BACKGROUND OF THE INVENTION
The HLA class I and class II loci are the most polymorphic genes in the human genome, with a complex pattern of patchwork polymorphism localized primarily in exon 2 for the class II genes and exons 2 and 3 for the class I genes. For current HLA typing methods, allele level resolution of HLA alleles, which is clinically important for hemapoetic stem cell transplantation, is technically challenging. Several large-scale studies have demonstrated that precise, allele-level HLA matching between donor and patient significantly improves overall transplant survival by reducing the incidence and severity of both acute and chronic graft versus host disease and improving the rates of successful engraftment. When, for example, 8 of 8 of the most significant HLA loci are matched vs 6 of 8, survival after transplant was enhanced by 60% after 12 months.
It is current practice to maintain bone marrow donor registries in which millions of potential donors are HLA typed at low-medium resolution for the A, B, and, in many cases, the DRB1 loci. Multiple potentially matched unrelated donors are selected, based on this initial typing, and then typed at allele level resolution at these and additional loci to identify the donor best matched to the recipient.
To date, the highest resolution HLA typing is obtained with fluorescent, Sanger-based DNA sequencing using capillary electrophoresis. Howver, ambiguities in the HLA typing data can persist due to multiple polymorphisms between alleles and the resultant phase ambiguities when both alleles are amplified and sequenced together. Resolving these ambiguities requires time-consuming approaches such as amplifying and then analyzing the two alleles separately.
Next-generation sequencing methods clonally propagate in parallel millions of single DNA molecules which are then also sequenced in parallel. Recently, the read lengths obtainable by one such next-generation pyrosequencing sequencing method (454 Life Sciences, Inc.) has increased to >250 nucleotides. The current invention provides improved HLA genotyping methods that are based on the discovery that clonal sequencing can be used for setting the phase of the linked polymorphisms within an exon and makes possible the unambiguous determination of the sequence of each HLA allele.
BRIEF SUMMARY OF THE INVENTION
The invention is based, in part, on the discovery that an 8-loci HLA genotyping can be performed on samples obtained from multiple subjections in a single sequencing run. In some embodiments, the invention therefore provides a method of determining the HLA genotypes for the HLA genes HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1 for more than one individual in parallel, the method comprising:
- (a) for each individual, amplifying the exons of the HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1 genes that comprises polymorphic sites to obtain HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1 amplicons for each individual, wherein each amplification reaction is performed with a forward primer and a reverse primer to amplify an HLA gene exon, where:
(i) the forward primer comprises the following sequences, from 5′ to 3″: an adapter sequence, a molecular identification sequence, and an HLA sequence; and
(ii) the reverse primer comprises the following sequences, from 5′ to 3″: an adapter sequence, a molecular identification sequence, and an HLA sequence;
- (b) pooling HLA amplicons from more than one individual and performing emulsion PCR;
- (c) determining the sequence of the HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1 amplicon for each individual using pyrosequencing in parallel; and
- (d) assigning the HLA alleles to each individual by comparing the sequence of the HLA amplicons to the known HLA sequence to determine which HLA alleles are present in the individual. In some embodiments, the forward or reverse primer for amplifying an HLA amplicon has the sequence of an HLA-hybridizing region of a primer set forth in Table 1. Such a primer may additionally comprise the sequence of an adapter region of a primer of Table 1. In further embodiments, the primer may also comprise an individual identification tag of a primer set forth in Table 1. In particular embodiments, the primer has a sequence of a primer set forth in Table 1.
In other embodiments, the invention provides a kit comprising primer pairs for obtaining HLA amplicons to determine the HLA genotypes for the HLA genes HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1 for more than one individual in parallel, wherein the primer pairs comprise a forward primer and a reverse primer to amplify an HLA gene exon, where: (i) the forward primer comprises the following sequences, from 5′ to 3″: an adapter sequence, a molecular identification sequence, and an HLA sequence; and (ii) the reverse primer comprises the following sequences, from 5′ to 3′: an adapter sequence, a molecular identification sequence, and an HLA sequence. In some embodiments, the kit comprises one ore more of the forward and reverse primers set forth in Table 1. In some embodiments, the kit comprises primer pairs to amplify exons for genotyping HLA genes HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1, wherein each of the primer pairs is selected from the primers set forth in Table 1.
The invention additionally provides a kit comprising one or more primer pairs, wherein each primer pair comprises a forward primer for obtaining an HLA amplicon that has the sequence of an HLA-hybridizing region of a primer set forth in Table 1; and a reverse primer for obtaining the HLA amplicon that has the sequence of an HLA-hybridizing region of a primer set forth in Table 1. Such a primer may additionally comprise an adapter region having a sequence set forth in Table 1. In further embodiments, the primer may have an individual identification tag of a primer set forth in Table 1. In particular embodiments, the forward primer has a sequence of a primer set forth in Table 1 and the reverse primer has a sequence of a primer set forth in Table 1.
In some embodiments, the invention provides a kit, wherein the kit comprises fifteen HLA primer pairs, where the primer pairs amplify exon 2, exon 3, and exon 4 of HLA-A; exon 2, exon 3, and exon 4 of HLA-B; exon 2, exon 3, and exon 4 of HLA-C; exon 2 of DRB1, exon 2 of DPB1, exon 2 of DPA1, exon 2 of DQA1; and exon 2 and exon 3 of DQB1. In some embodiments, the invention provides a kit that comprises at least six of the primer pairs, or at least, eight, nine, ten, eleven, twelve, thirteen, of fourteen of the primer pairs. In some embodiments, the primer pairs are selected from the primers set forth in Table 1.
In some embodiments, a kit of the invention comprises multiple primer pairs for each primer pair that amplifies exon 2, exon 3, and exon 4 of HLA-A; exon 2, exon 3, and exon 4 of HLA-B; exon 2, exon 3, and exon 4 of HLA-C; exon 2 of DRB1, exon 2 of DPB1, exon 2 of DPA1, exon 2 of DQA1; and exon 2 and exon 3 of DQB1, wherein the multiple primer pairs that amplify an individual exonic region of interest have the same HLA hybridizing region and the same adapter region, but different identification tags. In some embodiments, there are 12 or more multiple primer pairs for each exonic region of interest, where the primer pairs have different multiple identification tags.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 provides a schematic depicting a forward and reverse fusion primer of the invention.
FIG. 2 provides a histogram of the read length.
FIG. 3 shows the read depth for the total of forward and reverse reads.
DETAILED DESCRIPTION OF THE INVENTION
The term “allele”, as used herein, refers to a sequence variant of a gene. One or more genetic differences can constitute an allele. For HLA alleles, multiple genetic differences typically constitute an allele. Examples of HLA allele sequences are set out in Mason and Parham (1998) Tissue Antigens 51: 417-66, which list HLA-A, HLA-B, and HLA-C alleles and Marsh et al. (1992) Hum. Immunol. 35:1, which list HLA Class II alleles for DRA, DRB, DQA1, DQB1, DPA1, and DPB1.
The terms “polymorphic” and “polymorphism”, as used herein, refer to the condition in which two or more variants of a specific genomic sequence, or the encoded amino acid sequence, can be found in a population. A polymorphic position refers to a site in the nucleic acid where the nucleotide difference that distinguishes the variants occurs. As used herein, a “single nucleotide polymorphism”, or SNP, refers to a polymorphic site consisting of a single nucleotide position.
The term “genotype” refers to a description of the alleles of a gene or genes contained in an individual or a sample. As used herein, no distinction is made between the genotype of an individual and the genotype of a sample originating from the individual.
As used herein, “determining the genotype” of an HLA gene refers to determining the HLA polymorphisms present in the individual alleles of a subject. In the current invention, “determining the genotype of an HLA-A gene” refers to identifying the polymorphic residues present in at least exon 2 and exon 3, and typically exon 4, at positions that are allelic determinants of an HLA-A gene allele. In the current invention, “determining the genotype of an HLA-B gene” refers to identifying the polymorphic residues present in at least exon 2 and exon 3, and typically exon 4, at positions that are allelic determinants of an HLA-B gene allele; and “determining the genotype of an HLA-C gene” refers to identifying polymorphic residues present in at least exon 2 and exon 3, and typically exon 4, at positions that are allelic determinants of an HLA-C gene. Similarly, in the current invention, “determining the genotype” of a DRB1, DPB1, DPA1, or DQA1 gene refers to identifying the polymorphic residues present in exon 2 at t″ refers to identifying the polymorphic residues present in exon 2 and exon 3 at positions that are allelic determinants of a DQB1 allele.
As used herein an “allelic determinant” refers to a polymorphic site where the presence of variation results in variation in the HLA antigen.
The term “target region” refers to a region of a nucleic acid, in the current invention, an HLA gene, that is to be analyzed for the presence of polymorphic sites.
By “oligonucleotide” is meant a single-stranded nucleotide polymer made of more than 2 nucleotide subunits covalently joined together. An oligonucleotide primer as used herein is typically between about 10 and 100 nucleotides in length, usually from 20 to 60 nucleotides in length. The sugar groups of the nucleotide subunits may be ribose, deoxyribose or modified derivatives thereof such as o-methyl ribose. The nucleotide subunits of an oligonucleotide may be joined by phosphodiester linkages, phosphorothioate linkages, methyl phosphonate linkages or by other linkages, including but not limited to rare or non-naturally-occurring linkages, that do not prevent hybridization of the oligonucleotide. Furthermore, an oligonucleotide may have uncommon nucleotides or non-nucleotide moieties. An oligonucleotide as defined herein is a nucleic acid, preferably DNA, but may be RNA or have a combination of ribo- and deoxyribonucleotides covalently linked. Oligonucleotides of a defined sequence may be produced by techniques known to those of ordinary skill in the art, such as by chemical or biochemical synthesis, and by in vitro or in vivo expression from recombinant nucleic acid molecules.
The term “primer” refers to an oligonucleotide that acts as a point of initiation of DNA synthesis under conditions in which synthesis of a primer extension product complementary to a nucleic acid strand is induced in an appropriate buffer and at a suitable temperature. A primer is preferably a single-stranded oligodeoxyribonucleotide. In the current invention, a primer includes an “HLA-binding region” or HLA-hybridizing region” exactly or substantially complementary to the HLA sequence of interest. This region of the primer is typically about 15 to about 25, 30, 35 or 40 nucleotides in length.
As used herein, an “adapter region” of a primer refers to the region of a primer sequence at the 5′ end that is universal to the HLA amplicons obtained in accordance with the procedures described herein and provides sequences that anneal to an oligonucleotide present on a microparticle or other solid surface for emulsion PCR. The “adapter region” can further serve as a site to which a sequencing primer binds. The adapter region is typically from 15 to 30 nucleotides in length.
The terms “individual identifier tag”, “barcode”, “identification tag”, “multiplex identification tag”, “molecular identification tag” or “MID” are used interchangeably herein to refer to a nucleotide sequence present in a primer that serves as a marker of the DNA obtained from a particular subject.
As used herein, the terms “nucleic acid,” “polynucleotide” and “oligonucleotide” refer to primers and oligomer fragments. The terms are not limited by length and are generic to linear polymers of polydeoxyribonucleotides (containing 2-deoxy-D-ribose), polyribonucleotides (containing D-ribose), and any other N-glycoside of a purine or pyrimidine base, or modified purine or pyrimidine bases. These terms include double- and single-stranded DNA, as well as double- and single-stranded RNA.
A nucleic acid, polynucleotide or oligonucleotide can comprise phosphodiester linkages or modified linkages including, but not limited to phosphotriester, phosphoramidate, siloxane, carbonate, carboxymethylester, acetamidate, carbamate, thioether, bridged phosphoramidate, bridged methylene phosphonate, phosphorothioate, methylphosphonate, phosphorodithioate, bridged phosphorothioate or sulfone linkages, and combinations of such linkages.
