgacgacgacctggacaacctgctggcccagat

cggcgaccagtacgccgacctgtttctggccgccaagaacctgtccga

cgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaa

ggcccccctgagcgcctctatgatcaagagatacgacgagcaccacca

ggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaa

gtacaaagagattttcttcgaccagagcaagaacggctacgccggcta

cattgacggcggagccagccaggaagagttctacaagttcatcaagcc

catcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaa

cagagaggacctgctgcggaagcagcggaccttcgacaacggcagcat

cccccaccagatccacctgggagagctgcacgccattctgcggcggca

ggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaa

gatcctgaccttccgcatcccctactacgtgggccctctggccagggg

aaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcac

cccctggaacttcgaggaagtggtggacaagggcgcttccgcccagag

cttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaa

ggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataa

cgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgc

cttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaa

gaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcg

gttcaacgcctccctgggcacataccacgatctgctgaaaattatcaa

ggacaaggacttcctggacaatgaggaaaacgaggacattctggaaga

tatcgtgctgaccctgacactgtttgaggacagagagatgatcgagga

acggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagca

gctgaagcggcggagatacaccggctggggcaggctgagccggaagct

gatcaacggcatccgggacaagcagtccggcaagacaatcctggattt

cctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcca

cgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtc

cggccagggcgatagcctgcacgagcacattgccaatctggccggcag

ccccgccattaagaagggcatcctgcagacagtgaaggtggtggacga

gctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcga

aatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcca

gatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaa

gctgtacctgtactacctgcagaatggggggatatgtacgtggaccag

gaactggacatcaaccggctgtccgactacgatgtggacgctatcgtg

cctcagagctttctgaaggacgactccatcgacaacaaggtgctgacc

agaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagag

gtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaag

ctgattacccagagaaagttcgacaatctgaccaaggccgagagaggc

ggcctgagcgaactggataaggccggcttcatcaagagacagctggtg

gaaacccggcagatcacaaagcacgtggcacagatactagattcccga

atgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaa

gtaatcactttaaagtcaaaattggtgtcggacttcagaaaggatttt

caattctataaagttagggagataaataactaccaccatgcgcacgac

gcttatcttaatgccgtcgtagggaccgcactcattaagaaatacccg

aagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtc

cgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcc

aaatacttcttttattctaacattatgaatttctttaagacggaaatc

actctggcaaacggagagatacgcaaacgacctttaattgaaaccaat

ggggagacaggtgaaatcgtatgggataagggccgggacttcgcgacg

gtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaact

gaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagg

aatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaag

tacggtggcttcgtgagccctacagttgcctattctgtcctagtagtg

gcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaa

ttattggggataacgattatggagcgctcgtcttttgaaaagaacccc

atcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctc

ataattaaactaccaaagtatagtctgtttgagttagaaaatggccga

aaacggatgttggctagcgccagagagcttcaaaaggggaacgaactc

gcactaccgtctaaatacgtgaatttcctgtatttagcgtcccattac

gagaagttgaaaggttcacctgaagataacgaacagaagcaacttttt

gttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcg

gaattcagtaagagagtcatcctagctgatgccaatctggacaaagta

ttaagcgcatacaacaagcacagggataaacccatacgtgagcaggcg

gaaaatattatccatttgtttactcttaccaacctcggcgctccagcc

gcattcaagtattttgacacaacgatagatcgcaaagagtacagatct

accaaggaggtgctagacgcgacactgattcaccaatccatcacggga

ttatatgaaactcggatagatttgtcacagcttgggggtgacggatcc

cccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatcc

ccctcccggctccagatgttcttcgctaataaccacgaccaggaattt

gaccctccaaaggtttacccacctgtcccagctgagaagaggaagccc

atccgggtgctgtctctctttgatggaatcgctacagggctcctggtg

ctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtg

tgtgaggactccatcacggtgggcatggtgcggcaccaggggaagatc

atgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggag

tggggcccattcgatctggtgattgggggcagtccctgcaatgacctc

tccatcgtcaaccctgctcgcaagggcctctacgagggcactggccgg

ctcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggag

ggagatgatcgccccttcttctggctctttgagaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccct

gtgatgattgatgccaaagaagtgtcagctgcacacagggcccgctac

ttctggggtaaccttcccggtatgaacaggccgttggcatccactgtg

aatgataagctggagctgcaggagtgtctggagcatggcaggatagcc

aagttcagcaaagtgaggaccattactacgaggtcaaactccataaag

cagggcaaagaccagcattttcctgtgttcatgaatgagaaagaggac

atcttatggtgcactgaaatggaaagggtatttggtttcccagtccac

tatactgacgtgtccaacatgagccgcttggcgaggcagagactgctg

ggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctg

aaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaaca

aacttctctctgctgaaacaagccggagatgtcgaagagaatcctgga

ccgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtc

cccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgcc

acgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgag

ctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtg

tgggtcgcggacgacggcgccgcggtggcggtctggaccacgccggag

agcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggcc

gagttgagcggttcccggctggccgcgcagcaacagatggaaggcctc

ctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtc

ggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtg

ctccccggagtggaggcggccgagcgcgccggggtgcccgccttcctg

gagacctccgcgccccgcaacctccccttctacgagcggctcggcttc

accgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgc

atgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacct

ctggattacaaaatttgtgaaagattgactggtattcttaactatgtt

gctccttttacgctatgtggatacgctgctttaatgcctttgtatcat

gctattgcttcccgtatggctttcattttctcctccttgtataaatcc

tggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgt

ggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggc

attgccaccacctgtcagctcctttccgggactttcgctttccccctc

cctattgccacggcggaactcatcgccgcctgccttgcccgctgctgg

acaggggctcggctgttgggcactgacaattccgtggtgttgtcgggg

aaatcatcgtcctttccttggctgctcgcctgtgttgccacctggatt

ctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcg

gaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgt

cttcgccttcgccctcagacgagtcggatctccctttgggccgcctcc

ccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctta

gccactttttaaaagaaaaggggggactggaagggctaattcactccc

aacgaagacaagatctgctttttgcttgtactgggtctctctggttag

accagatctgagcctgggagctctctggctaactagggaacccactgc

ttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgccc

gtctgttgtgtgactctggtaactagagatccctcagacccttttagt

cagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcc

tcgactgtgccttctagttgccagccatctgttgtttgcccctccccc

gtgccttccttgaccctggaaggtgccactcccactgtcctttcctaa

taaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctatt

ctggggggtggggggggcaggacagcaagggggaggattgggaagaca

atagcaggcatgctggggatgcggtgggctctatggcttctgaggcgg

aaagaaccagctggggctctagggggtatccccacgcgccctgtagcg

gcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgcta

cacttgccagcgccctagcgcccgctcctttcgctttcttcccttcct

ttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggc

tccctttagggttccgatttagtgctttacggcacctcgaccccaaaa

aacttgattagggtgatggttcacgtagtgggccatcgccctgataga

cggtttttcgccctttgacgttggagtccacgttctttaatagtggac

tcttgttccaaactggaacaacactcaaccctatctcggtctattctt

ttgatttataagggattttgccgatttcggcctattggttaaaaaatg

agctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtg

tcagttagggtgtggaaagtccccaggctccccagcaggcagaagtat

gcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccc

aggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtc

agcaaccatagtcccgcccctaactccgcccatcccgcccctaactcc

gcccagttccgcccattctccgccccatggctgactaattttttttat

ttatgcagaggccgaggccgcctctgcctctgagctattccagaagta

gtgaggaggcttttttggaggcctaggcttttgcaaaaagctccctac

cgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttc

ctgtgtgaaattgttatccgctcacaattccacacaacatacgagccg

gaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactca

cattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgt

cgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtt

tgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgct

cggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaa

tacggttatccacagaatcaggggataacgcaggaaagaacatgtgag

caaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgac

gctcaagtcagaggtggcgaaacccgacaggactataaagataccagg

cgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgc

cgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgc

tttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttc

gctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgct

gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcga

ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacg

gctacactagaagaacagtatttggtatctgcgctctgctgaagccag

ttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgca

gaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagat

tatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagtt

ttaaatcaatctaaagtatatatgagtaaacttggtctgacagttacc

aatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt

catccatagttgcctgactccccgtcgtgtagataactacgatacggg

agggcttaccatctggccccagtgctgcaatgataccgcgagacccac

gctcaccggctccagatttatcagcaataaaccagccagccggaaggg

ccgagcgcagaagtggtcctgcaactttatccgcctccatccagtcta

ttaattgttgccgggaagctagagtaagtagttcgccagttaatagtt

tgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgt

cgtttggtatggcttcattcagctccggttcccaacgatcaaggcgag

ttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtc

ctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatgg

ttatggcagcactgcataattctcttactgtcatgccatccgtaagat

gcttttctgtgactggtgagtactcaaccaagtcattctgagaatagt

gtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataata

ccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgtt

cttcggggcgaaaactctcaaggatcttaccgctgttgagatccagtt

cgatgtaacccactcgtgcacccaactgatcttcagcatcttttactt

tcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaa

aaaagggaataagggcgacacggaaatgttgaatactcatactcttcc

tttttcaatattattgaagcatttatcagggttattgtctcatgagcg

gatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1428 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 55)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacgggggaggtctatataagcag

cgcgttttgcctgtactgggtctctctggttagaccagatctgagcct

gggagctctctggctaactagggaacccactgcttaagcctcaataaa

gcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgact

ctggtaactagagatccctcagacccttttagtcagtgtggaaaatct

ctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccaga

ggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaag

aggcgaggggcggcgactggtgagtacgccaaaaattttgactagcgg

aggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggg

gagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaa

gaaaaaatataaattaaaacatatagtatgggcaagcagggagctaga

acgattcgcagttaatcctggcctgttagaaacatcagaaggctgtag

acaaatactgggacagctacaaccatcccttcagacaggatcagaaga

acttagatcattatataatacagtagcaaccctctattgtgtgcatca

aaggatagagataaaagacaccaaggaagctttagacaagatagagga

agagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggggaacggtaccgagggcctat

ttcccatgattccttcatatttgcatatacgatacaaggctgttagag

agataattagaattaatttgactgtaaacacaaagatattagtacaaa

atacgtgacgtagaaagtaataatttcttgggtagtttgcagttttaa

aattatgttttaaaatggactatcatatgcttaccgtaacttgaaagt

atttcgatttcttggctttatatatcttgtggaaaggacgaaacaccg

gcggtactgcaccaggcggcgttttagagctagaaatagcaagttaaa

ataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtgc

ttttttgaattcgctagctaggtcttgaaaggagtgggaattggctcc

ggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaag

ttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgc

ggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttccc

gaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttc

tttttcgcaacgggtttgccgccagaacacaggaccggtgccaccatg

gactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggt

atccacggagtcccagcagccgacaagaagtacagcatcggcctggcc

atcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaag

gtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacagc

atcaagaagaacctgatcggagccctgctgttcgacagcggcgaaaca

gccgaggccacccggctgaagagaaccgccagaagaagatacaccaga

cggaagaaccggatctgctatctgcaagagatcttcagcaacgagatg

gccaaggtggacgacagcttcttccacagactggaagagtccttcctg

gtggaagaggataagaagcacgagcggcaccccatcttcggcaacatc

gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctg

agaaagaaactggtggacagcaccgacaaggccgacctgcggctgatc

tatctggccctggcccacatgatcaagttccggggccacttcctgatc

gagggcgacctgaaccccgacaacagcgacgtggacaagctgttcatc

cagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaac

gccagcggcgtggacgccaaggccatcctgtctgccagactgagcaag

agcagacggctggaaaatctgatcgcccagctgcccggcgagaagaag

aatggcctgttcggcaacctgattgccctgagcctgggcctgaccccc

aacttcaagagcaacttcgacctggccgaggatgccaaactgcagctg

agcaaggacacctacgacgacgacctggacaacctgctggcccagatc

ggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgac

gccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaag

gcccccctgagcgcctctatgatcaagagatacgacgagcaccaccag

gacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaag

tacaaagagattttcttcgaccagagcaagaacggctacgccggctac

attgacggcggagccagccaggaagagttctacaagttcatcaagccc

atcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaac

agagaggacctgctgcggaagcagcggaccttcgacaacggcagcatc

ccccaccagatccacctgggagagctgcacgccattctgcggcggcag

gaagatttttacccattcctgaaggacaaccgggaaaagatcgagaag

atcctgaccttccgcatcccctactacgtgggccctctggccagggga

aacagcagattcgcctggatgaccagaaagagcgaggaaaccatcacc

ccctggaacttcgaggaagtggtggacaagggcgcttccgcccagagc

ttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaag

gtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataac

gagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgcc

ttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaag

accaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaag

aaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcgg

ttcaacgcctccctgggcacataccacgatctgctgaaaattatcaag

gacaaggacttcctggacaatgaggaaaacgaggacattctggaagat

atcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaa

cggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcag

ctgaagcggcggagatacaccggctggggcaggctgagccggaagctg

atcaacggcatccgggacaagcagtccggcaagacaatcctggatttc

ctgaagtccgacggcttcgccaacagaaacttcatgcagctgatccac

gacgacagcctgacctttaaagaggacatccagaaagcccaggtgtcc

ggccagggcgatagcctgcacgagcacattgccaatctggccggcagc

cccgccattaagaagggcatcctgcagacagtgaaggtggtggacgag

ctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcgaa

atggccagagagaaccagaccacccagaagggacagaagaacagccgc

gagagaatgaagcggatcgaagagggcatcaaagagctgggcagccag

atcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaag

ctgtacctgtactacctgcagaatggggggatatgtacgtggaccagg

aactggacatcaaccggctgtccgactacgatgtggacgctatcgtgc

ctcagagctttctgaaggacgactccatcgacaacaaggtgctgacca

gaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagagg

tcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagc

tgattacccagagaaagttcgacaatctgaccaaggccgagagaggcg

gcctgagcgaactggataaggccggcttcatcaagagacagctggtgg

aaacccggcagatcacaaagcacgtggcacagatactagattcccgaa

tgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaag

taatcactttaaagtcaaaattggtgtcggacttcagaaaggattttc

aattctataaagttagggagataaataactaccaccatgcgcacgacg

cttatcttaatgccgtcgtagggaccgcactcattaagaaatacccga

agctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtcc

gtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcca

aatacttcttttattctaacattatgaatttctttaagacggaaatca

ctctggcaaacggagagatacgcaaacgacctttaattgaaaccaatg

gggagacaggtgaaatcgtatgggataagggccgggacttcgcgacgg

tgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaactg

aggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagga

atagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaagt

acggtggcttcgtgagccctacagttgcctattctgtcctagtagtgg

caaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaat

tattggggataacgattatggagcgctcgtcttttgaaaagaacccca

tcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctca

taattaaactaccaaagtatagtctgtttgagttagaaaatggccgaa

aacggatgttggctagcgccagagagcttcaaaaggggaacgaactcg

cactaccgtctaaatacgtgaatttcctgtatttagcgtcccattacg

agaagttgaaaggttcacctgaagataacgaacagaagcaactttttg

ttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcgg

aattcagtaagagagtcatcctagctgatgccaatctggacaaagtat

taagcgcatacaacaagcacagggataaacccatacgtgagcaggcgg

aaaatattatccatttgtttactcttaccaacctcggcgctccagccg

cattcaagtattttgacacaacgatagatcgcaaagagtacagatcta

ccaaggaggtgctagacgcgacactgattcaccaatccatcacgggat

tatatgaaactcggatagatttgtcacagcttgggggtgacggatccc

ccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatccc

cctcccggctccagatgttcttcgctaataaccacgaccaggaatttg

accctccaaaggtttacccacctgtcccagctgagaagaggaagccca

tccgggtgctgtctctctttgatggaatcgctacagggctcctggtgc

tgaaggacttgggcattcaggtggaccgctacattgcctcggaggtgt

gtgaggactccatcacggtgggcatggtgcggcaccaggggaagatca

tgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggagt

ggggcccattcgatctggtgattgggggcagtccctgcaatgacctct

ccatcgtcaaccctgctcgcaagggcctctacgagggcactggccggc

tcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggagg

gagatgatcgccccttcttctggctctttgcgaatgtggtggccatgg

gcgttagtgacaagagggacatctcgcgatttctcgagtccaaccctg

tgatgattgatgccaaagaagtgtcagctgcacacagggcccgctact

tctggggtaaccttcccggtatgaacaggccgttggcatccactgtga

atgataagctggagctgcaggagtgtctggagcatggcaggatagcca

agttcagcaaagtgaggaccattactacgaggtcaaactccataaagc

agggcaaagaccagcattttcctgtgttcatgaatgagaaagaggaca

tcttatggtgcactgaaatggaaagggtatttggtttcccagtccact

atactgacgtgtccaacatgagccgcttggcgaggcagagactgctgg

gccggtcatggagcgtgccagtcatccgccacctcttcgctccgctga

aggagtattttgcgtgtgtgtccggccggcccggatccggcgcaacaa

acttctctctgctgaaacaagccggagatgtcgaagagaatcctggac

cgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtcc

ccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgcca

cgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgagc

tgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtgt

gggtcgcggacgacggcgccgcggtggcggtctggaccacgccggaga

gcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggccg

agttgagcggttcccggctggccgcgcagcaacagatggaaggcctcc

tggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtcg

gagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtgc

tccccggagtggaggcggccgagcgcgccggggtgcccgccttcctgg

agacctccgcgccccgcaacctccccttctacgagcggctcggcttca

ccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgca

tgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacctc

tggattacaaaatttgtgaaagattgactggtattcttaactatgttg

ctccttttacgctatgtggatacgctgctttaatgcctttgtatcatg

ctattgcttcccgtatggctttcattttctcctccttgtataaatcct

ggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtg

gcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggca

ttgccaccacctgtcagctcctttccgggactttcgctttccccctcc

ctattgccacggcggaactcatcgccgcctgccttgcccgctgctgga

caggggctcggctgttgggcactgacaattccgtggtgttgtcgggga

aatcatcgtcctttccttggctgctcgcctgtgttgccacctggattc

tgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcgg

accttccttcccgcggcctgctgccggctctgcggcctcttccgcgtc

ttcgccttcgccctcagacgagtcggatctccctttgggccgcctccc

cgcgtcgactttaagaccaatgacttacaaggcagctgtagatcttag

ccactttttaaaagaaaaggggggactggaagggctaattcactccca

acgaagacaagatctgctttttgcttgtactgggtctctctggttaga

ccagatctgagcctgggagctctctggctaactagggaacccactgct

taagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccg

tctgttgtgtgactctggtaactagagatccctcagacccttttagtc

agtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcct

cgactgtgccttctagttgccagccatctgttgtttgcccctcccccg

tgccttccttgaccctggaaggtgccactcccactgtcctttcctaat

aaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctattc

tggggggtggggggggcaggacagcaagggggaggattgggaagacaa

tagcaggcatgctggggatgcggtgggctctatggcttctgaggcgga

aagaaccagctggggctctagggggtatccccacgcgccctgtagcgg

cgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgctac

acttgccagcgccctagcgcccgctcctttcgctttcttcccttcctt

tctcgccacgttcgccggctttccccgtcaagctctaaatcgggggct

ccctttagggttccgatttagtgctttacggcacctcgaccccaaaaa

acttgattagggtgatggttcacgtagtgggccatcgccctgatagac

ggtttttcgccctttgacgttggagtccacgttctttaatagtggact

cttgttccaaactggaacaacactcaaccctatctcggtctattcttt

tgatttataagggattttgccgatttcggcctattggttaaaaaatga

gctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtgt

cagttagggtgtggaaagtccccaggctccccagcaggcagaagtatg

caaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccca

ggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtca

gcaaccatagtcccgcccctaactccgcccatcccgcccctaactccg

cccagttccgcccattctccgccccatggctgactaattttttttatt

tatgcagaggccgaggccgcctctgcctctgagctattccagaagtag

tgaggaggcttttttggaggcctaggcttttgcaaaaagctccctacc

gtcgacctctagctagagcttggcgtaatcatggtcatagctgtttcc

tgtgtgaaattgttatccgctcacaattccacacaacatacgagccgg

aagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcac

attaattgcgttgcgctcactgcccgctttccagtcgggaaacctgtc

gtgccagctgcattaatgaatcggccaacgcgcggggagaggcggttt

gcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctc

ggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaat

acggttatccacagaatcaggggataacgcaggaaagaacatgtgagc

aaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggc

gtttttccataggctccgcccccctgacgagcatcacaaaaatcgacg

ctcaagtcagaggtggcgaaacccgacaggactataaagataccaggc

gtttccccctggaagctccctcgtgcgctctcctgttccgaccctgcc

gcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgct

ttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcg

ctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctg

cgccttatccggtaactatcgtcttgagtccaacccggtaagacacga

cttatcgccactggcagcagccactggtaacaggattagcagagcgag

gtatgtaggcggtgctacagagttcttgaagtggtggcctaactacgg

ctacactagaagaacagtatttggtatctgcgctctgctgaagccagt

taccttcggaaaaagagttggtagctcttgatccggcaaacaaaccac

cgctggtagcggtggtttttttgtttgcaagcagcagattacgcgcag

aaaaaaaggatctcaagaagatcctttgatcttttctacggggtctga

cgctcagtggaacgaaaactcacgttaagggattttggtcatgagatt

atcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttt

taaatcaatctaaagtatatatgagtaaacttggtctgacagttacca

atgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttc

atccatagttgcctgactccccgtcgtgtagataactacgatacggga

gggcttaccatctggccccagtgctgcaatgataccgcgagacccacg

ctcaccggctccagatttatcagcaataaaccagccagccggaagggc

cgagcgcagaagtggtcctgcaactttatccgcctccatccagtctat

taattgttgccgggaagctagagtaagtagttcgccagttaatagttt

gcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtc

gtttggtatggcttcattcagctccggttcccaacgatcaaggcgagt

tacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcc

tccgatcgttgtcagaagtaagttggccgcagtgttatcactcatggt

tatggcagcactgcataattctcttactgtcatgccatccgtaagatg

cttttctgtgactggtgagtactcaaccaagtcattctgagaatagtg

tatgcggcgaccgagttgctcttgcccggcgtcaatacgggataatac

cgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttc

ttcggggcgaaaactctcaaggatcttaccgctgttgagatccagttc

gatgtaacccactcgtgcacccaactgatcttcagcatcttttacttt

caccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaaa

aaagggaataagggcgacacggaaatgttgaatactcatactcttcct

ttttcaatattattgaagcatttatcagggttattgtctcatgagcgg

atacatatttgaatgtatttagaaaaataaacaaataggggttccgcg

cacatttccccgaaaagtgccacct.

In an aspect, a disclosed pBK1428 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 56)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