A nucleic acid, polynucleotide or oligonucleotide can comprise the five biologically occurring bases (adenine, guanine, thymine, cytosine and uracil) and/or bases other than the five biologically occurring bases. These bases may serve a number of purposes, e.g., to stabilize or destabilize hybridization; to promote or inhibit probe degradation; or as attachment points for detectable moieties or quencher moieties. For example, a polynucleotide of the invention can contain one or more modified, non-standard, or derivatized base moieties, including, but not limited to, N6-methyl-adenine, N6-tert-butyl-benzyl-adenine, imidazole, substituted imidazoles, 5-fluorouracil, 5 bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xanthine, 4-acetylcytosine, 5 (carboxyhydroxymethyl)uracil, 5 carboxymethylaminomethyl-2-thiouridine, 5 carboxymethylaminomethyluracil, dihydrouracil, beta-D-galactosylqueosine, inosine, N6 isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methyladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-methyladenine, 7-methylguanine, 5-methylaminomethyluracil, 5-methoxyaminomethyl-2-thiouracil, beta-D mannosylqueosine, 5′-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N6-isopentenyladenine, uracil-5-oxyacetic acid (v), wybutoxosine, pseudouracil, queosine, 2 thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, uracil-5-oxyacetic acidmethylester, 3-(3-amino-3-N-2-carboxypropyl) uracil, (acp3)w, 2,6-diaminopurine, and 5-propynyl pyrimidine. Other examples of modified, non-standard, or derivatized base moieties may be found in U.S. Pat. Nos. 6,001,611; 5,955,589; 5,844,106; 5,789,562; 5,750,343; 5,728,525; and 5,679,785, each of which is incorporated herein by reference in its entirety. Furthermore, a nucleic acid, polynucleotide or oligonucleotide can comprise one or more modified sugar moieties including, but not limited to, arabinose, 2-fluoroarabinose, xylulose, and a hexose.
The term “amplification conditions” refers to conditions in an amplification reaction (e.g., a PCR amplification) that allow for hybridization of an extendable polynucleotide (e.g., a primer) with a target nucleotide, and the template-dependent extension of the extendable polynucleotide. As used herein, “amplification conditions” or conditions sufficient for amplifying a target nucleic acid are well known in the art. See, e.g., PCR Primer: A Laboratory Manual, by Dieffenbach and Dveksler, eds., 2003, Cold Spring Harbor Press; and PCR Protocols, Bartlett and Stirling, eds., 2003, Humana Press.
The term “amplification” as used here in the context of a nucleic acid amplification reaction refers to a reaction that increases the copies of a nucleic acid template, e.g., the target nucleic acid sequence.
Introduction
The current invention provides methods of HLA genotyping based the discovery that a multiplex, parallel clonal sequencing analysis can be used to genotype at least 3, typically at least 6, and preferably at least 8 HLA loci in multiple individuals at the same time. Next-generation sequencing methods clonally propagate in parallel millions of single DNA molecules which are then also sequenced in parallel. Recently, the read lengths obtainable by one such next-generation sequencing method (454 Life Sciences, Inc.) have increased to >250 nucleotides. These clonal read lengths make possible setting the phase of the linked polymorphisms within an exon and thus the unambiguous determination of the sequence of each HLA allele. In the current invention, the system is sufficiently high throughput to enable a complete, 8-locus HLA typing for multiple individuals, e.g., 24 or 48 subjects, in a single sequencing run using a pyrosequencing platform as described herein.
The highly multiplexed amplicon sequencing of the invention employs sample-specific internal sequence tags (barcode tags or MIDs) in the primers that allow pooling of samples yet maintain the ability to assign sequences to a specific individual. In the current invention, the HLA genotypes for at least eight loci (HLA-A, B, C, DRB1, DQA1, DQB1, DPA1, DPB1), as well as for DRB3,4, and 5 can be obtained from the data generated by sequencing. This HLA sequencing system can also detect chimeric mixtures, e.g., the detection of the rare non-transmitted maternal allele present in the blood of SCID patients.
HLA genes
The human leukocyte antigen system (HLA) complex spans approximately 3.5 million base pairs on the short arm of chromosome 6. The major regions are the class I and class II regions. The major Class I antigens are HLA-A, HLA-B, and HLA-C and the major Class II antigens are HLA-DP, HLA-DQ and HLA-DR. The HLA-DP, HLA-DQ and HLA-DR loci encode the α and β chains of the HLA-DR, DP and DQ antigens. The HLA genes are among the most polymorphic genes. Polymorphisms that are expressed in the HLA antigen (and therefore of great interest for typing for transplantation) are localized primarily in exon 2 for the class II genes and exons 2 and 3 for the class I genes.
In the current invention, the genotype of an HLA gene as described herein refers to determining the polymorphisms present in that HLA gene. For HLA-A, the polymorphisms present in exon 2 and exon 3 are determined by sequencing amplicons generated by PCR from an individual. In typical embodiments, the sequence of exon 4 is also determined. Exon 2, exon 3, and exon 4, or regions thereof that comprise the allelic determinants, are each amplified in individual PCR reactions to obtain amplicons. Similarly, amplicons are obtained for exon 2 and exon 3, and in some embodiments, exon 4, for the HLA-B and HLA-C alleles for an individual. For genotyping HLA class II alleles, amplicons are obtained for exon 2 of DRB1, DPB1, DPA1, DQA1 and exons 2 and 3 of DQB1. Each exon can be sequenced completely by sequencing both strands with sufficient overlap between the reads from either end that specific HLA alleles can be unambiguously assigned.
Each sample from an individual is amplified at each exon individually using primers that target the exon of interest, or the polymorphic region of the exon of interest, for amplification. The primers employed in the amplification reaction include additional sequences: adapter sequences for emulsion PCR and an identifying sequence that serves as a marker for the DNA from a single individual.
Amplification Primers
The invention employs amplification primers that amplify the exons of interest of the HLA genes. Typically, the primers are designed to ensure that the entire polymorphic portion of the exon is obtained.
In the current invention, primer sequences for the multiplex amplification of the invention are designed to include sequences that can be used to facilitate the clonal sequencing and the analysis. The amplification primers of the invention (also referred to herein as “fusion primers”) therefore include the following components: an adaptor, a unique identification tag and a sequence that hybridizes to an HLA gene of interest to use in an amplification reaction to obtain an HLA amplicon. FIG. 1 provides a schematic showing a fusion primer of the invention.
The adaptor portions of the primer sequences are present at the 5′ end of the amplicon fusion primers. The adapter regions comprise sequences that serve as the site of annealing of primers for the sequencing reaction and also correspond to sequences present on beads, or a solid surface, so that the amplicon can be annealed to the surface for emulsion PCR. The forward primer for amplifying an HLA exon includes an adapter sequence at the 5′ end, referred to here as the adapter region A. The reverse primer comprises a region that contains an adapter sequence at the 5′ end, referred to here as adapter reigon B. As noted, the sequences present in the adaptor region and their complements allow for annealing of the amplicons to beads for emulsion PCR. Optionally, the adaptor may further include a unique discriminating key sequence comprised of a non-repeating nucleotide sequence (i.e., ACGT, CAGT, etc.). This key sequence is typically incorporated to distinguish the amplicons for HLA genotyping from control sequences that are included in the reaction. Such sequences are described, e.g., in WO/2004/069849 and WO 2005/073410 Additional guidance for configuring adapter primers is provided, e.g., in WO/2006/110855.
In some embodiments, the adapter sequences for use in the invention are the primer A and primer B sequences for the 454 GS-FLX 454 sequencing system (Roche Diagnostics). The primer A sequence is 5′ GCCTCCCTCGCGCCA 3′. The primer B sequence is 5′ GCCTTGCCAGCCCGC 3′. As noted above, the primers typically contain additional “key” sequences that provide identifying sequencing to distinguish the amplicons from control sequences.
PCR primers for use in the HLA genotyping methods of the invention further comprise individual identifier tags. These individual identifier tags are used to mark the HLA amplicons from each individual who is being tested. The HLA sequences of interest are amplified from a nucleic acid sample from a subject to be genotyped. As explained above, the HLA exons, or regions of the exons, comprising the polymorphisms that act as allelic determinants are individually amplified. The amplicons obtained from the subject are marked with the same identification tag. The tag is included in the fusion primers that are used to amplify each amplicon for that subject. Accordingly, the identification tags are also sequenced in the sequencing reaction. The ID tags are present in the fusion primers used to obtain the HLA amplicons between the adapter region and the HLA priming region of the fusion primer.
Identification tags may vary in length. Typically, the tag is at least 4 or 5 nucleotides in length. In some applications, it may be desirable to have longer identification sequences, e.g., 6, 8, or 10 or more nucleotides in length. The use of such sequences is well know in the art. (see, eg., Thomas, et al. Nat. Med., 12:852-855, 2006; Parameswaran et al,. Nucl. Acids Res., 35:e130, 2007; Hofmann et al., Nucl. Acids Res. 35:e91, 2007). In most embodiments of this invention, the identification tag is from 4 to 10 nucleotides in length.
Individual identifier sequences can be designed taking into account certain parameters. For example, in designing a 4-residue ID tag, it is desirable to choose 4 bases that take into account the flow cycle of the nucleotides in the sequencing reaction. For example, if the nucleotides are added in the order T, A, C, and G, it is typically desirable to design the tag sequence such that a residue that is positive is followed by a residue that would be negative. Accordingly, in this example, if a tag sequence begins with an “A” residue such that the nucleotide incorporated in the sequencing reaction is T, the second residue in the tag sequence would be a nucleotide such that A would not be incorporated. In addition, it is desirable to avoid forming homopolymers, either within the tag sequence or through creating them based on the last base of the adapter region or the first base of the HLA-specific region of the fusion primer.
The HLA priming region (also referred to herein as HLA binding region, or HLA hybridizing region) of the fusion primers is the region of the primer that hybridizes to the HLA sequence of interest to amplify the desired exon (or in some embodiments, region of the exon). Typically, the HLA region of the fusion primer hybridizes to intronic sequence adjacent to the exon to be amplified in order to obtain the entire exon sequence. The HLA sequences are preferably selected to selectively amplify the HLA exon of interest, although in some embodiments, a primer pair may also amplify a highly similar region of a related HLA gene. For example, the primers for exon 2 of DRB1 described in the example section below also amplify the DRB3, DRB4, and DRB5 loci. The primers are selected such that the exon is amplified with sufficient specificity to allow unambiguous determination of the HLA genotype from the sequence.
Sequences of HLA genes and alleles are known and available through various databases, including GenBank and other gene databases and have been published (see e.g., Mason and Parham (1998) Tissue Antigens 51: 417-66, listing HLA-A, HLA-B, and HLA-C alleles; Marsh et al. (1992) Hum. Immunol. 35:1, listing HLA Class II alleles-DRA, DRB, DQA1, DQB1, DPA1, and DPB1).
The PCR primers can be designed based on principles known in the art. Strategies for primer design may be found throughout the scientific literature, for example, in Rubin, E. and A. A. Levy, Nucleic Acids Res, 1996.24 (18): p. 3538-45; and Buck et al., Biotechniques, 1999.27 (3): p. 528-36. For example, the HLA-specific region of the primer is typically about 20 nucleotides or greater, e.g., 20 to 35 nucleotides in length. Other parameters that are considered are G/C content, design considerations to avoid internal secondary structure, and prevent the formation of primer dimers, as well as melting temperatures (Tm).
Examples of primers for use in this invention are provided in Table 1. In Table 1, the forward primers have the 454 sequencing system “A” primer sequence at the 5′ end, followed by a four nucleotide key (TCAG), which together comprise the adapter region; followed by the identifier tag (4 nucleotides, unless otherwise noted); which is then followed by the region that hybridizes to the HLA gene indicated. The reverse primers have the 454 sequencing system “B” primer sequence at the 5′ end followed by the four nucleotide key TCAG″, which together comprise the adapter region, followed by the identifier tag region, followed by the HLA-specific region.