ggggcgcggacatggaggacggttttagagctagaaatagcaagttaa

aataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtg

cttttttgaattcgctagctaggtcttgaaaggagtgggaattggctc

cggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaa

gttggggggaggggtcggcaattgatccggtgcctagagaaggtggcg

cggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttcc

cgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgtt

ctttttcgcaacgggtttgccgccagaacacaggaccggtgccaccat

ggactataaggaccacgacggagactacaaggatcatgatattgatta

caaagacgatgacgataagatggccccaaagaagaagcggaaggtcgg

tatccacggagtcccagcagccgacaagaagtacagcatcggcctggc

catcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaa

ggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacag

catcaagaagaacctgatcggagccctgctgttcgacagcggcgaaac

agccgaggccacccggctgaagagaaccgccagaagaagatacaccag

acggaagaaccggatctgctatctgcaagagatcttcagcaacgagat

ggccaaggtggacgacagcttcttccacagactggaagagtccttcct

ggtggaagaggataagaagcacgagcggcaccccatcttcggcaacat

cgtggacgaggtggcctaccacgagaagtaccccaccatctaccacct

gagaaagaaactggtggacagcaccgacaaggccgacctgcggctgat

ctatctggccctggcccacatgatcaagttccggggccacttcctgat

cgagggcgacctgaaccccgacaacagcgacgtggacaagctgttcat

ccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaa

cgccagcggcgtggacgccaaggccatcctgtctgccagactgagcaa

gagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaa

gaatggcctgttcggcaacctgattgccctgagcctgggcctgacccc

caacttcaagagcaacttcgacctggccgaggatgccaaactgcagct

gagcaaggacacctacgacgacgacctggacaacctgctggcccagat

cggcgaccagtacgccgacctgtttctggccgccaagaacctgtccga

cgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaa

ggcccccctgagcgcctctatgatcaagagatacgacgagcaccacca

ggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaa

gtacaaagagattttcttcgaccagagcaagaacggctacgccggcta

cattgacggcggagccagccaggaagagttctacaagttcatcaagcc

catcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaa

cagagaggacctgctgcggaagcagcggaccttcgacaacggcagcat

cccccaccagatccacctgggagagctgcacgccattctgcggcggca

ggaagatttttacccattcctgaaggacaaccgggaaaagatcgagaa

gatcctgaccttccgcatcccctactacgtgggccctctggccagggg

aaacagcagattcgcctggatgaccagaaagagcgaggaaaccatcac

cccctggaacttcgaggaagtggtggacaagggcgcttccgcccagag

cttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaa

ggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataa

cgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgc

cttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaa

gaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaa

gaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcg

gttcaacgcctccctgggcacataccacgatctgctgaaaattatcaa

ggacaaggacttcctggacaatgaggaaaacgaggacattctggaaga

tatcgtgctgaccctgacactgtttgaggacagagagatgatcgagga

acggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagca

gctgaagcggcggagatacaccggctggggcaggctgagccggaagct

gatcaacggcatccgggacaagcagtccggcaagacaatcctggattt

cctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcca

cgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtc

cggccagggcgatagcctgcacgagcacattgccaatctggccggcag

ccccgccattaagaagggcatcctgcagacagtgaaggtggtggacga

gctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcga

aatggccagagagaaccagaccacccagaagggacagaagaacagccg

cgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcca

gatcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaa

gctgtacctgtactacctgcagaatggggggatatgtacgtggaccag

gaactggacatcaaccggctgtccgactacgatgtggacgctatcgtg

cctcagagctttctgaaggacgactccatcgacaacaaggtgctgacc

agaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagag

gtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaag

ctgattacccagagaaagttcgacaatctgaccaaggccgagagaggc

ggcctgagcgaactggataaggccggcttcatcaagagacagctggtg

gaaacccggcagatcacaaagcacgtggcacagatactagattcccga

atgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaa

gtaatcactttaaagtcaaaattggtgtcggacttcagaaaggatttt

caattctataaagttagggagataaataactaccaccatgcgcacgac

gcttatcttaatgccgtcgtagggaccgcactcattaagaaatacccg

aagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtc

cgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcc

aaatacttcttttattctaacattatgaatttctttaagacggaaatc

actctggcaaacggagagatacgcaaacgacctttaattgaaaccaat

ggggagacaggtgaaatcgtatgggataagggccgggacttcgcgacg

gtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaact

gaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagg

aatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaag

tacggtggcttcgtgagccctacagttgcctattctgtcctagtagtg

gcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaa

ttattggggataacgattatggagcgctcgtcttttgaaaagaacccc

atcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctc

ataattaaactaccaaagtatagtctgtttgagttagaaaatggccga

aaacggatgttggctagcgccagagagcttcaaaaggggaacgaactc

gcactaccgtctaaatacgtgaatttcctgtatttagcgtcccattac

gagaagttgaaaggttcacctgaagataacgaacagaagcaacttttt

gttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcg

gaattcagtaagagagtcatcctagctgatgccaatctggacaaagta

ttaagcgcatacaacaagcacagggataaacccatacgtgagcaggcg

gaaaatattatccatttgtttactcttaccaacctcggcgctccagcc

gcattcaagtattttgacacaacgatagatcgcaaagagtacagatct

accaaggaggtgctagacgcgacactgattcaccaatccatcacggga

ttatatgaaactcggatagatttgtcacagcttgggggtgacggatcc

cccaagaagaagaggaaagtcctcgagggcggaggcgggagcggatcc

ccctcccggctccagatgttcttcgctaataaccacgaccaggaattt

gaccctccaaaggtttacccacctgtcccagctgagaagaggaagccc

atccgggtgctgtctctctttgatggaatcgctacagggctcctggtg

ctgaaggacttgggcattcaggtggaccgctacattgcctcggaggtg

tgtgaggactccatcacggtgggcatggtgcggcaccaggggaagatc

atgtacgtcggggacgtccgcagcgtcacacagaagcatatccaggag

tggggcccattcgatctggtgattgggggcagtccctgcaatgacctc

tccatcgtcaaccctgctcgcaagggcctctacgagggcactggccgg

ctcttctttgagttctaccgcctcctgcatgatgcgcggcccaaggag

ggagatgatcgccccttcttctggctctttgcgaatgtggtggccatg

ggcgttagtgacaagagggacatctcgcgatttctcgagtccaaccct

gtgatgattgatgccaaagaagtgtcagctgcacacagggcccgctac

ttctggggtaaccttcccggtatgaacaggccgttggcatccactgtg

aatgataagctggagctgcaggagtgtctggagcatggcaggatagcc

aagttcagcaaagtgaggaccattactacgaggtcaaactccataaag

cagggcaaagaccagcattttcctgtgttcatgaatgagaaagaggac

atcttatggtgcactgaaatggaaagggtatttggtttcccagtccac

tatactgacgtgtccaacatgagccgcttggcgaggcagagactgctg

ggccggtcatggagcgtgccagtcatccgccacctcttcgctccgctg

aaggagtattttgcgtgtgtgtccggccggcccggatccggcgcaaca

aacttctctctgctgaaacaagccggagatgtcgaagagaatcctgga

ccgaccgagtacaagcccacggtgcgcctcgccacccgcgacgacgtc

cccagggccgtacgcaccctcgccgccgcgttcgccgactaccccgcc

acgcgccacaccgtcgatccggaccgccacatcgagcgggtcaccgag

ctgcaagaactcttcctcacgcgcgtcgggctcgacatcggcaaggtg

tgggtcgcggacgacggcgccgcggtggcggtctggaccacgccggag

agcgtcgaagcgggggcggtgttcgccgagatcggcccgcgcatggcc

gagttgagcggttcccggctggccgcgcagcaacagatggaaggcctc

ctggcgccgcaccggcccaaggagcccgcgtggttcctggccaccgtc

ggagtctcgcccgaccaccagggcaagggtctgggcagcgccgtcgtg

ctccccggagtggaggcggccgagcgcgccggggtgcccgccttcctg

gagacctccgcgccccgcaacctccccttctacgagcggctcggcttc

accgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacctggtgc

atgacccgcaagcccggtgcctgaacgcgttaagtcgacaatcaacct

ctggattacaaaatttgtgaaagattgactggtattcttaactatgtt

gctccttttacgctatgtggatacgctgctttaatgcctttgtatcat

gctattgcttcccgtatggctttcattttctcctccttgtataaatcc

tggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgt

ggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggc

attgccaccacctgtcagctcctttccgggactttcgctttccccctc

cctattgccacggcggaactcatcgccgcctgccttgcccgctgctgg

acaggggctcggctgttgggcactgacaattccgtggtgttgtcgggg

aaatcatcgtcctttccttggctgctcgcctgtgttgccacctggatt

ctgcgcgggacgtccttctgctacgtcccttcggccctcaatccagcg

gaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgt

cttcgccttcgccctcagacgagtcggatctccctttgggccgcctcc

ccgcgtcgactttaagaccaatgacttacaaggcagctgtagatctta

gccactttttaaaagaaaaggggggactggaagggctaattcactccc

aacgaagacaagatctgctttttgcttgtactgggtctctctggttag

accagatctgagcctgggagctctctggctaactagggaacccactgc

ttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgccc

gtctgttgtgtgactctggtaactagagatccctcagacccttttagt

cagtgtggaaaatctctagcagggcccgtttaaacccgctgatcagcc

tcgactgtgccttctagttgccagccatctgttgtttgcccctccccc

gtgccttccttgaccctggaaggtgccactcccactgtcctttcctaa

taaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctatt

ctggggggtggggggggcaggacagcaagggggaggattgggaagaca

atagcaggcatgctggggatgcggtgggctctatggcttctgaggcgg

aaagaaccagctggggctctagggggtatccccacgcgccctgtagcg

gcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgaccgcta

cacttgccagcgccctagcgcccgctcctttcgctttcttcccttcct

ttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggc

tccctttagggttccgatttagtgctttacggcacctcgaccccaaaa

aacttgattagggtgatggttcacgtagtgggccatcgccctgataga

cggtttttcgccctttgacgttggagtccacgttctttaatagtggac

tcttgttccaaactggaacaacactcaaccctatctcggtctattctt

ttgatttataagggattttgccgatttcggcctattggttaaaaaatg

agctgatttaacaaaaatttaacgcgaattaattctgtggaatgtgtg

tcagttagggtgtggaaagtccccaggctccccagcaggcagaagtat

gcaaagcatgcatctcaattagtcagcaaccaggtgtggaaagtcccc

aggctccccagcaggcagaagtatgcaaagcatgcatctcaattagtc

agcaaccatagtcccgcccctaactccgcccatcccgcccctaactcc

gcccagttccgcccattctccgccccatggctgactaattttttttat

ttatgcagaggccgaggccgcctctgcctctgagctattccagaagta

gtgaggaggcttttttggaggcctaggcttttgcaaaaagctccctac

cgtcgacctctagctagagcttggcgtaatcatggtcatagctgtttc

ctgtgtgaaattgttatccgctcacaattccacacaacatacgagccg

gaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactca

cattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgt

cgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtt

tgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgct

cggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaa

tacggttatccacagaatcaggggataacgcaggaaagaacatgtgag

caaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgac

gctcaagtcagaggtggcgaaacccgacaggactataaagataccagg

cgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgc

cgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgc

tttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttc

gctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgct

gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcga

ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacg

gctacactagaagaacagtatttggtatctgcgctctgctgaagccag

ttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgca

gaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctg

acgctcagtggaacgaaaactcacgttaagggattttggtcatgagat

tatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagtt

ttaaatcaatctaaagtatatatgagtaaacttggtctgacagttacc

aatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt

catccatagttgcctgactccccgtcgtgtagataactacgatacggg

agggcttaccatctggccccagtgctgcaatgataccgcgagacccac

gctcaccggctccagatttatcagcaataaaccagccagccggaaggg

ccgagcgcagaagtggtcctgcaactttatccgcctccatccagtcta

ttaattgttgccgggaagctagagtaagtagttcgccagttaatagtt

tgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgt

cgtttggtatggcttcattcagctccggttcccaacgatcaaggcgag

ttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtc

ctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatgg

ttatggcagcactgcataattctcttactgtcatgccatccgtaagat

gcttttctgtgactggtgagtactcaaccaagtcattctgagaatagt

gtatgcggcgaccgagttgctcttgcccggcgtcaatacgggataata

ccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgtt

cttcggggcgaaaactctcaaggatcttaccgctgttgagatccagtt

cgatgtaacccactcgtgcacccaactgatcttcagcatcttttactt

tcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgcaa

aaaagggaataagggcgacacggaaatgttgaatactcatactcttcc

tttttcaatattattgaagcatttatcagggttattgtctcatgagcg

gatacatatttgaatgtatttagaaaaataaacaaataggggttccgc

gcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1531 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 59)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cggcggtactgcaccaggcggcgttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

cccaaagaagaaacggaaggtgggtggaggaagtggcgggtcaggtgg

ctctagacggacactggtgaccttcaaggatgtatttgtggacttcac

cagggaggagtggaagctgctggacactgctcagcagatcgtgtacag

aaatgtgatgctggagaactataagaacctggtttccttgggttatca

gcttactaagccagatgtgatcctccggttggagaagggagaagagcc

ctcgggaggtggttcgggaggtggttcggagggtgtgcaggtgaaaag

ggtcctggagaaaagtcctgggaagctccttgtcaagatgccttttca

aacttcgccagggggcaaggctgaggggggtggggccaccacatccac

ccaggtcatggtgatcaaacgccccggcaggaagcgaaaagctgaggc

cgaccctcaggccattcccaagaaacggggccgaaagccggggagtgt

ggtggcagccgctgccgccgaggccaaaaagaaagccgtgaaggagtc

ttctatccgatctgtgcaggagacagtactccccatcaagaagcgcaa

gacccgggagggcgcgcccaagaagaagaggaaagtctccggatccgg

cgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaa

tcctggaccgaccgagtacaagcccacggtgcgcctcgccacccgcga

cgacgtccccagggccgtacgcaccctcgccgccgcgttcgccgacta

ccccgccacgcgccacaccgtcgatccggaccgccacatcgagcgggt

caccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcgg

caaggtgtgggtcgcggacgacggcgccgcggtggcggtctggaccac

gccggagagcgtcgaagcgggggcggtgttcgccgagatcggcccgcg

catggccgagttgagcggttcccggctggccgcgcagcaacagatgga

aggcctcctggcgccgcaccggcccaaggagcccgcgtggttcctggc

caccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgc

cgtcgtgctccccggagtggaggcggccgagcgcgccggggtgcccgc

cttcctggagacctccgcgccccgcaacctccccttctacgagcggct

cggcttcaccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcac

ctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaa

tcaacctctggattacaaaatttgtgaaagattgactggtattcttaa

ctatgttgctccttttacgctatgtggatacgctgctttaatgccttt

gtatcatgctattgcttcccgtatggctttcattttctcctccttgta

taaatcctggttgctgtctctttatgaggagttgtggcccgttgtcag

gcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactgg

ttggggcattgccaccacctgtcagctcctttccgggactttcgcttt

ccccctccctattgccacggcggaactcatcgccgcctgccttgcccg

ctgctggacaggggctcggctgttgggcactgacaattccgtggtgtt

gtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccac

ctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaa

tccagcggaccttccttcccgcggcctgctgccggctctgcggcctct

tccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggc

cgcctccccgcgtcgactttaagaccaatgacttacaaggcagctgta

gatcttagccactttttaaaagaaaaggggggactggaagggctaatt

cactcccaacgaagacaagatctgctttttgcttgtactgggtctctc

tggttagaccagatctgagcctgggagctctctggctaactagggaac

ccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtg

tgtgcccgtctgttgtgtgactctggtaactagagatccctcagaccc

ttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctg

atcagcctcgactgtgccttctagttgccagccatctgttgtttgccc

ctcccccgtgccttccttgaccctggaaggtgccactcccactgtcct

ttcctaataaaatgaggaaattgcatcgcattgtctgagtaggtgtca

ttctattctggggggtggggggggcaggacagcaagggggaggattgg

gaagacaatagcaggcatgctggggatgcggtgggctctatggcttct

gaggcggaaagaaccagctggggctctagggggtatccccacgcgccc

tgtagcggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtg

accgctacacttgccagcgccctagcgcccgctcctttcgctttcttc

ccttcctttctcgccacgttcgccggctttccccgtcaagctctaaat

cgggggctccctttagggttccgatttagtgctttacggcacctcgac

cccaaaaaacttgattagggtgatggttcacgtagtgggccatcgccc

tgatagacggtttttcgccctttgacgttggagtccacgttctttaat

agtggactcttgttccaaactggaacaacactcaaccctatctcggtc

tattcttttgatttataagggattttgccgatttcggcctattggtta

aaaaatgagctgatttaacaaaaatttaacgcgaattaattctgtgga

atgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggca

gaagtatgcaaagcatgcatctcaattagtcagcaaccaggtgtggaa

agtccccaggctccccagcaggcagaagtatgcaaagcatgcatctca

attagtcagcaaccatagtcccgcccctaactccgcccatcccgcccc

taactccgcccagttccgcccattctccgccccatggctgactaattt

tttttatttatgcagaggccgaggccgcctctgcctctgagctattcc

agaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagc

tccctaccgtcgacctctagctagagcttggcgtaatcatggtcatag

ctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagc

taactcacattaattgcgttgcgctcactgcccgctttccagtcggga

aacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggaga

ggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcg

ctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaag

gcggtaatacggttatccacagaatcaggggataacgcaggaaagaac

atgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcg

ttgctggcgtttttccataggctccgcccccctgacgagcatcacaaa

aatcgacgctcaagtcagaggtggcgaaacccgacaggactataaaga

taccaggcgtttccccctggaagctccctcgtgcgctctcctgttccg

accctgccgcttaccggatacctgtccgcctttctcccttcgggaagc

gtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtag

gtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagccc

gaccgctgcgccttatccggtaactatcgtcttgagtccaacccggta

agacacgacttatcgccactggcagcagccactggtaacaggattagc

agagcgaggtatgtaggcggtgctacagagttcttgaagtggtggcct

aactacggctacactagaagaacagtatttggtatctgcgctctgctg

aagccagttaccttcggaaaaagagttggtagctcttgatccggcaaa

caaaccaccgctggtagcggtggtttttttgtttgcaagcagcagatt

acgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacg

gggtctgacgctcagtggaacgaaaactcacgttaagggattttggtc

atgagattatcaaaaaggatcttcacctagatccttttaaattaaaaa

tgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtcta

tttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcga

gacccacgctcaccggctccagatttatcagcaataaaccagccagcc

ggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatc

cagtctattaattgttgccgggaagctagagtaagtagttcgccagtt

aatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtca

cgctcgtcgtttggtatggcttcattcagctccggttcccaacgatca

aggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctcc

ttcggtcctccgatcgttgtcagaagtaagttggccgcagtgttatca

ctcatggttatggcagcactgcataattctcttactgtcatgccatcc

gtaagatgcttttctgtgactggtgagtactcaaccaagtcattctga

gaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgg

gataataccgcgccacatagcagaactttaaaagtgctcatcattgga

aaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgaga

tccagttcgatgtaacccactcgtgcacccaactgatcttcagcatct

tttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaat

gccgcaaaaaagggaataagggcgacacggaaatgttgaatactcata

ctcttcctttttcaatattattgaagcatttatcagggttattgtctc

atgagcggatacatatttgaatgtatttagaaaaataaacaaataggg

gttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1532 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth

(SEQ ID NO: 60)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaacaaaagtaagaccaccgcacagcaagcggccgctgatc

ttcagacctggaggaggagatatgagggacaattggagaagtgaatta

tataaatataaagtagtaaaaattgaaccattaggagtagcacccacc

aaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaata

ggagctttgttccttgggttcttgggagcagcaggaagcactatgggc

gcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt

atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacag

catctgttgcaactcacagtctggggcatcaagcagctccaggcaaga

atcctggctgtggaaagatacctaaaggatcaacagctcctggggatt

tggggttgctctggaaaactcatttgcaccactgctgtgccttggaat

gctagttggagtaataaatctctggaacagatttggaatcacacgacc

tggatggagtgggacagagaaattaacaattacacaagcttaatacac

tccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaa

ttattggaattagataaatgggcaagtttgtggaattggtttaacata

acaaattggctgtggtatataaaattattcataatgatagtaggaggc

ttggtaggtttaagaatagtttttgctgtactttctatagtgaataga

gttaggcagggatattcaccattatcgtttcagacccacctcccaacc

ccgaggggacccgacaggcccgaaggaatagaagaagaaggtggagag

agagacagagacagatccattcgattagtgaacggatcggcactgcgt

gcgccaattctgcagacaaatggcagtattcatccacaattttaaaag

aaaaggggggattggggggtacagtgcaggggaaagaatagtagacat

aatagcaacagacatacaaactaaagaattacaaaaacaaattacaaa

aattcaaaattttcgggtttattacagggacagcagagatccagtttg

gttaattaatggggggacgttaacggggcggaacggtaccgagggcct

atttcccatgattccttcatatttgcatatacgatacaaggctgttag

agagataattagaattaatttgactgtaaacacaaagatattagtaca

aaatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttt

aaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaa

gtatttcgatttcttggctttatatatcttgtggaaaggacgaaacac

cggggcgcggacatggaggacggttttagagctagaaatagcaagtta

aaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggt

gcttttttgaattcgctagctaggtcttgaaaggagtgggaattggct

ccggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgaga

agttggggggaggggtcggcaattgatccggtgcctagagaaggtggc

gcggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttc

ccgaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgt

tctttttcgcaacgggtttgccgccagaacacaggaccggtgccacca

tggactataaggaccacgacggagactacaaggatcatgatattgatt

acaaagacgatgacgataagatggccccaaagaagaagcggaaggtcg

gtatccacggagtcccagcagccgacaagaagtacagcatcggcctgg

ccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtaca

aggtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcaca

gcatcaagaagaacctgatcggagccctgctgttcgacagcggcgaaa

cagccgaggccacccggctgaagagaaccgccagaagaagatacacca

gacggaagaaccggatctgctatctgcaagagatcttcagcaacgaga

tggccaaggtggacgacagcttcttccacagactggaagagtccttcc

tggtggaagaggataagaagcacgagcggcaccccatcttcggcaaca

tcgtggacgaggtggcctaccacgagaagtaccccaccatctaccacc

tgagaaagaaactggtggacagcaccgacaaggccgacctgcggctga

tctatctggccctggcccacatgatcaagttccggggccacttcctga

tcgagggcgacctgaaccccgacaacagcgacgtggacaagctgttca

tccagctggtgcagacctacaaccagctgttcgaggaaaaccccatca

acgccagcggcgtggacgccaaggccatcctgtctgccagactgagca

agagcagacggctggaaaatctgatcgcccagctgcccggcgagaaga

agaatggcctgttcggcaacctgattgccctgagcctgggcctgaccc

ccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagc

tgagcaaggacacctacgacgacgacctggacaacctgctggcccaga

tcggcgaccagtacgccgacctgtttctggccgccaagaacctgtccg

acgccatcctgctgagcgacatcctgagagtgaacaccgagatcacca

aggcccccctgagcgcctctatgatcaagagatacgacgagcaccacc

aggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgaga

agtacaaagagattttcttcgaccagagcaagaacggctacgccggct

acattgacggcggagccagccaggaagagttctacaagttcatcaagc

ccatcctggaaaagatggacggcaccgaggaactgctcgtgaagctga

acagagaggacctgctgcggaagcagcggaccttcgacaacggcagca

tcccccaccagatccacctgggagagctgcacgccattctgcggcggc

aggaagatttttacccattcctgaaggacaaccgggaaaagatcgaga

agatcctgaccttccgcatcccctactacgtgggccctctggccaggg

gaaacagcagattcgcctggatgaccagaaagagcgaggaaaccatca

ccccctggaacttcgaggaagtggtggacaagggcgcttccgcccaga

gcttcatcgagcggatgaccaacttcgataagaacctgcccaacgaga

aggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtata

acgagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccg

ccttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttca

agaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttca

agaaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatc

ggttcaacgcctccctgggcacataccacgatctgctgaaaattatca

aggacaaggacttcctggacaatgaggaaaacgaggacattctggaag

atatcgtgctgaccctgacactgtttgaggacagagagatgatcgagg

aacggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagc

agctgaagcggcggagatacaccggctggggcaggctgagccggaagc

tgatcaacggcatccgggacaagcagtccggcaagacaatcctggatt

tcctgaagtccgacggcttcgccaacagaaacttcatgcagctgatcc

acgacgacagcctgacctttaaagaggacatccagaaagcccaggtgt

ccggccagggcgatagcctgcacgagcacattgccaatctggccggca

gccccgccattaagaagggcatcctgcagacagtgaaggtggtggacg

agctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcg

aaatggccagagagaaccagaccacccagaagggacagaagaacagcc

gcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagcc

agatcctgaaagaacaccccgtggaaaacacccagctgcagaacgaga

agctgtacctgtactacctgcagaatggggggatatgtacgtggacca

ggaactggacatcaaccggctgtccgactacgatgtggacgctatcgt

gcctcagagctttctgaaggacgactccatcgacaacaaggtgctgac

cagaagcgacaagaaccggggcaagagcgacaacgtgccctccgaaga

ggtcgtgaagaagatgaagaactactggcggcagctgctgaacgccaa

gctgattacccagagaaagttcgacaatctgaccaaggccgagagagg

cggcctgagcgaactggataaggccggcttcatcaagagacagctggt

ggaaacccggcagatcacaaagcacgtggcacagatactagattcccg

aatgaatacgaaatacgacgagaacgataagctgattcgggaagtcaa

agtaatcactttaaagtcaaaattggtgtcggacttcagaaaggattt

tcaattctataaagttagggagataaataactaccaccatgcgcacga

cgcttatcttaatgccgtcgtagggaccgcactcattaagaaataccc

gaagctagaaagtgagtttgtgtatggtgattacaaagtttatgacgt

ccgtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagc

caaatacttcttttattctaacattatgaatttctttaagacggaaat

cactctggcaaacggagagatacgcaaacgacctttaattgaaaccaa

tggggagacaggtgaaatcgtatgggataagggccgggacttcgcgac

ggtgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaac

tgaggtgcagaccggagggttttcaaaggaatcgattcttccaaaaag

gaatagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaa

gtacggtggcttcgtgagccctacagttgcctattctgtcctagtagt

ggcaaaagttgagaagggaaaatccaagaaactgaagtcagtcaaaga

attattggggataacgattatggagcgctcgtcttttgaaaagaaccc

catcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatct

cataattaaactaccaaagtatagtctgtttgagttagaaaatggccg

aaaacggatgttggctagcgccagagagcttcaaaaggggaacgaact

cgcactaccgtctaaatacgtgaatttcctgtatttagcgtcccatta

cgagaagttgaaaggttcacctgaagataacgaacagaagcaactttt

tgttgagcagcacaaacattatctcgacgaaatcatagagcaaatttc

ggaattcagtaagagagtcatcctagctgatgccaatctggacaaagt

attaagcgcatacaacaagcacagggataaacccatacgtgagcaggc