A primer used in the methods of the invention may comprise an HLA-hybridizing region of a primer set forth in Table 1. In other embodiments, such a primer may comprise a portion that is substantially identical to the sequence of an HLA hybridizing region set forth in Table 1. Thus, for example, a primer of the invention may comprise at least 10, 15, or 20 or more contiguous nucleotides of an HLA hybridizing region of a primer set forth in Table 1.
The HLA amplifications for each subject to be HLA genotyped are performed separately. The amplicons from the individual subject are then pooled for subsequent emulsion PCR and sequence analysis.
The template nucleic acid used to amplify the HLA amplicon of interest is typically from genomic DNA isolated from a subject to be genotyped. In the current method, more than one subject is HLA genotyped in parallel reactions. In the current invention, at least 12 subjects, and typically at least 16, 20, 24, 30, 36, or 48 subjects are HLA genotyped.
The HLA amplicons may be obtained using any type of amplification reaction. In the current invention, multiplex amplicons are typically made by PCR using primer pairs as described herein. It is typically desirable to use a polymerase with a low error rate, e.g., such as a high-fidelity Taq polymerase (Roche Diagnostics).
The PCR conditions can be optimized to determine suitable conditions for obtaining HLA amplicons from a subject. Each HLA amplicon may be individually amplified in separate PCR reactions. In some embodiments, the HLA amplicons for a subject may be obtained in one or more multiplex reactions that comprise primer pairs to amplify individual amplicons
Emulsion PCR
The HLA amplicons are attached to beads and subject to emulsion PCR. Emulsion PCR is known in the art (see, e.g., WO/2004/9849, WO 2005/073410, U.S. Patent Application Publication No. 20050130173, WO/2007/086935 and WO/2008/076842). In emulsion PCR, amplification is performed by attaching a template to be amplified, in the current invention, an HLA amplicon, to a solid support, preferably in the form of a generally spherical bead.
The HLA amplicon is attached to the bead by annealing the amplicon, via the adaptor region, to a primer attached to a bead. Thus, the bead is linked to a large number of a single primer species that is complementary to the HLA amplicon in the adapter portion. The beads are suspended in aqueous reaction mixture and then encapsulated in a water-in-oil emulsion. The emulsion is composed of discrete aqueous phase microdroplets, e.g., approximately 60 to 200 μm in diameter, enclosed by a thermostable oil phase. Oil is added and emulsion droplets are formed such that on average, the emulsion comprises only one target nucleic acid and one bead. Each microdroplet contains, preferably, amplification reaction solution (i.e., the reagents necessary for nucleic acid amplification, such as polymerase, salts, and appropriate primers, e.g., corresponding to the adaptor region).
In the current invention, emulsion PCR is typically performed with two populations of beads, as the HLA amplicons are sequenced in both directions. In one population of beads, a first primer corresponding to the adapter sequence present on the reverse primer is attached to a bead. In the second population, a second primer corresponding to the adapter sequence present on the forward primer is attached to a bead. Thus, a primer for use in the emulsion amplification reaction typically has the sequence of the adapter region, without additional sequences such as “key” sequences. The emulsion amplification reaction is typically performed asymmetrically. For example, a the PCR primers may be present in a 8:1 or 16:1 ratio (i.e., 8 or 16 of one primer to 1 of the second primer) to perform asymmetric PCR.
Following emulsion amplification, the beads that have the singled-stranded HLA amplicon template are isolated, e.g., via a moiety such as a biotin that is present on an amplification primer during the emulsion PCR, and the template is sequenced using DNA sequencing technology that is based on the detection of base incorporation by the release of a pyrophosphate and simultaneous enzymatic nucleotide degradation (described, e.g., in U.S. Pat. Nos. 6,274,320, 6,258,568 and 6,210,891).
Clonal amplicons are sequenced using a sequencing primer (e.g., primer A or primer B) and adding four different dNTPs or ddNTPs subjected to a polymerase reaction. As each dNTP or ddNTP is added to the primer extension product, a pyrophosphate molecule is released. Pyrophosphate release can be detected enzymatically, such as, by the generation of light in a luciferase-luciferin reaction. Additionally, a nucleotide degrading enzyme, such as apyrase, can be present during the reaction in order to degrade unincorporated nucleotides (see, e.g., U.S. Pat. No. 6,258,568.) In other embodiments, the reaction can be carried out in the presence of a sequencing primer, polymerase, a nucleotide degrading enzyme, deoxynucleotide triphosphates, and a pyrophosphate detection system comprising ATP sulfurylase and luciferase (see, e.g., U.S. Pat. No. 6,258,568).
Once the sequencing data is obtained for the sequence of the individual DNA molecules, the unambiguous exon sequence can be determined by comparing these sequence files to an HLA sequence database for the two HLA alleles The read lengths achieved by the GSFLX system (454 Life Sciences) (avg=250 bp) allow sufficient overlap for this determination of each exon. The assignment of genotypes at each locus based on the exon sequence data files can be performed, e.g., by a software developed by Conexio Genomics. An important aspect of the software is the ability to filter out related sequence reads (pseudogenes and other unwanted HLA genes) that were co-amplified by the primers along with the target sequence.
Kits
The compositions and reagents described herein can be packaged into kits. A kit of the invention typically comprises multiple primer pairs as described herein that are suitable for amplifying the regions of interest in an HLA allele. The primer pairs comprise a forward primer comprising an adapter region, an individual identification tag and an HLA hybridizing region; and a reverse primer that comprises an adaptter region, an individual identification tag, and an HLA hybridizing region. The kits of the invention often comprise primer pairs to amplify amplicons for determining the genotype of multiple subjects for at least HLA-A, HLA-B, and DRB1. Often, a kit of the invention comprises sufficient primer pairs to determine the genotype of HLA-A, HLA-B, HLA-C, DRB1, DQA1, DQB1, DPA1, and DPB1 genes for multiple individuals, e.g., 12 or more individuals.
In some embodiments, a kit can additionally comprise one or more populations of beads that have a primer attached that corresponds to an adapter regions that can be used in emulsion PCR. In some embodiments, a kit can comprise one or more reaction compartments comprising reagents suitable for performing a reaction selected at the discretion of a practitioner. For example, in some embodiments, a kit can comprise one or more reaction compartments comprising one more sequencing reagents.
The various components included in the kit are typically contained in separate containers, however, in some embodiments, one or more of the components can be present in the same container. Additionally, kits can comprise any combination of the compositions and reagents described herein. In some embodiments, kits can comprise additional reagents that may be necessary or optional for performing the disclosed methods. Such reagents include, but are not limited to, buffers, control polynucleotides, and the like.
In this application, the use of the singular includes the plural unless specifically stated otherwise. The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described in any way. While the present teachings are described in conjunction with various examples, it is not intended that the present teachings be limited to such embodiments.
EXAMPLES
Multiplex Pyrosequencing
The analysis of multiple HLA loci for multiple samples in a single 454 run is facilitated by the incorporation of molecular ID (MID) tags into the PCR primers. Table 1 shows the sequences of the 454 HLA-specific fusion primers with the adapter sequence (for bead capture) and a 4-base MID tag. Additional sequences are provided that gave a 5-base MID tag.
In an initial experiment 24 cell lines having known HLA genotypes (Table 2) were analyzed. In a subsequent experiment, 48 samples were analyzed.
Fifteen primer pairs were designed for the exons 2,3, and 4 of HLA-A, B and C loci, exon 2 of DRB1, DPB1, DPA1, DQA1, and exons 2 and 3 of DQB1. Primers with twelve different MID tags for each target sequence were designed for a total of 180 (15×12). The primers for exon 2 of DRB1 also amplify the DRB3, DRB4, and DRB5 loci, genes that are present on specific DRB1 haplotypes. Following amplification of the various samples, the PCR products were quantified by BioAnalyzer analysis, diluted to the appropriate concentration, and pooled for the emulsion PCR. Pyrosequencing runs of 24 and 48 individuals were achieved using 2 or 4 picotitre plate regions, respectively. The distribution of read lengths for all amplicons is shown in FIG. 2. The average length was 250 bp. This length is sufficient for the forward and reverse sequence reads to overlap, allowing unambiguous assignment of sequence to each exon and, ultimately, to each allele. The numbers of reads for each exon per individual are shown in FIG. 3.
Genotyping Software
To facilitate genotype assignment from these complex sequence data files, a software program was developed (Conexio Genomics) that compares the forward and reverse sequence reads derived from each exon to an HLA sequence database. The database also contains the sequence of HLA pseudogenes and related genes, allowing the filtering out of sequences generated from pseudogenes or from non classical HLA class I genes (e.g. HLA-E, F, G, and H).
Twenty four cell-line derived DNA samples of known HLA type, based on probe hybridization HLA typing and Sanger sequencing, were sequenced at all 8 loci (HLA-A, -B,-C,-DRB1,-DQA1,-DQB1, DPA1, DPB1). Exon 2 sequences of DRB3, DRB4, and DRB5 were also identified in the amplicons generated by the DRB primer pair. Subsequently, a run of 48 samples (24 cell line DNAs and 24 DNAs extracted from blood samples) were sequenced at the same loci and genotype assignments were generated from the sequence data by Conexio ATF software. The concordance of software genotype calls and previously determined HLA types was 99.4%.
Analysis of Chimeric Mixtures (Rare Variant Detection)
The very high number of sequence reads (n=300-350K) generated in a typical GSFLX run makes possible the detection of rare variant sequences present in the sample. To estimate the sensitivity to detect such sequences, we prepared mixtures of PCR products for exons 2 and 3 of HLA-A and HLA-B and exon 2 of DRB1 from two HLA homozygote samples in various proportions (1/1, 1/10,1/100, 1/1000). The rare variant present in mixtures of 1/00 could be detected reproducibly.
The blood of certain individuals is chimeric, with residual maternal cells present at very low levels in the child's circulation or rare fetal cells maintained in the mother's circulation (ref.) SCID patients often retain maternal cells at a very low level. When such patients are recipients of hemapoetic stem cell transplant, characterizing the level of this potential chimerism is clinically important. To mimic the SCIDS situation, in which maternal cells may be present in a child, we prepared mixtures of two heterozygous samples, which shared one allele, in various proportions. In this experiment, the rare variant could be detected.
Two SCIDS patients, who were recipients of HST transplants were also analyzed, along with their parents. In each case, the presence of the non-transmitted maternal allele could be detected.
Clonal sequencing, the analysis of amplicons generated from individual DNA molecules amplified in turn from HLA exons allows the unambiguous exon sequence determination and, by comparing these sequence files to an HLA sequence database, determination of the two HLA alleles The read lengths achieved by the GSFLX system (454 Life Sciences) (avg=250 bp) allow sufficient overlap for this determination of each exon. In the present examples, the assignment of genotypes at each locus based on the exon sequence data files was performed by a software (ATF) developed by Conexio Genomics. An important aspect of the software is the ability to filter out related sequence reads (pseudogenes and other unwanted HLA genes) that were co-amplified by the primers along with the target sequence. The software also filters out very rare sequence reads that may have been generated by an error in the initial PCR amplification of the target sequence from genomic DNA, errors in the emulsion PCR, or pyrosequencing errors. One well-documented category of pyrosequencing errors is in the length determination of homopolymer tracts. For example, we have observed, rare sequence reads containing a run of Gs when most sequence reads contained the correct run of—Gs.
The cost of a single GSFLX run is considerable. To make this system cost-effective for high resolution clinical HLA typing, multiple samples are analyzed at multiple loci in a single run. The use of MID tags and multiple regions of the picotitre plate makes running 24 or 48 samples analyzed at 8 loci possible, as described in these examples.
It is the very large number of sequence reads generated in parallel that allows this multiplex analysis of multiple individuals at multiple loci It also provides the capacity to detect rare variants sequences. In mixtures of PCR products from two different genomic DNA samples, HLA exon sequences present at 1/100 were reliably detected. Related but unwanted sequences as well as rare sequences containing errors can also be filtered out. (Most HLA alleles differ from one another by multiple polymorphisms while the sequences containing errors typically differ from the correct sequence by only one nucleotide.)
Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be readily apparent to those of ordinary skill in the art in light of the teachings of this invention that certain changes and modifications may be made thereto without departing from the spirit or scope of the appended claims.
All publications, patents, accession number, and patent applications cited in this specification are herein incorporated by reference as if each individual publication or patent application were specifically and individually indicated to be incorporated by reference.
TABLE 1
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|
(SEQ ID NOS:5-592)
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dna_designation
sequence
n_mer
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|
HLA-A Exon 1-2 Forward
PM1216TCAT
GCCTCCCTCGCGCCATCAGTCATCTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216TGAT
GCCTCCCTCGCGCCATCAGTGATCTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216TGCT
GCCTCCCTCGCGCCATCAGTGCTCTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216TGCA
GCCTCCCTCGCGCCATCAGTGCACTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216CAGA
GCCTCCCTCGCGCCATCAGCAGACTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216CTCT
GCCTCCCTCGCGCCATCAGCTCTCTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216CTCA
GCCTCCCTCGCGCCATCAGCTCACTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216CTGA
GCCTCCCTCGCGCCATCAGCTGACTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216ATCA
GCCTCCCTCGCGCCATCAGATCACTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216ATCT
GCCTCCCTCGCGCCATCAGATCTCTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216ATGA
GCCTCCCTCGCGCCATCAGATGACTCCCCAGACSCCGAGGATGGCC
46
|
|
PM1216AGCA
GCCTCCCTCGCGCCATCAGAGCACTCCCCAGACSCCGAGGATGGCC
46
|
|
HLA-A Exon 1-2 Reverse
PM1219TCAT
GCCTTGCCAGCCCGCTCAGTCATGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219TGAT
GCCTTGCCAGCCCGCTCAGTGATGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219TGCT
GCCTTGCCAGCCCGCTCAGTGCTGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219TGCA
GCCTTGCCAGCCCGCTCAGTGCAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219CAGA
GCCTTGCCAGCCCGCTCAGCAGAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219CTCT
GCCTTGCCAGCCCGCTCAGCTCTGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219CTCA
GCCTTGCCAGCCCGCTCAGCTCAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219CTGA
GCCTTGCCAGCCCGCTCAGCTGAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219ATCA
GCCTTGCCAGCCCGCTCAGATCAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219ATCT
GCCTTGCCAGCCCGCTCAGATCTGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219ATGA
GCCTTGCCAGCCCGCTCAGATGAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
PM1219AGCA
GCCTTGCCAGCCCGCTCAGAGCAGGTGGATCTCGGACCCGGAGACTGT
48
|
|
HLA-A Exon 3 Forward
PM1230TCAT
GCCTCCCTCGCGCCATCAGTCATGACTGGGCTGACCGTGGGGT
43
|
|
PM1230TGAT
GCCTCCCTCGCGCCATCAGTGATGACTGGGCTGACCGTGGGGT
43
|
|
PM1230TGCT
GCCTCCCTCGCGCCATCAGTGCTGACTGGGCTGACCGTGGGGT
43
|
|
PM1230TGCA
GCCTCCCTCGCGCCATCAGTGCAGACTGGGCTGACCGTGGGGT
43
|
|
PM1230CAGA
GCCTCCCTCGCGCCATCAGCAGAGACTGGGCTGACCGTGGGGT
43
|
|
PM1230CTCT
GCCTCCCTCGCGCCATCAGCTCTGACTGGGCTGACCGTGGGGT
43
|
|
PM1230CTCA
GCCTCCCTCGCGCCATCAGCTCAGACTGGGCTGACCGTGGGGT
43
|
|
PM1230CTGA
GCCTCCCTCGCGCCATCAGCTGAGACTGGGCTGACCGTGGGGT
43
|
|
PM1230ATCA
GCCTCCCTCGCGCCATCAGATCAGACTGGGCTGACCGTGGGGT
43
|
|
PM1230ATCT
GCCTCCCTCGCGCCATCAGATCTGACTGGGCTGACCGTGGGGT
43
|
|
PM1230ATGA
GCCTCCCTCGCGCCATCAGATGAGACTGGGCTGACCGTGGGGT
43
|
|
PM1230AGCA
GCCTCCCTCGCGCCATCAGAGCAGACTGGGCTGACCGTGGGGT
43
|
|
HLA-A Exon 3 Reverse
PS102TCAT
GCCTTGCCAGCCCGCTCAGTCATCCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102TGAT
GCCTTGCCAGCCCGCTCAGTGATCCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102TGCT
GCCTTGCCAGCCCGCTCAGTGCTCCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102TGCA
GCCTTGCCAGCCCGCTCAGTGCACCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102CAGA
GCCTTGCCAGCCCGCTCAGCAGACCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102CTCT
GCCTTGCCAGCCCGCTCAGCTCTCCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102CTCA
GCCTTGCCAGCCCGCTCAGCTCACCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102CTGA
GCCTTGCCAGCCCGCTCAGCTGACCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102ATCA
GCCTTGCCAGCCCGCTCAGATCACCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102ATCT
GCCTTGCCAGCCCGCTCAGATCTCCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102ATGA
GCCTTGCCAGCCCGCTCAGATGACCCCTGGTACCVGTGCGCTGCA
45
|
|
PS102AGCA
GCCTTGCCAGCCCGCTCAGAGCACCCCTGGTACCVGTGCGCTGCA
45
|
|
HLA-A Exon 4 Forward
PB1001TCTC
GCCTCCCTCGCGCCATCAGTCTCTGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001TCTG
GCCTCCCTCGCGCCATCAGTCTGTGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001TGAG
GCCTCCCTCGCGCCATCAGTGAGTGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001TGCA
GCCTCCCTCGCGCCATCAGTGCATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001CAGA
GCCTCCCTCGCGCCATCAGCAGATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001CATG
GCCTCCCTCGCGCCATCAGCATGTGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001CTCA
GCCTCCCTCGCGCCATCAGCTCATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001CTGA
GCCTCCCTCGCGCCATCAGCTGATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001ATCA
GCCTCCCTCGCGCCATCAGATCATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001CTGC
GCCTCCCTCGCGCCATCAGCTGCTGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001ATGA
GCCTCCCTCGCGCCATCAGATGATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
PB1001AGCA
GCCTCCCTCGCGCCATCAGAGCATGCCTGAATGWTCTGACTCTTCCCGTMAGA
53
|
|
HLA-A Exon 4 Reverse
PM1226TCAG
GCCTTGCCAGCCCGCTCAGTCAGTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226TCTC
GCCTTGCCAGCCCGCTCAGTCTCTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226TCTG
GCCTTGCCAGCCCGCTCAGTCTGTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226TGAG
GCCTTGCCAGCCCGCTCAGTGAGTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226TGCA
GCCTTGCCAGCCCGCTCAGTGCATGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CAGA
GCCTTGCCAGCCCGCTCAGCAGATGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CAGC
GCCTTGCCAGCCCGCTCAGCAGCTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CATC
GCCTTGCCAGCCCGCTCAGCATCTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CATG
GCCTTGCCAGCCCGCTCAGCATGTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CTCA
GCCTTGCCAGCCCGCTCAGCTCATGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CTGA
GCCTTGCCAGCCCGCTCAGCTGATGACCCTGCTAAAGGTCTCCAGAG
47
|
|
PM1226CTGC
GCCTTGCCAGCCCGCTCAGCTGCTGACCCTGCTAAAGGTCTCCAGAG
47
|
|
HLA-A Exon 4 Reverse
PM1228TCAG
GCCTTGCCAGCCCGCTCAGTCAGTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228TCTC
GCCTTGCCAGCCCGCTCAGTCTCTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228TCTG
GCCTTGCCAGCCCGCTCAGTCTGTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228TGAG
GCCTTGCCAGCCCGCTCAGTGAGTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228TGCA
GCCTTGCCAGCCCGCTCAGTGCATGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CAGA
GCCTTGCCAGCCCGCTCAGCAGATGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CAGC
GCCTTGCCAGCCCGCTCAGCAGCTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CATC
GCCTTGCCAGCCCGCTCAGCATCTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CATG
GCCTTGCCAGCCCGCTCAGCATGTGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CTCA
GCCTTGCCAGCCCGCTCAGCTCATGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CTGA
GCCTTGCCAGCCCGCTCAGCTGATGACCCTGCTAAAGGTCAGAG
44
|
|
PM1228CTGC
GCCTTGCCAGCCCGCTCAGCTGCTGACCCTGCTAAAGGTCAGAG
44
|
|
HLA-B Exon 2 Forward
FJCC148TCAG
GCCTCCCTCGCGCCATCAGTCAGAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148TCAT
GCCTCCCTCGCGCCATCAGTCATAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148TCTC
GCCTCCCTCGCGCCATCAGTCTCAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148TCTG
GCCTCCCTCGCGCCATCAGTCTGAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148TGAT
GCCTCCCTCGCGCCATCAGTGATAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148TGAG
GCCTCCCTCGCGCCATCAGTGAGAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148TGCT
GCCTCCCTCGCGCCATCAGTGCTAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148CAGC
GCCTCCCTCGCGCCATCAGCAGCAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148CATC
GCCTCCCTCGCGCCATCAGCATCAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148CATG
GCCTCCCTCGCGCCATCAGCATGAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148CTCT
GCCTCCCTCGCGCCATCAGCTCTAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
FJCC148CTGC
GCCTCCCTCGCGCCATCAGCTGCAGAGCTCGGGAGGAGCGAGGGGACCSCAG
52
|
|
HLA-B Exon 2 Reverse
RRAP423TCAG
GCCTTGCCAGCCCGCTCAGTCAGACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423TCAT
GCCTTGCCAGCCCGCTCAGTCATACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423TCTC
GCCTTGCCAGCCCGCTCAGTCTCACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423TCTG
GCCTTGCCAGCCCGCTCAGTCTGACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423TGAT
GCCTTGCCAGCCCGCTCAGTGATACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423TGAG
GCCTTGCCAGCCCGCTCAGTGAGACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423TGCT
GCCTTGCCAGCCCGCTCAGTGCTACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423CAGC
GCCTTGCCAGCCCGCTCAGCAGCACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423CATC
GCCTTGCCAGCCCGCTCAGCATCACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423CATG
GCCTTGCCAGCCCGCTCAGCATGACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423CTCT
GCCTTGCCAGCCCGCTCAGCTCTACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
RRAP423CTGC
GCCTTGCCAGCCCGCTCAGCTGCACTCGAGGCCTCGCTCTGGTTGTAGTA
50
|
|
HLA-B Exon 3 Forward
FJCC146TCAG
GCCTCCCTCGCGCCATCAGTCAGAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146TCAT
GCCTCCCTCGCGCCATCAGTCATAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146TCTC
GCCTCCCTCGCGCCATCAGTCTCAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146TCTG
GCCTCCCTCGCGCCATCAGTCTGAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146TGAT
GCCTCCCTCGCGCCATCAGTGATAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146TGAG
GCCTCCCTCGCGCCATCAGTGAGAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146TGCT
GCCTCCCTCGCGCCATCAGTGCTAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146CAGC
GCCTCCCTCGCGCCATCAGCAGCAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146CATC
GCCTCCCTCGCGCCATCAGCATCAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146CATG
GCCTCCCTCGCGCCATCAGCATGAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146CTCT
GCCTCCCTCGCGCCATCAGCTCTAGAGCTCGGGCCAGGGTCTCACA
46
|
|
FJCC146CTGC
GCCTCCCTCGCGCCATCAGCTGCAGAGCTCGGGCCAGGGTCTCACA
46
|
|
HLA-B Exon 3 Reverse
RJCC149TCAG
GCCTTGCCAGCCCGCTCAGTCAGACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149TCAT
GCCTTGCCAGCCCGCTCAGTCATACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149TCTC
GCCTTGCCAGCCCGCTCAGTCTCACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149TCTG
GCCTTGCCAGCCCGCTCAGTCTGACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149TGAT
GCCTTGCCAGCCCGCTCAGTGATACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149TGAG
GCCTTGCCAGCCCGCTCAGTGAGACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149TGCT
GCCTTGCCAGCCCGCTCAGTGCTACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149CAGC
GCCTTGCCAGCCCGCTCAGCAGCACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149CATC
GCCTTGCCAGCCCGCTCAGCATCACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149CATG
GCCTTGCCAGCCCGCTCAGCATGACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149CTCT
GCCTTGCCAGCCCGCTCAGCTCTACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
RJCC149CTGC
GCCTTGCCAGCCCGCTCAGCTGCACTCGAGGGAGGCCATCCCCGGCGACCTAT
53
|
|
HLA-B Exon 4 Forward
FJCC126TCAT
GCCTCCCTCGCGCCATCAGTCATGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126TCTC
GCCTCCCTCGCGCCATCAGTCTCGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126TGAT
GCCTCCCTCGCGCCATCAGTGATGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126TGCT
GCCTCCCTCGCGCCATCAGTGCTGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126TGCA
GCCTCCCTCGCGCCATCAGTGCAGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CAGA
GCCTCCCTCGCGCCATCAGCAGAGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CAGC
GCCTCCCTCGCGCCATCAGCAGCGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CATC
GCCTCCCTCGCGCCATCAGCATCGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CTCT
GCCTCCCTCGCGCCATCAGCTCTGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CTCA
GCCTCCCTCGCGCCATCAGCTCAGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CTGA
GCCTCCCTCGCGCCATCAGCTGAGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
FJCC126CTGC
GCCTCCCTCGCGCCATCAGCTGCGCGCCTGAATTTTCTGACTCTTCCCA
49
|
|
HLA-B Exon 4 Reverse
RCM117TCAT
GCCTTGCCAGCCCGCTCAGTCATGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117TCTC
GCCTTGCCAGCCCGCTCAGTCTCGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117TGAT
GCCTTGCCAGCCCGCTCAGTGATGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117TGCT
GCCTTGCCAGCCCGCTCAGTGCTGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117TGCA
GCCTTGCCAGCCCGCTCAGTGCAGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CAGA
GCCTTGCCAGCCCGCTCAGCAGAGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CAGC
GCCTTGCCAGCCCGCTCAGCAGCGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CATC
GCCTTGCCAGCCCGCTCAGCATCGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CTCT
GCCTTGCCAGCCCGCTCAGCTCTGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CTCA
GCCTTGCCAGCCCGCTCAGCTCAGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CTGA
GCCTTGCCAGCCCGCTCAGCTGAGGCTCCTGCTTTCCCTGAGAA
44
|
|
RCM117CTGC
GCCTTGCCAGCCCGCTCAGCTGCGGCTCCTGCTTTCCCTGAGAA
44
|
|
HLA-C Exon 1-2 Forward
FBA412TCAT
GCCTCCCTCGCGCCATCAGTCATATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412TCTC
GCCTCCCTCGCGCCATCAGTCTCATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412TGAT
GCCTCCCTCGCGCCATCAGTGATATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412TGCT
GCCTCCCTCGCGCCATCAGTGCTATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412CAGC
GCCTCCCTCGCGCCATCAGCAGCATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412CATC
GCCTCCCTCGCGCCATCAGCATCATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412CTCT
GCCTCCCTCGCGCCATCAGCTCTATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412CTGC
GCCTCCCTCGCGCCATCAGCTGCATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412ATCT
GCCTCCCTCGCGCCATCAGATCTATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412ATGC
GCCTCCCTCGCGCCATCAGATGCATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412AGCT
GCCTCCCTCGCGCCATCAGAGCTATGCGGGTCATGGCGCCCCRA
44
|
|
FBA412AGAT
GCCTCCCTCGCGCCATCAGAGATATGCGGGTCATGGCGCCCCRA
44
|
|
HLA-C Exon 1-2 Reverse
RBA414TCAT
GCCTTGCCAGCCCGCTCAGTCATGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414TCTC
GCCTTGCCAGCCCGCTCAGTCTCGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414TGAT
GCCTTGCCAGCCCGCTCAGTGATGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414TGCT
GCCTTGCCAGCCCGCTCAGTGCTGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414CAGC
GCCTTGCCAGCCCGCTCAGCAGCGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414CATC
GCCTTGCCAGCCCGCTCAGCATCGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414CTCT
GCCTTGCCAGCCCGCTCAGCTCTGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414CTGC
GCCTTGCCAGCCCGCTCAGCTGCGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414ATCT
GCCTTGCCAGCCCGCTCAGATCTGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414ATGC
GCCTTGCCAGCCCGCTCAGATGCGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414AGCT
GCCTTGCCAGCCCGCTCAGAGCTGAAAATGAAACCGGGTAAAGGYGA
47
|
|
RBA414AGAT
GCCTTGCCAGCCCGCTCAGAGATGAAAATGAAACCGGGTAAAGGYGA
47
|
|
HLA-C Exon 2 Reverse
EDB1313TCAG
GCCTTGCCAGCCCGCTCAGTCAGACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313TCAT
GCCTTGCCAGCCCGCTCAGTCATACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313TCTC
GCCTTGCCAGCCCGCTCAGTCTCACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313TCTG
GCCTTGCCAGCCCGCTCAGTCTGACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313TGAT
GCCTTGCCAGCCCGCTCAGTGATACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313TGAG
GCCTTGCCAGCCCGCTCAGTGAGACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313TGCT
GCCTTGCCAGCCCGCTCAGTGCTACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313CAGC
GCCTTGCCAGCCCGCTCAGCAGCACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313CATC
GCCTTGCCAGCCCGCTCAGCATCACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313CATG
GCCTTGCCAGCCCGCTCAGCATGACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313CTCT
GCCTTGCCAGCCCGCTCAGCTCTACTCGAGGGGCYGGGGTCACTCAC
47
|
|
RDB1313CTGC
GCCTTGCCAGCCCGCTCAGCTGCACTCGAGGGGCYGGGGTCACTCAC
47
|
|
HLA-C Exon 3 Forward
FDB1180TCAG
GCCTCCCTCGCGCCATCAGTCAGACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180TCAT
GCCTCCCTCGCGCCATCAGTCATACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180TCTC
GCCTCCCTCGCGCCATCAGTCTCACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180TCTG
GCCTCCCTCGCGCCATCAGTCTGACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180TGAT
GCCTCCCTCGCGCCATCAGTGATACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180TGAG
GCCTCCCTCGCGCCATCAGTGAGACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180TGCT
GCCTCCCTCGCGCCATCAGTGCTACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180CAGC
GCCTCCCTCGCGCCATCAGCAGCACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180CATC
GCCTCCCTCGCGCCATCAGCATCACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180CATG
GCCTCCCTCGCGCCATCAGCATGACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180CTCT
GCCTCCCTCGCGCCATCAGCTCTACGTCGACGGGCCAGGKTCTCACA
47
|
|
FDB1180CTGC
GCCTCCCTCGCGCCATCAGCTGCACGTCGACGGGCCAGGKTCTCACA
47
|
|
HLA-C Exon 3 Reverse
RDB1053TCAG
GCCTTGCCAGCCCGCTCAGTCAGACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053TCAT
GCCTTGCCAGCCCGCTCAGTCATACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053TCTG
GCCTTGCCAGCCCGCTCAGTCTGACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053TGAT
GCCTTGCCAGCCCGCTCAGTGATACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053TGAG
GCCTTGCCAGCCCGCTCAGTGAGACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053TGCT
GCCTTGCCAGCCCGCTCAGTGCTACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053CATG
GCCTTGCCAGCCCGCTCAGCATGACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053CTCT
GCCTTGCCAGCCCGCTCAGCTCTACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053ATCT
GCCTTGCCAGCCCGCTCAGATCTACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053ATGC
GCCTTGCCAGCCCGCTCAGATGCACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053AGCT
GCCTTGCCAGCCCGCTCAGAGCTACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
RDB1053AGAT
GCCTTGCCAGCCCGCTCAGAGATACCTCGAGGTCAGCAGCCTGACCACA
49
|
|
HLA-C Exon 4 Forward
FBNH277TCAG
GCCTCCCTCGCGCCATCAGTCAGCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277TCAT
GCCTCCCTCGCGCCATCAGTCATCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277TCTG
GCCTCCCTCGCGCCATCAGTCTGCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277TGAT
GCCTCCCTCGCGCCATCAGTGATCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277TGAG
GCCTCCCTCGCGCCATCAGTGAGCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277TGCT
GCCTCCCTCGCGCCATCAGTGCTCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277CATG
GCCTCCCTCGCGCCATCAGCATGCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277CTCT
GCCTCCCTCGCGCCATCAGCTCTCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277ATCT
GCCTCCCTCGCGCCATCAGATCTCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277AGCA
GCCTCCCTCGCGCCATCAGAGCACAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277AGCT
GCCTCCCTCGCGCCATCAGAGCTCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
FBNH277AGAT
GCCTCCCTCGCGCCATCAGAGATCAAAGTGTCTGAATTTTCTGACTCTTCCC
52
|
|
HLA-C Exon 4 Reverse
RBNH287TCAG
GCCTTGCCAGCCCGCTCAGTCAGTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287TCTC
GCCTTGCCAGCCCGCTCAGTCTCTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287TCTG
GCCTTGCCAGCCCGCTCAGTCTGTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287TGAG
GCCTTGCCAGCCCGCTCAGTGAGTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287TGCA
GCCTTGCCAGCCCGCTCAGTGCATGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CAGA
GCCTTGCCAGCCCGCTCAGCAGATGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CAGC
GCCTTGCCAGCCCGCTCAGCAGCTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CATC
GCCTTGCCAGCCCGCTCAGCATCTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CATG
GCCTTGCCAGCCCGCTCAGCATGTGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CTCA
GCCTTGCCAGCCCGCTCAGCTCATGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CTGA