ggaaaatattatccatttgtttactcttaccaacctcggcgctccagc

cgcattcaagtattttgacacaacgatagatcgcaaagagtacagatc

taccaaggaggtgctagacgcgacactgattcaccaatccatcacggg

attatatgaaactcggatagatttgtcacagcttgggggtgacggatc

cccaaagaagaaacggaaggtgggtggaggaagtggcgggtcaggtgg

ctctagacggacactggtgaccttcaaggatgtatttgtggacttcac

cagggaggagtggaagctgctggacactgctcagcagatcgtgtacag

aaatgtgatgctggagaactataagaacctggtttccttgggttatca

gcttactaagccagatgtgatcctccggttggagaagggagaagagcc

ctcgggaggtggttcgggaggtggttcggagggtgtgcaggtgaaaag

ggtcctggagaaaagtcctgggaagctccttgtcaagatgccttttca

aacttcgccagggggcaaggctgaggggggtggggccaccacatccac

ccaggtcatggtgatcaaacgccccggcaggaagcgaaaagctgaggc

cgaccctcaggccattcccaagaaacggggccgaaagccggggagtgt

ggtggcagccgctgccgccgaggccaaaaagaaagccgtgaaggagtc

ttctatccgatctgtgcaggagacagtactccccatcaagaagcgcaa

gacccgggagggcgcgcccaagaagaagaggaaagtctccggatccgg

cgcaacaaacttctctctgctgaaacaagccggagatgtcgaagagaa

tcctggaccgaccgagtacaagcccacggtgcgcctcgccacccgcga

cgacgtccccagggccgtacgcaccctcgccgccgcgttcgccgacta

ccccgccacgcgccacaccgtcgatccggaccgccacatcgagcgggt

caccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcgg

caaggtgtgggtcgcggacgacggcgccgcggtggcggtctggaccac

gccggagagcgtcgaagcgggggcggtgttcgccgagatcggcccgcg

catggccgagttgagcggttcccggctggccgcgcagcaacagatgga

aggcctcctggcgccgcaccggcccaaggagcccgcgtggttcctggc

caccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgc

cgtcgtgctccccggagtggaggcggccgagcgcgccggggtgcccgc

cttcctggagacctccgcgccccgcaacctccccttctacgagcggct

cggcttcaccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcac

ctggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaa

tcaacctctggattacaaaatttgtgaaagattgactggtattcttaa

ctatgttgctccttttacgctatgtggatacgctgctttaatgccttt

gtatcatgctattgcttcccgtatggctttcattttctcctccttgta

taaatcctggttgctgtctctttatgaggagttgtggcccgttgtcag

gcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactgg

ttggggcattgccaccacctgtcagctcctttccgggactttcgcttt

ccccctccctattgccacggcggaactcatcgccgcctgccttgcccg

ctgctggacaggggctcggctgttgggcactgacaattccgtggtgtt

gtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccac

ctggattctgcgcgggacgtccttctgctacgtcccttcggccctcaa

tccagcggaccttccttcccgcggcctgctgccggctctgcggcctct

tccgcgtcttcgccttcgccctcagacgagtcggatctccctttgggc

cgcctccccgcgtcgactttaagaccaatgacttacaaggcagctgta

gatcttagccactttttaaaagaaaaggggggactggaagggctaatt

cactcccaacgaagacaagatctgctttttgcttgtactgggtctctc

tggttagaccagatctgagcctgggagctctctggctaactagggaac

ccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtg

tgtgcccgtctgttgtgtgactctggtaactagagatccctcagaccc

ttttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctg

atcagcctcgactgtgccttctagttgccagccatctgttgtttgccc

ctcccccgtgccttccttgaccctggaaggtgccactcccactgtcct

ttcctaataaaatgaggaaattgcatcgcattgtctgagtaggtgtca

ttctattctggggggtggggggggcaggacagcaagggggaggattgg

gaagacaatagcaggcatgctggggatgcggtgggctctatggcttct

gaggcggaaagaaccagctggggctctagggggtatccccacgcgccc

tgtagcggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtg

accgctacacttgccagcgccctagcgcccgctcctttcgctttcttc

ccttcctttctcgccacgttcgccggctttccccgtcaagctctaaat

cgggggctccctttagggttccgatttagtgctttacggcacctcgac

cccaaaaaacttgattagggtgatggttcacgtagtgggccatcgccc

tgatagacggtttttcgccctttgacgttggagtccacgttctttaat

agtggactcttgttccaaactggaacaacactcaaccctatctcggtc

tattcttttgatttataagggattttgccgatttcggcctattggtta

aaaaatgagctgatttaacaaaaatttaacgcgaattaattctgtgga

atgtgtgtcagttagggtgtggaaagtccccaggctccccagcaggca

gaagtatgcaaagcatgcatctcaattagtcagcaaccaggtgtggaa

agtccccaggctccccagcaggcagaagtatgcaaagcatgcatctca

attagtcagcaaccatagtcccgcccctaactccgcccatcccgcccc

taactccgcccagttccgcccattctccgccccatggctgactaattt

tttttatttatgcagaggccgaggccgcctctgcctctgagctattcc

agaagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagc

tccctaccgtcgacctctagctagagcttggcgtaatcatggtcatag

ctgtttcctgtgtgaaattgttatccgctcacaattccacacaacata

cgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagc

taactcacattaattgcgttgcgctcactgcccgctttccagtcggga

aacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggaga

ggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcg

ctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaag

gcggtaatacggttatccacagaatcaggggataacgcaggaaagaac

atgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcg

ttgctggcgtttttccataggctccgcccccctgacgagcatcacaaa

aatcgacgctcaagtcagaggtggcgaaacccgacaggactataaaga

taccaggcgtttccccctggaagctccctcgtgcgctctcctgttccg

accctgccgcttaccggatacctgtccgcctttctcccttcgggaagc

gtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtag

gtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagccc

gaccgctgcgccttatccggtaactatcgtcttgagtccaacccggta

agacacgacttatcgccactggcagcagccactggtaacaggattagc

agagcgaggtatgtaggcggtgctacagagttcttgaagtggtggcct

aactacggctacactagaagaacagtatttggtatctgcgctctgctg

aagccagttaccttcggaaaaagagttggtagctcttgatccggcaaa

caaaccaccgctggtagcggtggtttttttgtttgcaagcagcagatt

acgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacg

gggtctgacgctcagtggaacgaaaactcacgttaagggattttggtc

atgagattatcaaaaaggatcttcacctagatccttttaaattaaaaa

tgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgac

agttaccaatgcttaatcagtgaggcacctatctcagcgatctgtcta

tttcgttcatccatagttgcctgactccccgtcgtgtagataactacg

atacgggagggcttaccatctggccccagtgctgcaatgataccgcga

gacccacgctcaccggctccagatttatcagcaataaaccagccagcc

ggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatc

cagtctattaattgttgccgggaagctagagtaagtagttcgccagtt

aatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtca

cgctcgtcgtttggtatggcttcattcagctccggttcccaacgatca

aggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctcc

ttcggtcctccgatcgttgtcagaagtaagttggccgcagtgttatca

ctcatggttatggcagcactgcataattctcttactgtcatgccatcc

gtaagatgcttttctgtgactggtgagtactcaaccaagtcattctga

gaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacgg

gataataccgcgccacatagcagaactttaaaagtgctcatcattgga

aaacgttcttcggggcgaaaactctcaaggatcttaccgctgttgaga

tccagttcgatgtaacccactcgtgcacccaactgatcttcagcatct

tttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaat

gccgcaaaaaagggaataagggcgacacggaaatgttgaatactcata

ctcttcctttttcaatattattgaagcatttatcagggttattgtctc

atgagcggatacatatttgaatgtatttagaaaaataaacaaataggg

gttccgcgcacatttccccgaaaagtgccacctgac.

In an aspect, a disclosed pBK1536 plasmid can comprise the sequence set forth below or a sequence having 70%, 75%, 80%, 85%, 90%, 95%, or more identity to the sequence set forth below:

(SEQ ID NO: 61)