GCCTTGCCAGCCCGCTCAGCTGATGAAGGGCTCCAGAAGGACTT
44
|
|
RBNH287CTGC
GCCTTGCCAGCCCGCTCAGCTGCTGAAGGGCTCCAGAAGGACTT
44
|
|
HLA-C Exon 4 Reverse
RBNH288TCAG
GCCTTGCCAGCCCGCTCAGTCAGTGAAGGGCTCCAGGACTT
41
|
|
RBNH288TCTC
GCCTTGCCAGCCCGCTCAGTCTCTGAAGGGCTCCAGGACTT
41
|
|
RBNH288TCTG
GCCTTGCCAGCCCGCTCAGTCTGTGAAGGGCTCCAGGACTT
41
|
|
RBNH288TGAG
GCCTTGCCAGCCCGCTCAGTGAGTGAAGGGCTCCAGGACTT
41
|
|
RBNH288TGCA
GCCTTGCCAGCCCGCTCAGTGCATGAAGGGCTCCAGGACTT
41
|
|
RBNH288CAGA
GCCTTGCCAGCCCGCTCAGCAGATGAAGGGCTCCAGGACTT
41
|
|
RBNH288CAGC
GCCTTGCCAGCCCGCTCAGCAGCTGAAGGGCTCCAGGACTT
41
|
|
RBNH288CATC
GCCTTGCCAGCCCGCTCAGCATCTGAAGGGCTCCAGGACTT
41
|
|
RBNH288CATG
GCCTTGCCAGCCCGCTCAGCATGTGAAGGGCTCCAGGACTT
41
|
|
RBNH288CTCA
GCCTTGCCAGCCCGCTCAGCTCATGAAGGGCTCCAGGACTT
41
|
|
RBNH288CTGA
GCCTTGCCAGCCCGCTCAGCTGATGAAGGGCTCCAGGACTT
41
|
|
RBNH288CTGC
GCCTTGCCAGCCCGCTCAGCTGCTGAAGGGCTCCAGGACTT
41
|
|
DQB1 Exon 2 Forward
FBA400TCAG
GCCTCCCTCGCGCCATCAGTCAGAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400TCTC
GCCTCCCTCGCGCCATCAGTCTCAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400TCTG
GCCTCCCTCGCGCCATCAGTCTGAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400TGAG
GCCTCCCTCGCGCCATCAGTGAGAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400CAGC
GCCTCCCTCGCGCCATCAGCAGCAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400CATC
GCCTCCCTCGCGCCATCAGCATCAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400CATG
GCCTCCCTCGCGCCATCAGCATGAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400CTGC
GCCTCCCTCGCGCCATCAGCTGCAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400ATGC
GCCTCCCTCGCGCCATCAGATGCAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400AGAC
GCCTCCCTCGCGCCATCAGAGACAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400CTCT
GCCTCCCTCGCGCCATCAGCTCTAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
FBA400ATCT
GCCTCCCTCGCGCCATCAGATCTAGGATCCCCGCAGAGGATTTCGTGTACCA
52
|
|
DQB1 Exon 2 Reverse
RDB380TCAG
GCCTTGCCAGCCCGCTCAGTCAGTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380TCTC
GCCTTGCCAGCCCGCTCAGTCTCTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380TCTG
GCCTTGCCAGCCCGCTCAGTCTGTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380TGAG
GCCTTGCCAGCCCGCTCAGTGAGTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380CAGC
GCCTTGCCAGCCCGCTCAGCAGCTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380CATC
GCCTTGCCAGCCCGCTCAGCATCTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380CATG
GCCTTGCCAGCCCGCTCAGCATGTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380CTGC
GCCTTGCCAGCCCGCTCAGCTGCTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380ATGC
GCCTTGCCAGCCCGCTCAGATGCTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380AGAC
GCCTTGCCAGCCCGCTCAGAGACTCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380CTCA
GCCTTGCCAGCCCGCTCAGCTCATCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
RDB380ATGA
GCCTTGCCAGCCCGCTCAGATGATCCTGCAGGACGCTCACCTCTCCGCTGCA
52
|
|
DQB1 Exon 3 Forward
FBNH86TCTC
GCCTCCCTCGCGCCATCAGTCTCTGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86TGCA
GCCTCCCTCGCGCCATCAGTGCATGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86CAGA
GCCTCCCTCGCGCCATCAGCAGATGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86CAGC
GCCTCCCTCGCGCCATCAGCAGCTGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86CATC
GCCTCCCTCGCGCCATCAGCATCTGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86CTCA
GCCTCCCTCGCGCCATCAGCTCATGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86CTGA
GCCTCCCTCGCGCCATCAGCTGATGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86CTGC
GCCTCCCTCGCGCCATCAGCTGCTGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86ATCA
GCCTCCCTCGCGCCATCAGATCATGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86ATGA
GCCTCCCTCGCGCCATCAGATGATGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86ATGC
GCCTCCCTCGCGCCATCAGATGCTGGAGCCCACAGTGACCATCTCC
46
|
|
FBNH86AGCA
GCCTCCCTCGCGCCATCAGAGCATGGAGCCCACAGTGACCATCTCC
46
|
|
DQB1 Exon 3 Reverse
RBA411TCTC
GCCTTGCCAGCCCGCTCAGTCTCGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411TGCA
GCCTTGCCAGCCCGCTCAGTGCAGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411CAGA
GCCTTGCCAGCCCGCTCAGCAGAGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411CAGC
GCCTTGCCAGCCCGCTCAGCAGCGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411CATC
GCCTTGCCAGCCCGCTCAGCATCGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411CTCA
GCCTTGCCAGCCCGCTCAGCTCAGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411CTGA
GCCTTGCCAGCCCGCTCAGCTGAGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411CTGC
GCCTTGCCAGCCCGCTCAGCTGCGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411ATCA
GCCTTGCCAGCCCGCTCAGATCAGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411ATGA
GCCTTGCCAGCCCGCTCAGATGAGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411ATGC
GCCTTGCCAGCCCGCTCAGATGCGCTGGGGTGCTCCACGTGGCA
44
|
|
RBA411AGCA
GCCTTGCCAGCCCGCTCAGAGCAGCTGGGGTGCTCCACGTGGCA
44
|
|
DPA1 Exon 2 Forward
FPM058BTCAT
GCCTCCCTCGCGCCATCAGTCATCGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BTGAT
GCCTCCCTCGCGCCATCAGTGATCGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BTGCT
GCCTCCCTCGCGCCATCAGTGCTCGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BTGCA
GCCTCCCTCGCGCCATCAGTGCACGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BCAGA
GCCTCCCTCGCGCCATCAGCAGACGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BCTCT
GCCTCCCTCGCGCCATCAGCTCTCGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BCTCA
GCCTCCCTCGCGCCATCAGCTCACGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BCTGA
GCCTCCCTCGCGCCATCAGCTGACGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BATCA
GCCTCCCTCGCGCCATCAGATCACGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BATCT
GCCTCCCTCGCGCCATCAGATCTCGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BATGA
GCCTCCCTCGCGCCATCAGATGACGGATCCATGTGTCAACTTATGCC
47
|
|
FPM058BAGCA
GCCTCCCTCGCGCCATCAGAGCACGGATCCATGTGTCAACTTATGCC
47
|
|
DPA1 Exon 2 Reverse
RPM059BTCAT
GCCTTGCCAGCCCGCTCAGTCATGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BTGAT
GCCTTGCCAGCCCGCTCAGTGATGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BTGCT
GCCTTGCCAGCCCGCTCAGTGCTGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BTGCA
GCCTTGCCAGCCCGCTCAGTGCAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BCAGA
GCCTTGCCAGCCCGCTCAGCAGAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BCTCT
GCCTTGCCAGCCCGCTCAGCTCTGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BCTCA
GCCTTGCCAGCCCGCTCAGCTCAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BCTGA
GCCTTGCCAGCCCGCTCAGCTGAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BATCA
GCCTTGCCAGCCCGCTCAGATCAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BATCT
GCCTTGCCAGCCCGCTCAGATCTGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BATGA
GCCTTGCCAGCCCGCTCAGATGAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
RPM059BAGCA
GCCTTGCCAGCCCGCTCAGAGCAGGCTACAGAGGAAGAGGCAAAGATAGG
50
|
|
DPB1 Exon 2 Forward
FUG19BTGAT
GCCTCCCTCGCGCCATCAGTGATGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BTGCT
GCCTCCCTCGCGCCATCAGTGCTGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BTGCA
GCCTCCCTCGCGCCATCAGTGCAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BCAGA
GCCTCCCTCGCGCCATCAGCAGAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BCTCT
GCCTCCCTCGCGCCATCAGCTCTGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BCTCA
GCCTCCCTCGCGCCATCAGCTCAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BCTGA
GCCTCCCTCGCGCCATCAGCTGAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BATCA
GCCTCCCTCGCGCCATCAGATCAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BATCT
GCCTCCCTCGCGCCATCAGATCTGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BATGA
GCCTCCCTCGCGCCATCAGATGAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BAGCA
GCCTCCCTCGCGCCATCAGAGCAGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
FUG19BAGCT
GCCTCCCTCGCGCCATCAGAGCTGCTGCAGGAGAGTGGCGCCTCCGCTCAT
51
|
|
DPB1 Exon 2 Reverse
RUG1BTGAT
GCCTTGCCAGCCCGCTCAGTGATCGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BTGCT
GCCTTGCCAGCCCGCTCAGTGCTCGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BTGCA
GCCTTGCCAGCCCGCTCAGTGCACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BCAGA
GCCTTGCCAGCCCGCTCAGCAGACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BCTCT
GCCTTGCCAGCCCGCTCAGCTCTCGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BCTCA
GCCTTGCCAGCCCGCTCAGCTCACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BCTGA
GCCTTGCCAGCCCGCTCAGCTGACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BATCA
GCCTTGCCAGCCCGCTCAGATCACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BATCT
GCCTTGCCAGCCCGCTCAGATGACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BATGA
GCCTTGCCAGCCCGCTCAGATGACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BAGCA
GCCTTGCCAGCCCGCTCAGAGCACGGATCCGGCCCAAAGCCCTCACTC
48
|
|
RUG1BAGCT
GCCTTGCCAGCCCGCTCAGAGCTCGGATCCGGCCCAAAGCCCTCACTC
48
|
|
DQA1 Exon 2 Forward
FPM066BTGAG
GCCTCCCTCGCGCCATCAGTCATGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BTGCT
GCCTCCCTCGCGCCATCAGTCTCGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BTGCA
GCCTCCCTCGCGCCATCAGTGATGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BCAGA
GCCTCCCTCGCGCCATCAGTGCTGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BCATG
GCCTCCCTCGCGCCATCAGCAGCGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BCTCT
GCCTCCCTCGCGCCATCAGCATCGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BCTCA
GCCTCCCTCGCGCCATCAGCTCTGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BCTGA
GCCTCCCTCGCGCCATCAGCTCAGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BATCA
GCCTCCCTCGCGCCATCAGCTGCGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BATCT
GCCTCCCTCGCGCCATCAGATCAGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BATGA
GCCTCCCTCGCGCCATCAGATCTGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
FPM066BAGCA
GCCTCCCTCGCGCCATCAGATGCGTTTCTTCCATCATTTTGTGTATTAAGGT
52
|
|
DQA1 Exon 2 Reverse
RRR060BTGAG2
GCCTTGCCAGCCCGCTCAGTGAGCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BTGCT2
GCCTTGCCAGCCCGCTCAGTGCTCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BTGCA2
GCCTTGCCAGCCCGCTCAGTGCACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCAGA2
GCCTTGCCAGCCCGCTCAGCAGACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCATG2
GCCTTGCCAGCCCGCTCAGCATGCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCTCT2
GCCTTGCCAGCCCGCTCAGCTCTCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCTCA2
GCCTTGCCAGCCCGCTCAGCTCACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCTGA2
GCCTTGCCAGCCCGCTCAGCTGACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BATCA2
GCCTTGCCAGCCCGCTCAGATCACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BATCT2
GCCTTGCCAGCCCGCTCAGATCTCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BATGA2
GCCTTGCCAGCCCGCTCAGATGACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BAGCA2
GCCTTGCCAGCCCGCTCAGAGCACGGTAGAGTTGTAGCGTTTA
43
|
|
HLA-A Exon 2 Forward
F5AIN1.46TCAT
GCCTCCCTCGCGCCATCAGTCATGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
pairs with HLA-A Exon 1-2
|
Reverse (PM1219)
|
|
F5AIN1.46TGAT
GCCTCCCTCGCGCCATCAGTGATGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46TGCT
GCCTCCCTCGCGCCATCAGTGCTGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46TGCA
GCCTCCCTCGCGCCATCAGTGCAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46CAGA
GCCTCCCTCGCGCCATCAGCAGAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46CTCT
GCCTCCCTCGCGCCATCAGCTCTGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46CTCA
GCCTCCCTCGCGCCATCAGCTCAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46CTGA
GCCTCCCTCGCGCCATCAGCTGAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46ATCA
GCCTCCCTCGCGCCATCAGATCAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46ATCT
GCCTCCCTCGCGCCATCAGATCTGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46ATGA
GCCTCCCTCGCGCCATCAGATGAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
F5AIN1.46AGCA
GCCTCCCTCGCGCCATCAGAGCAGAAACGGCCTCTGTGGGGAGAAGCAA
49
|
|
HLA-C Exon 2 Forward
FDB1215TCAT
GCCTCCCTCGCGCCATCAGTCATAGTCGACGAADCGGCCTCTGSGGA
47
|
pairs with HLA-C Exon 2
|
Reverse (DB1313)
|
|
FDB1215TCTC
GCCTCCCTCGCGCCATCAGTCTCAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215TGAT
GCCTCCCTCGCGCCATCAGTGATAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215TGCT
GCCTCCCTCGCGCCATCAGTGCTAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215CAGC
GCCTCCCTCGCGCCATCAGCAGCAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215CATC
GCCTCCCTCGCGCCATCAGCATCAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215CTCT
GCCTCCCTCGCGCCATCAGCTCTAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215CTGC
GCCTCCCTCGCGCCATCAGCTGCAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215ATCT
GCCTCCCTCGCGCCATCAGATCTAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215ATGC
GCCTCCCTCGCGCCATCAGATGCAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215AGCT
GCCTCCCTCGCGCCATCAGAGCTAGTCGACGAADCGGCCTCTGSGGA
47
|
|
FDB1215AGAT
GCCTCCCTCGCGCCATCAGAGATAGTCGACGAADCGGCCTCTGSGGA
47
|
|
DRB1 Exon 2 Forward
FCRX28TCAG
GCCTCCCTCGCGCCATCAGTCAGCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28TCTG
GCCTCCCTCGCGCCATCAGTCTGCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28TGAT
GCCTCCCTCGCGCCATCAGTGATCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28TGCT
GCCTCCCTCGCGCCATCAGTGCTCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28CAGA
GCCTCCCTCGCGCCATCAGCAGACCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28CATG
GCCTCCCTCGCGCCATCAGCATGCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28CTCT
GCCTCCCTCGCGCCATCAGCTCTCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28CTGA
GCCTCCCTCGCGCCATCAGCTGACCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28ATCA
GCCTCCCTCGCGCCATCAGATCACCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28ATGA
GCCTCCCTCGCGCCATCAGATGACCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28AGCA
GCCTCCCTCGCGCCATCAGAGCACCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
FCRX28AGAT
GCCTCCCTCGCGCCATCAGAGATCCGGATCCTTCGTGTCCCCACAGCACG
50
|
|
DRB1 Exon 2 Reverse
RAB60TCAG
GCCTTGCCAGCCCGCTCAGTCAGCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60TCTG
GCCTTGCCAGCCCGCTCAGTCTGCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60TGAT
GCCTTGCCAGCCCGCTCAGTGATCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60TGCT
GCCTTGCCAGCCCGCTCAGTGCTCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60CAGA
GCCTTGCCAGCCCGCTCAGCAGACCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60CATG
GCCTTGCCAGCCCGCTCAGCATGCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60CTCT
GCCTTGCCAGCCCGCTCAGCTCTCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60CTGA
GCCTTGCCAGCCCGCTCAGCTGACCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60ATCA
GCCTTGCCAGCCCGCTCAGATCACCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60ATGA
GCCTTGCCAGCCCGCTCAGATGACCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60AGCA
GCCTTGCCAGCCCGCTCAGAGCACCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
RAB60AGAT
GCCTTGCCAGCCCGCTCAGAGATCCGAATTCCGCTGCACTGTGAAGCTCTC
51
|
|
DPA1 Exon 2 Forward
FDPA1E2_TCAG
GCCTCCCTCGCGCCATCAGTCAGATGTTTGAATTTGATGAAGATGAG
47
|
pairs with DPA1 Exon 2
|
reverse (PM059B)
|
|
FDPA1E2_TCAT
GCCTCCCTCGCGCCATCAGTCATATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_TCTC
GCCTCCCTCGCGCCATCAGTCTCATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_TCTG
GCCTCCCTCGCGCCATCAGTCTGATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_TGAT
GCCTCCCTCGCGCCATCAGTGATATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_TGAG
GCCTCCCTCGCGCCATCAGTGAGATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_TGCT
GCCTCCCTCGCGCCATCAGTGCTATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_CAGC
GCCTCCCTCGCGCCATCAGCAGCATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_CATC
GCCTCCCTCGCGCCATCAGCATCATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_CATG
GCCTCCCTCGCGCCATCAGCATGATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_CTCT
GCCTCCCTCGCGCCATCAGCTCTATGTTTGAATTTGATGAAGATGAG
47
|
|
FDPA1E2_CTGC
GCCTCCCTCGCGCCATCAGCTGCATGTTTGAATTTGATGAAGATGAG
47
|
|
DQA1 Exon 2 Forward
FPM1240BTCAT
GCCTCCCTCGCGCCATCAGTCATGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BTGAT
GCCTCCCTCGCGCCATCAGTGATGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BTGCT
GCCTCCCTCGCGCCATCAGTGCTGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BCTCT
GCCTCCCTCGCGCCATCAGCTCTGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BCTCA
GCCTCCCTCGCGCCATCAGCTCAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BATCA
GCCTCCCTCGCGCCATCAGATCAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BATCT
GCCTCCCTCGCGCCATCAGATCTGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BTGCA
GCCTCCCTCGCGCCATCAGTGCAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BCAGA
GCCTCCCTCGCGCCATCAGCAGAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BCTGA
GCCTCCCTCGCGCCATCAGCTGAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BATGA
GCCTCCCTCGCGCCATCAGATGAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
FPM1240BAGCA
GCCTCCCTCGCGCCATCAGAGCAGTTTCTTYCATCATTTTGTGTATTAAGGT
52
|
|
DQA1 Exon 2 Reverse
RRR060BTCAT
GCCTTGCCAGCCCGCTCAGTCATCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BTGAT
GCCTTGCCAGCCCGCTCAGTGATCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BTGCT
GCCTTGCCAGCCCGCTCAGTGCTCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCTCT
GCCTTGCCAGCCCGCTCAGCTCTCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCTCA
GCCTTGCCAGCCCGCTCAGCTCACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BATCA
GCCTTGCCAGCCCGCTCAGATCACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BATCT
GCCTTGCCAGCCCGCTCAGATCTCGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BTGCA
GCCTTGCCAGCCCGCTCAGTGCACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCAGA
GCCTTGCCAGCCCGCTCAGCAGACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BCTGA
GCCTTGCCAGCCCGCTCAGCTGACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BATGA
GCCTTGCCAGCCCGCTCAGATGACGGTAGAGTTGTAGCGTTTA
43
|
|
RRR060BAGCA
GCCTTGCCAGCCCGCTCAGAGCACGGTAGAGTTGTAGCGTTTA
43
|
|
HLA-C Exon 3
RHLACE3TGAT
GCCTTGCCAGCCCGCTCAGTGATCTCCCCACTGCCCCTGGTAC
43
|
reverse primer for re-amp
|
from DB1180, DB1053
|
amplicon
|
|
RHLACE3TGCT
GCCTTGCCAGCCCGCTCAGTGCTCTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3TGCA
GCCTTGCCAGCCCGCTCAGTGCACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3CAGA
GCCTTGCCAGCCCGCTCAGCAGACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3CTCT
GCCTTGCCAGCCCGCTCAGCTCTCTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3CTCA
GCCTTGCCAGCCCGCTCAGCTCACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3CTGA
GCCTTGCCAGCCCGCTCAGCTGACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3ATCA
GCCTTGCCAGCCCGCTCAGATCACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3ATCT
GCCTTGCCAGCCCGCTCAGATCTCTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3ATGA
GCCTTGCCAGCCCGCTCAGATGACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3AGCA
GCCTTGCCAGCCCGCTCAGAGCACTCCCCACTGCCCCTGGTAC
43
|
|
RHLACE3AGCT
GCCTTGCCAGCCCGCTCAGAGCTCTCCCCACTGCCCCTGGTAC
43
|
|
HLA-A Exon 2, 3, 4
FPM1231TCAGA
GCCTTGCCAGCCCGCTCAGTCAGAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
2nd gen primers 5 base
|
MID's
|
|
FPM1231TCATC
GCCTTGCCAGCCCGCTCAGTCATCGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231TCTCA
GCCTTGCCAGCCCGCTCAGTCTCAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231TCTGA
GCCTTGCCAGCCCGCTCAGTCTGAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231TGATC
GCCTTGCCAGCCCGCTCAGTGATCGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231TGAGA
GCCTTGCCAGCCCGCTCAGTGAGAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231TGCTC
GCCTTGCCAGCCCGCTCAGTGCTCGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231TGCAT
GCCTTGCCAGCCCGCTCAGTGCATGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231CAGAT
GCCTTGCCAGCCCGCTCAGCAGATGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231CAGCA
GCCTTGCCAGCCCGCTCAGCAGCAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231CATCA
GCCTTGCCAGCCCGCTCAGCATCAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1231CATGA
GCCTTGCCAGCCCGCTCAGCATGAGGGAAACGGCCTCTGTGGGGAGAAGCA
51
|
|
FPM1229TCAGA
GCCTCCCTCGCGCCATCAGTCAGAGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TCATC
GCCTCCCTCGCGCCATCAGTCATCGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TCTCA
GCCTCCCTCGCGCCATCAGTCTCAGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TCTGA
GCCTCCCTCGCGCCATCAGTCTGAGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TGATC
GCCTCCCTCGCGCCATCAGTGATCGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TGAGA
GCCTCCCTCGCGCCATCAGTGAGAGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TGCTC