gtcgacggatcgggagatctcccgatcccctatggtgcactctcagta

caatctgctctgatgccgcatagttaagccagtatctgctccctgctt

gtgtgttggaggtcgctgagtagtgcgcgagcaaaatttaagctacaa

caaggcaaggcttgaccgacaattgcatgaagaatctgcttagggtta

ggcgttttgcgctgcttcgcgatgtacgggccagatatacgcgttgac

attgattattgactagttattaatagtaatcaattacggggtcattag

ttcatagcccatatatggagttccgcgttacataacttacggtaaatg

gcccgcctggctgaccgcccaacgacccccgcccattgacgtcaataa

tgacgtatgttcccatagtaacgccaatagggactttccattgacgtc

aatgggtggagtatttacggtaaactgcccacttggcagtacatcaag

tgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaat

ggcccgcctggcattatgcccagtacatgaccttatgggactttccta

cttggcagtacatctacgtattagtcatcgctattaccatggtgatgc

ggttttggcagtacatcaatgggcgtggatagcggtttgactcacggg

gatttccaagtctccaccccattgacgtcaatgggagtttgttttggc

accaaaatcaacgggactttccaaaatgtcgtaacaactccgccccat

tgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagca

gcgcgttttgcctgtactgggtctctctggttagaccagatctgagcc

tgggagctctctggctaactagggaacccactgcttaagcctcaataa

agcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgac

tctggtaactagagatccctcagacccttttagtcagtgtggaaaatc

tctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccag

aggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaa

gaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcg

gaggctagaaggagagagatgggtgcgagagcgtcagtattaagcggg

ggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaa

agaaaaaatataaattaaaacatatagtatgggcaagcagggagctag

aacgattcgcagttaatcctggcctgttagaaacatcagaaggctgta

gacaaatactgggacagctacaaccatcccttcagacaggatcagaag

aacttagatcattatataatacagtagcaaccctctattgtgtgcatc

aaaggatagagataaaagacaccaaggaagctttagacaagatagagg

aagagcaaaaaaaagtaagaccaccgcacagcaagcggccgctgatct

tcagacctggaggaggagatatgagggacaattggagaagtgaattat

ataaatataaagtagtaaaaattgaaccattaggagtagcacccacca

aggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaatag

gagctttgttccttgggttcttgggagcagcaggaagcactatgggcg

cagcgtcaatgacgctgacggtacaggccagacaattattgtctggta

tagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagc

atctgttgcaactcacagtctggggcatcaagcagctccaggcaagaa

tcctggctgtggaaagatacctaaaggatcaacagctcctggggattt

ggggttgctctggaaaactcatttgcaccactgctgtgccttggaatg

ctagttggagtaataaatctctggaacagatttggaatcacacgacct

ggatggagtgggacagagaaattaacaattacacaagcttaatacact

ccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaat

tattggaattagataaatgggcaagtttgtggaattggtttaacataa

caaattggctgtggtatataaaattattcataatgatagtaggaggct

tggtaggtttaagaatagtttttgctgtactttctatagtgaatagag

ttaggcagggatattcaccattatcgtttcagacccacctcccaaccc

cgaggggacccgacaggcccgaaggaatagaagaagaaggtggagaga

gagacagagacagatccattcgattagtgaacggatcggcactgcgtg

cgccaattctgcagacaaatggcagtattcatccacaattttaaaaga

aaaggggggattggggggtacagtgcaggggaaagaatagtagacata

atagcaacagacatacaaactaaagaattacaaaaacaaattacaaaa

attcaaaattttcgggtttattacagggacagcagagatccagtttgg

ttaattaatggggggacgttaacggggcggaacggtaccgagggccta

tttcccatgattccttcatatttgcatatacgatacaaggctgttaga

gagataattagaattaatttgactgtaaacacaaagatattagtacaa

aatacgtgacgtagaaagtaataatttcttgggtagtttgcagtttta

aaattatgttttaaaatggactatcatatgcttaccgtaacttgaaag

tatttcgatttcttggctttatatatcttgtggaaaggacgaaacacc

ggagacgtgtacacgtctctgttttagagctagaaatagcaagttaaa

ataaggctagtccgttatcaacttgaaaaagtggcaccgagtcggtgc

ttttttgaattcgctagctaggtcttgaaaggagtgggaattggctcc

ggtgcccgtcagtgggcagagcgcacatcgcccacagtccccgagaag

ttggggggaggggtcggcaattgatccggtgcctagagaaggtggcgc

ggggtaaactgggaaagtgatgtcgtgtactggctccgcctttttccc

gaggggggggagaaccgtatataagtgcagtagtcgccgtgaacgttc

tttttcgcaacgggtttgccgccagaacacaggaccggtgccaccatg

gactataaggaccacgacggagactacaaggatcatgatattgattac

aaagacgatgacgataagatggccccaaagaagaagcggaaggtcggt

atccacggagtcccagcagccgacaagaagtacagcatcggcctggcc

atcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaag

gtgcccagcaagaaattcaaggtgctgggcaacaccgaccggcacagc

atcaagaagaacctgatcggagccctgctgttcgacagcggcgaaaca

gccgaggccacccggctgaagagaaccgccagaagaagatacaccaga

cggaagaaccggatctgctatctgcaagagatcttcagcaacgagatg

gccaaggtggacgacagcttcttccacagactggaagagtccttcctg

gtggaagaggataagaagcacgagcggcaccccatcttcggcaacatc

gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctg

agaaagaaactggtggacagcaccgacaaggccgacctgcggctgatc

tatctggccctggcccacatgatcaagttccggggccacttcctgatc

gagggcgacctgaaccccgacaacagcgacgtggacaagctgttcatc

cagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaac

gccagcggcgtggacgccaaggccatcctgtctgccagactgagcaag

agcagacggctggaaaatctgatcgcccagctgcccggcgagaagaag

aatggcctgttcggcaacctgattgccctgagcctgggcctgaccccc

aacttcaagagcaacttcgacctggccgaggatgccaaactgcagctg

agcaaggacacctacgacgacgacctggacaacctgctggcccagatc

ggcgaccagtacgccgacctgtttctggccgccaagaacctgtccgac

gccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaag

gcccccctgagcgcctctatgatcaagagatacgacgagcaccaccag

gacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaag

tacaaagagattttcttcgaccagagcaagaacggctacgccggctac

attgacggcggagccagccaggaagagttctacaagttcatcaagccc

atcctggaaaagatggacggcaccgaggaactgctcgtgaagctgaac

agagaggacctgctgcggaagcagcggaccttcgacaacggcagcatc

ccccaccagatccacctgggagagctgcacgccattctgcggcggcag

gaagatttttacccattcctgaaggacaaccgggaaaagatcgagaag

atcctgaccttccgcatcccctactacgtgggccctctggccagggga

aacagcagattcgcctggatgaccagaaagagcgaggaaaccatcacc

ccctggaacttcgaggaagtggtggacaagggcgcttccgcccagagc

ttcatcgagcggatgaccaacttcgataagaacctgcccaacgagaag

gtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtataac

gagctgaccaaagtgaaatacgtgaccgagggaatgagaaagcccgcc

ttcctgagcggcgagcagaaaaaggccatcgtggacctgctgttcaag

accaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaag

aaaatcgagtgcttcgactccgtggaaatctccggcgtggaagatcgg

ttcaacgcctccctgggcacataccacgatctgctgaaaattatcaag

gacaaggacttcctggacaatgaggaaaacgaggacattctggaagat

atcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaa

cggctgaaaacctatgcccacctgttcgacgacaaagtgatgaagcag

ctgaagcggcggagatacaccggctggggcaggctgagccggaagctg

atcaacggcatccgggacaagcagtccggcaagacaatcctggatttc

ctgaagtccgacggcttcgccaacagaaacttcatgcagctgatccac

gacgacagcctgacctttaaagaggacatccagaaagcccaggtgtcc

ggccagggcgatagcctgcacgagcacattgccaatctggccggcagc

cccgccattaagaagggcatcctgcagacagtgaaggtggtggacgag

ctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcgaa

atggccagagagaaccagaccacccagaagggacagaagaacagccgc

gagagaatgaagcggatcgaagagggcatcaaagagctgggcagccag

atcctgaaagaacaccccgtggaaaacacccagctgcagaacgagaag

ctgtacctgtactacctgcagaatggggggatatgtacgtggaccagg

aactggacatcaaccggctgtccgactacgatgtggacgctatcgtgc

ctcagagctttctgaaggacgactccatcgacaacaaggtgctgacca

gaagcgacaagaaccggggcaagagcgacaacgtgccctccgaagagg

tcgtgaagaagatgaagaactactggcggcagctgctgaacgccaagc

tgattacccagagaaagttcgacaatctgaccaaggccgagagaggcg

gcctgagcgaactggataaggccggcttcatcaagagacagctggtgg

aaacccggcagatcacaaagcacgtggcacagatactagattcccgaa

tgaatacgaaatacgacgagaacgataagctgattcgggaagtcaaag

taatcactttaaagtcaaaattggtgtcggacttcagaaaggattttc

aattctataaagttagggagataaataactaccaccatgcgcacgacg

cttatcttaatgccgtcgtagggaccgcactcattaagaaatacccga

agctagaaagtgagtttgtgtatggtgattacaaagtttatgacgtcc

gtaagatgatcgcgaaaagcgaacaggagataggcaaggctacagcca

aatacttcttttattctaacattatgaatttctttaagacggaaatca

ctctggcaaacggagagatacgcaaacgacctttaattgaaaccaatg

gggagacaggtgaaatcgtatgggataagggccgggacttcgcgacgg

tgagaaaagttttgtccatgccccaagtcaacatagtaaagaaaactg

aggtgcagaccggagggttttcaaaggaatcgattcttccaaaaagga

atagtgataagctcatcgctcgtaaaaaggactgggacccgaaaaagt

acggtggcttcgtgagccctacagttgcctattctgtcctagtagtgg

caaaagttgagaagggaaaatccaagaaactgaagtcagtcaaagaat

tattggggataacgattatggagcgctcgtcttttgaaaagaacccca

tcgacttccttgaggcgaaaggttacaaggaagtaaaaaaggatctca

taattaaactaccaaagtatagtctgtttgagttagaaaatggccgaa

aacggatgttggctagcgccagagagcttcaaaaggggaacgaactcg

cactaccgtctaaatacgtgaatttcctgtatttagcgtcccattacg

agaagttgaaaggttcacctgaagataacgaacagaagcaactttttg

ttgagcagcacaaacattatctcgacgaaatcatagagcaaatttcgg

aattcagtaagagagtcatcctagctgatgccaatctggacaaagtat

taagcgcatacaacaagcacagggataaacccatacgtgagcaggcgg

aaaatattatccatttgtttactcttaccaacctcggcgctccagccg

cattcaagtattttgacacaacgatagatcgcaaagagtacagatcta

ccaaggaggtgctagacgcgacactgattcaccaatccatcacgggat

tatatgaaactcggatagatttgtcacagcttgggggtgacggatccc

caaagaagaaacggaaggtgggtggaggaagtggcgggtcaggtggct

ctagacggacactggtgaccttcaaggatgtatttgtggacttcacca

gggaggagtggaagctgctggacactgctcagcagatcgtgtacagaa

atgtgatgctggagaactataagaacctggtttccttgggttatcagc

ttactaagccagatgtgatcctccggttggagaagggagaagagccct

cgggaggtggttcgggaggtggttcggagggtgtgcaggtgaaaaggg

tcctggagaaaagtcctgggaagctccttgtcaagatgccttttcaaa

cttcgccagggggcaaggctgaggggggtggggccaccacatccaccc

aggtcatggtgatcaaacgccccggcaggaagcgaaaagctgaggccg

accctcaggccattcccaagaaacggggccgaaagccggggagtgtgg

tggcagccgctgccgccgaggccaaaaagaaagccgtgaaggagtctt

ctatccgatctgtgcaggagacagtactccccatcaagaagcgcaaga

cccgggagggcgcgcccaagaagaagaggaaagtctccggatccggcg

caacaaacttctctctgctgaaacaagccggagatgtcgaagagaatc

ctggaccgaccgagtacaagcccacggtgcgcctcgccacccgcgacg

acgtccccagggccgtacgcaccctcgccgccgcgttcgccgactacc

ccgccacgcgccacaccgtcgatccggaccgccacatcgagcgggtca

ccgagctgcaagaactcttcctcacgcgcgtcgggctcgacatcggca

aggtgtgggtcgcggacgacggcgccgcggtggcggtctggaccacgc

cggagagcgtcgaagcgggggcggtgttcgccgagatcggcccgcgca

tggccgagttgagcggttcccggctggccgcgcagcaacagatggaag

gcctcctggcgccgcaccggcccaaggagcccgcgtggttcctggcca

ccgtcggagtctcgcccgaccaccagggcaagggtctgggcagcgccg

tcgtgctccccggagtggaggcggccgagcgcgccggggtgcccgcct

tcctggagacctccgcgccccgcaacctccccttctacgagcggctcg

gcttcaccgtcaccgccgacgtcgaggtgcccgaaggaccgcgcacct

ggtgcatgacccgcaagcccggtgcctgaacgcgttaagtcgacaatc

aacctctggattacaaaatttgtgaaagattgactggtattcttaact

atgttgctccttttacgctatgtggatacgctgctttaatgcctttgt

atcatgctattgcttcccgtatggctttcattttctcctccttgtata

aatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggc

aacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggtt

ggggcattgccaccacctgtcagctcctttccgggactttcgctttcc

ccctccctattgccacggcggaactcatcgccgcctgccttgcccgct

gctggacaggggctcggctgttgggcactgacaattccgtggtgttgt

cggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacct

ggattctgcgcgggacgtccttctgctacgtcccttcggccctcaatc

cagcggaccttccttcccgcggcctgctgccggctctgcggcctcttc

cgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccg

cctccccgcgtcgactttaagaccaatgacttacaaggcagctgtaga

tcttagccactttttaaaagaaaaggggggactggaagggctaattca

ctcccaacgaagacaagatctgctttttgcttgtactgggtctctctg

gttagaccagatctgagcctgggagctctctggctaactagggaaccc

actgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtg

tgcccgtctgttgtgtgactctggtaactagagatccctcagaccctt

ttagtcagtgtggaaaatctctagcagggcccgtttaaacccgctgat

cagcctcgactgtgccttctagttgccagccatctgttgtttgcccct

cccccgtgccttccttgaccctggaaggtgccactcccactgtccttt

cctaataaaatgaggaaattgcatcgcattgtctgagtaggtgtcatt

ctattctggggggtggggggggcaggacagcaagggggaggattggga

agacaatagcaggcatgctggggatgcggtgggctctatggcttctga

ggcggaaagaaccagctggggctctagggggtatccccacgcgccctg

tagcggcgcattaagcgcggcgggtgtggtggttacgcgcagcgtgac

cgctacacttgccagcgccctagcgcccgctcctttcgctttcttccc

ttcctttctcgccacgttcgccggctttccccgtcaagctctaaatcg

ggggctccctttagggttccgatttagtgctttacggcacctcgaccc

caaaaaacttgattagggtgatggttcacgtagtgggccatcgccctg

atagacggtttttcgccctttgacgttggagtccacgttctttaatag

tggactcttgttccaaactggaacaacactcaaccctatctcggtcta

ttcttttgatttataagggattttgccgatttcggcctattggttaaa

aaatgagctgatttaacaaaaatttaacgcgaattaattctgtggaat

gtgtgtcagttagggtgtggaaagtccccaggctccccagcaggcaga

agtatgcaaagcatgcatctcaattagtcagcaaccaggtgtggaaag

tccccaggctccccagcaggcagaagtatgcaaagcatgcatctcaat

tagtcagcaaccatagtcccgcccctaactccgcccatcccgccccta

actccgcccagttccgcccattctccgccccatggctgactaattttt

tttatttatgcagaggccgaggccgcctctgcctctgagctattccag

aagtagtgaggaggcttttttggaggcctaggcttttgcaaaaagctc

cctaccgtcgacctctagctagagcttggcgtaatcatggtcatagct

gtttcctgtgtgaaattgttatccgctcacaattccacacaacatacg

agccggaagcataaagtgtaaagcctggggtgcctaatgagtgagcta

actcacattaattgcgttgcgctcactgcccgctttccagtcgggaaa

cctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagagg

cggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgct

gcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggc

ggtaatacggttatccacagaatcaggggataacgcaggaaagaacat

gtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgtt

gctggcgtttttccataggctccgcccccctgacgagcatcacaaaaa

tcgacgctcaagtcagaggtggcgaaacccgacaggactataaagata

ccaggcgtttccccctggaagctccctcgtgcgctctcctgttccgac

cctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgt

ggcgctttctcatagctcacgctgtaggtatctcagttcggtgtaggt

cgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccga

ccgctgcgccttatccggtaactatcgtcttgagtccaacccggtaag

acacgacttatcgccactggcagcagccactggtaacaggattagcag

agcgaggtatgtaggcggtgctacagagttcttgaagtggtggcctaa

ctacggctacactagaagaacagtatttggtatctgcgctctgctgaa

gccagttaccttcggaaaaagagttggtagctcttgatccggcaaaca

aaccaccgctggtagcggtggtttttttgtttgcaagcagcagattac

gcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggg

gtctgacgctcagtggaacgaaaactcacgttaagggattttggtcat

gagattatcaaaaaggatcttcacctagatccttttaaattaaaaatg

aagttttaaatcaatctaaagtatatatgagtaaacttggtctgacag

ttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatt

tcgttcatccatagttgcctgactccccgtcgtgtagataactacgat

acgggagggcttaccatctggccccagtgctgcaatgataccgcgaga

cccacgctcaccggctccagatttatcagcaataaaccagccagccgg

aagggccgagcgcagaagtggtcctgcaactttatccgcctccatcca

gtctattaattgttgccgggaagctagagtaagtagttcgccagttaa

tagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacg

ctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaag

gcgagttacatgatcccccatgttgtgcaaaaaagcggttagctcctt

cggtcctccgatcgttgtcagaagtaagttggccgcagtgttatcact

catggttatggcagcactgcataattctcttactgtcatgccatccgt

aagatgcttttctgtgactggtgagtactcaaccaagtcattctgaga

atagtgtatgcggcgaccgagttgctcttgcccggcgtcaatacggga

taataccgcgccacatagcagaactttaaaagtgctcatcattggaaa

acgttcttcggggcgaaaactctcaaggatcttaccgctgttgagatc

cagttcgatgtaacccactcgtgcacccaactgatcttcagcatcttt

tactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgc

cgcaaaaaagggaataagggcgacacggaaatgttgaatactcatact

cttcctttttcaatattattgaagcatttatcagggttattgtctcat

gagcggatacatatttgaatgtatttagaaaaataaacaaataggggt

tccgcgcacatttccccgaaaagtgccacctgac.

C. Methods of Administering Precision Gene Therapy

Disclosed herein is a method of administering precision gene therapy, the method comprising contacting one or more cells with a therapeutically effective amount of a disclosed isolated nucleic acid molecule, and reducing the activity and/or expression of APOE in one or more cells.

Disclosed herein is a method of administering precision gene therapy, the method comprising contacting one or more cells with a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing the activity and/or expression of APOE in one or more cells.

Disclosed herein is a method of administering precision gene therapy, the method comprising contacting one or more cells with a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing the activity and/or expression of the APOE e4 allele in one or more cells.

In an aspect of a disclosed method, increased APOE expression and/or activity can be mediated by a coding mutation in exon 4, gene dysregulation, or a combination thereof.

In an aspect, a disclosed method can reduce expression and/or activity of APOE regardless of the subject's genotype.

In an aspect, the disclosed cells can be neurons such as, for example, cholinergic neurons. In an aspect, the disclosed cells can be in a subject.

In an aspect, a disclosed viral vector can be a lentiviral vector. In an aspect of a disclosed method, a disclosed viral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In an aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect of a disclosed method, a disclosed viral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule and one or more regulatory elements. Regulatory elements are known in the art and can comprise one or more of the following: a Sp1 responsive element, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR. In an aspect, a disclosed viral vector can comprise two Sp1 response elements, a p2A signal, a woodchuck hepatitis virus post-transcriptional regulatory element, a Phi signal-packaging signal, a rev responsive element, a 5′-LTR, and a 3′-LTR.

In an aspect, a disclosed Cas endonuclease can be fused to a disclosed polypeptide having an enzymatic activity. In an aspect, a disclosed Cas endonuclease can be dCas9 and the polypeptide can be DNMT3A. In an aspect, a dCas9-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:19. In an aspect, a dCas9-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:20. In an aspect, a disclosed dCas9-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO: 19 or SEQ ID NO:20 or a fragment thereof. In an aspect, a disclosed Cas endonuclease can be dVRER and the polypeptide can be DNMT3A. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:38. In an aspect, a disclosed dVRER-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:37. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:38 or SEQ ID NO:37 or a fragment thereof.

In an aspect, a disclosed method can comprise reducing the pathological phenotype associated with Alzheimer's disease. In an aspect, reducing the pathological phenotype associated with Alzheimer's disease can comprise reducing the A042/40 ratio and reducing the level of Tau. In an aspect, a disclosed method can comprise diagnosing the subject with Alzheimer's disease.

In an aspect, a disclosed method can comprise repeating one or more steps of the method and/or modifying one or more steps of the method.

In an aspect of a disclosed method, administering a disclosed viral vector can comprise intravenous administration, intracerebral administration, intra-CSF administration, intracerebroventricular (ICV) administration, intraventricular administration, intra-cistema magna (ICM) administration, intraparenchymal administration, intrathecal (lumbar, cistemal, or both) administration, or any combination thereof.

In an aspect, a disclosed method can comprise administering to the subject a therapeutically effective amount of a therapeutic agent, an effective amount of an immune modulator, or a combination thereof.

Disclosed herein is a method of administering precision gene therapy, the method comprising administering to a subject in need thereof a therapeutically effective amount of a viral vector, wherein the viral vector comprises an isolated nucleic acid molecule comprising a nucleic acid sequence encoding (i) a Cas endonuclease, (ii) at least one polypeptide having an enzymatic activity, and (iii) at least one guide RNA, and reducing expression of the APOE e4 allele.

In an aspect, the disclosed cells can be neurons such as, for example, cholinergic neurons. In an aspect, the disclosed cells can be in a subject.

In an aspect, a disclosed viral vector can be a lentiviral vector. In an aspect of a disclosed method, a disclosed viral vector can comprise one or more promoters operably linked to the isolated nucleic acid molecule. In an aspect, a disclosed promoter can drive the expression of a gRNA, the Cas9 endonuclease, a polypeptide, or a combination thereof. In an aspect, a disclosed promoter can be a hU6 promoter and a disclosed hU6 promoter can drive expression of a gRNA. In an aspect, a disclosed promoter can be an EFS-NC promoter and a disclosed EFS-NC promoter can drive expression of the Cas endonuclease. In an aspect, a disclosed promoter can comprise a hU6 promoter, an EFS-NC promoter, or a combination thereof.

In an aspect, a disclosed Cas endonuclease can be dVRER and a disclosed polypeptide can be DNMT3A. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have the amino acid sequence set forth SEQ ID NO:38. In an aspect, a disclosed dVRER-DNMT3A fusion protein can be encoded by the sequence set forth in SEQ ID NO:37. In an aspect, a disclosed dVRER-DNMT3A fusion protein can have a sequence having at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identity to the sequence set forth in SEQ ID NO:38 or SEQ ID NO:37 or a fragment thereof.

In an aspect, a disclosed method can comprise repeating one or more steps of the method and/or modifying one or more steps of the method.

In an aspect of a disclosed method, administering a disclosed vector can comprise intravenous administration, intracerebral administration, intra-CSF administration, intracerebroventricular (ICV) administration, intraventricular administration, intra-cistema magna (ICM) administration, intraparenchymal administration, intrathecal (lumbar, cistemal, or both) administration, or any combination thereof.

D. Methods of Treating and/or Preventing Alzheimer's Disease Progression