GCCTCCCTCGCGCCATCAGTGCTCGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229TGCAT
GCCTCCCTCGCGCCATCAGTGCATGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229CAGAT
GCCTCCCTCGCGCCATCAGCAGATGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229CAGCA
GCCTCCCTCGCGCCATCAGCAGCAGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229CATCA
GCCTCCCTCGCGCCATCAGCATCAGACTGGGCTGACCKYGGGGT
44
|
|
FPM1229CATGA
GCCTCCCTCGCGCCATCAGCATGAGACTGGGCTGACCKYGGGGT
44
|
|
RPB1003TCAGA
GCCTTGCCAGCCCGCTCAGTCAGAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TCATC
GCCTTGCCAGCCCGCTCAGTCATCGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TCTCA
GCCTTGCCAGCCCGCTCAGTCTCAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TCTGA
GCCTTGCCAGCCCGCTCAGTCTGAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TGATC
GCCTTGCCAGCCCGCTCAGTGATCGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TGAGA
GCCTTGCCAGCCCGCTCAGTGAGAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TGCTC
GCCTTGCCAGCCCGCTCAGTGCTCGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003TGCAT
GCCTTGCCAGCCCGCTCAGTGCATGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003CAGAT
GCCTTGCCAGCCCGCTCAGCAGATGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003CAGCA
GCCTTGCCAGCCCGCTCAGCAGCAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003CATCA
GCCTTGCCAGCCCGCTCAGCATCAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
RPB1003CATGA
GCCTTGCCAGCCCGCTCAGCATGAGAGGGTGATATTCTAGTGTTGGTCCCAA
52
|
|
FPM1252TCAGA
GCCTCCCTCGCGCCATCAGTCAGACTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TCATG
GCCTCCCTCGCGCCATCAGTCATGCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TCTCT
GCCTCCCTCGCGCCATCAGTCTCTCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TCTGA
GCCTCCCTCGCGCCATCAGTCTGACTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TGATG
GCCTCCCTCGCGCCATCAGTGATGCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TGAGA
GCCTCCCTCGCGCCATCAGTGAGACTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TGCTG
GCCTCCCTCGCGCCATCAGTGCTGCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252TGCAT
GCCTCCCTCGCGCCATCAGTGCATCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252CAGAT
GCCTCCCTCGCGCCATCAGCAGATCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252CAGCT
GCCTCCCTCGCGCCATCAGCAGCTCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252CATCT
GCCTCCCTCGCGCCATCAGCATCTCTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
FPM1252CATGA
GCCTCCCTCGCGCCATCAGCATGACTGGGTTCTGTGCTCYCTTCCCCAT
49
|
|
RPM1248TCAGA
GCCTTGCCAGCCCGCTCAGTCAGACTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TCATG
GCCTTGCCAGCCCGCTCAGTCATGCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TCTCT
GCCTTGCCAGCCCGCTCAGTCTCTCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TCTGA
GCCTTGCCAGCCCGCTCAGTCTGACTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TGATG
GCCTTGCCAGCCCGCTCAGTGATGCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TGAGA
GCCTTGCCAGCCCGCTCAGTGAGACTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TGCTG
GCCTTGCCAGCCCGCTCAGTGCTGCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248TGCAT
GCCTTGCCAGCCCGCTCAGTGCATCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248CAGAT
GCCTTGCCAGCCCGCTCAGCAGATCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248CAGCT
GCCTTGCCAGCCCGCTCAGCAGCTCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248CATCT
GCCTTGCCAGCCCGCTCAGCATCTCTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
RPM1248CATGA
GCCTTGCCAGCCCGCTCAGCATGACTCCAGAGAGGCTCCTGCTTTCCSTA
50
|
|
HLA-B Exon 4
FCM141TCAGA
GCCTCCCTCGCGCCATCAGTCAGACTGGTCACATGGGTGGTCC
43
|
2nd gen primer 5 base
|
MID's
|
|
FCM141TCATG
GCCTCCCTCGCGCCATCAGTCATGCTGGTCACATGGGTGGTCC
43
|
|
FCM141TCTCT
GCCTCCCTCGCGCCATCAGTCTCTCTGGTCACATGGGTGGTCC
43
|
|
FCM141TCTGA
GCCTCCCTCGCGCCATCAGTCTGACTGGTCACATGGGTGGTCC
43
|
|
FCM141TGATG
GCCTCCCTCGCGCCATCAGTGATGCTGGTCACATGGGTGGTCC
43
|
|
FCM141TGAGA
GCCTCCCTCGCGCCATCAGTGAGACTGGTCACATGGGTGGTCC
43
|
|
FCM141TGCTG
GCCTCCCTCGCGCCATCAGTGCTGCTGGTCACATGGGTGGTCC
43
|
|
FCM141TGCAT
GCCTCCCTCGCGCCATCAGTGCATCTGGTCACATGGGTGGTCC
43
|
|
FCM141CAGAT
GCCTCCCTCGCGCCATCAGCAGATCTGGTCACATGGGTGGTCC
43
|
|
FCM141CAGCT
GCCTCCCTCGCGCCATCAGCAGCTCTGGTCACATGGGTGGTCC
43
|
|
FCM141CATCT
GCCTCCCTCGCGCCATCAGCATCTCTGGTCACATGGGTGGTCC
43
|
|
FCM141CATGA
GCCTCCCTCGCGCCATCAGCATGACTGGTCACATGGGTGGTCC
43
|
|
FCM141TCAGC
GCCTTGCCAGCCCGCTCAGTCAGCAGATATGACCCCTCATCCC
43
|
|
FCM141TCATG
GCCTTGCCAGCCCGCTCAGTCATGAGATATGACCCCTCATCCC
43
|
|
FCM141TCTCT
GCCTTGCCAGCCCGCTCAGTCTCTAGATATGACCCCTCATCCC
43
|
|
FCM141TCTGC
GCCTTGCCAGCCCGCTCAGTCTGCAGATATGACCCCTCATCCC
43
|
|
FCM141TGATG
GCCTTGCCAGCCCGCTCAGTGATGAGATATGACCCCTCATCCC
43
|
|
FCM141TGAGC
GCCTTGCCAGCCCGCTCAGTGAGCAGATATGACCCCTCATCCC
43
|
|
FCM141TGCTG
GCCTTGCCAGCCCGCTCAGTGCTGAGATATGACCCCTCATCCC
43
|
|
FCM141TGCAG
GCCTTGCCAGCCCGCTCAGTGCAGAGATATGACCCCTCATCCC
43
|
|
FCM141CAGAG
GCCTTGCCAGCCCGCTCAGCAGAGAGATATGACCCCTCATCCC
43
|
|
FCM141CAGCT
GCCTTGCCAGCCCGCTCAGCAGCTAGATATGACCCCTCATCCC
43
|
|
FCM141CATCT
GCCTTGCCAGCCCGCTCAGCATCTAGATATGACCCCTCATCCC
43
|
|
FCM141CATGC
GCCTTGCCAGCCCGCTCAGCATGCAGATATGACCCCTCATCCC
43
|
|
HLA-C Exon 4
FBA765TCAGA
GCCTCCCTCGCGCCATCAGTCAGAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
2nd gen primer 5 base
|
MID's
|
|
FBA765TCATC
GCCTCCCTCGCGCCATCAGTCATCGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765TCTCA
GCCTCCCTCGCGCCATCAGTCTCAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765TCTGA
GCCTCCCTCGCGCCATCAGTCTGAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765TGATC
GCCTCCCTCGCGCCATCAGTGATCGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765TGAGA
GCCTCCCTCGCGCCATCAGTGAGAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765TGCTC
GCCTCCCTCGCGCCATCAGTGCTCGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765TGCAT
GCCTCCCTCGCGCCATCAGTGCATGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765CAGAT
GCCTCCCTCGCGCCATCAGCAGATGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765CAGCA
GCCTCCCTCGCGCCATCAGCAGCAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765CATCA
GCCTCCCTCGCGCCATCAGCATCAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
FBA765CATGA
GCCTCCCTCGCGCCATCAGCATGAGTGTCGCAAGAGAGATGCAAAGTGT
49
|
|
RBA767TCAGA
GCCTTGCCAGCCCGCTCAGTCAGAGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TCATC
GCCTTGCCAGCCCGCTCAGTCATCGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TCTCA
GCCTTGCCAGCCCGCTCAGTCTCAGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TCTGA
GCCTTGCCAGCCCGCTCAGTCTGAGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TGATC
GCCTTGCCAGCCCGCTCAGTGATCGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TGAGA
GCCTTGCCAGCCCGCTCAGTGAGAGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TGCTC
GCCTTGCCAGCCCGCTCAGTGCTCGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767TGCAT
GCCTTGCCAGCCCGCTCAGTGCATGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767CAGAT
GCCTTGCCAGCCCGCTCAGCAGATGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767CAGCA
GCCTTGCCAGCCCGCTCAGCAGCAGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767CATCA
GCCTTGCCAGCCCGCTCAGCATCAGAGGGGAAGGTGAGGGGCC
43
|
|
RBA767CATGA
GCCTTGCCAGCCCGCTCAGCATGAGAGGGGAAGGTGAGGGGCC
43
|
|
DQB1 Exon 3
RBA762TCAGC
GCCTTGCCAGCCCGCTCAGTCAGCAGTGACATCAGGGATAAGAGATGGGAA
51
|
2nd gen primer 5 base
|
MID's
|
|
RBA762TCATG
GCCTTGCCAGCCCGCTCAGTCATGAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762TCTCT
GCCTTGCCAGCCCGCTCAGTCTCTAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762TCTGC
GCCTTGCCAGCCCGCTCAGTCTGCAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762TGATG
GCCTTGCCAGCCCGCTCAGTGATGAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762TGAGC
GCCTTGCCAGCCCGCTCAGTGAGCAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762TGCTG
GCCTTGCCAGCCCGCTCAGTGCTGAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762TGCAG
GCCTTGCCAGCCCGCTCAGTGCAGAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762CAGAG
GCCTTGCCAGCCCGCTCAGCAGAGAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762CAGCT
GCCTTGCCAGCCCGCTCAGCAGCTAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762CATCT
GCCTTGCCAGCCCGCTCAGCATCTAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
RBA762CATGC
GCCTTGCCAGCCCGCTCAGCATGCAGTGACATCAGGGATAAGAGATGGGAA
51
|
|
TABLE 2
|
|
CellLineName
LocusName
Allele1
Allele2
|
|
JW5
DRB1
0103
03
|
JW5
DRB3
0101
0101
|
JW5
DQA1
0101 (1.1)
0501 (4.1)
|
JW5
DQB1
0201/2
0501
|
JW5
DPA1
010301
020102
|
JW5
DPB1
010101
020102
|
JW5
HLA-A
0101
2301
|
JW5
HLA-B
0801/4
18
|
JW5
HLA-C
05
07
|
RAJI
DRB1
0301
100101
|
RAJI
DRB3
02
02
|
RAJI
DQA1
0101 (1.1)
0501 (4.1)
|
RAJI
DQB1
0201/2
0501
|
RAJI
DPA1
020202
020202
|
RAJI
DPB1
010101
010101
|
RAJI
HLA-A
03
03
|
RAJI
HLA-B
1510
1510
|
RAJI
HLA-C
030402
04
|
NAMALWA
DRB1
0405
1503
|
NAMALWA
DRB4
01
0
|
NAMALWA
DRB5
0101
0
|
NAMALWA
DQA1
0102 (1.2)
0301 (3)
|
NAMALWA
DQB1
0302
0602
|
NAMALWA
DPA1
010301
020202
|
NAMALWA
DPB1
0201
0201
|
NAMALWA
HLA-A
03
6802
|
NAMALWA
HLA-B
0702
4901
|
NAMALWA
HLA-C
0701/6
0702/3
|
APA
DRB1
1405
150101/102
|
APA
DRB3
02/0302
0
|
APA
DRB5
01
0
|
APA
DQA1
0101 (1.1)
0102 (1.2)
|
APA
DQB1
050301
0501
|
APA
DPA1
020202
020202
|
APA
DPB1
0501
0501
|
APA
HLA-A
2403
1101
|
APA
HLA-B
1502
5502
|
APA
HLA-C
08
1203/6
|
MG
DRB1
0401/16
1001
|
MG
DRB4
01
01
|
MG
DQA1
0101 (1.1)
0301 (3)
|
MG
DQB1
0302/7
0501
|
MG
DPA1
010301
010301
|
MG
DPB1
0401
0601
|
MG
HLA-A
0101
0201
|
MG
HLA-B
15
3701
|
MG
HLA-C
03
0602
|
TTL
DRB1
1301
1501
|
TTL
DQA1
0102 (1.2)
0103 (1.3)
|
TTL
DQB1
0502
0603
|
TTL
DPA1
010301
020101
|
TTL
DPB1
0201
1301
|
TTL
HLA-A
1102
3303
|
TTL
HLA-B
51
5401
|
TTL
HLA-C
0102
0302
|
FH6
DRB1
160101
1001
|
FH6
DRB5
01/02
02
|
FH6
DQA1
0101 (1.1)
0102 (1.2)
|
FH6
DQB1
0501
0502
|
FH6
DPA1
010301
010301
|
FH6
DPB1
020102
0401
|
FH6
HLA-A
24
2901
|
FH6
HLA-B
0705/6
2702
|
JY
DRB1
0404
1301
|
JY
DRB3
0101
0
|
JY
DRB4
01
0
|
JY
DQA1
0103 (1.3)
0301 (3)
|
JY
DQB1
0302
0603
|
JY
DPA1
010301
010301
|
JY
DPB1
020102
0401
|
JY
HLA-A
020101
020101
|
JY
HLA-B
070201
070201
|
JY
HLA-C
0702
0702
|
Z3232
DRB1
010201
1001
|
Z3232
DQA1
0101 (1.1)
0101 (1.1)
|
Z3232
DQB1
0501
0501
|
Z3232
DPA1
010301
020101
|
Z3232
DPB1
020102
1301
|
Z3232
HLA-A
2902
3002
|
Z3232
HLA-B
5702
7801/2
|
Z3232
HLA-C
1601
1801/2
|
LH
DRB1
0301
0404
|
LH
DRB3
01
0
|
LH
DRB4
01
0
|
LH
DQA1
0301 (3)
0501 (4.1)
|
LH
DQB1
0201
0402
|
LH
DPA1
020101
020102
|
LH
DPB1
010101
0501
|
LH
HLA-A
2402
2402
|
LH
HLA-B
0802
2708
|
LH
HLA-C
0102
0701/6
|
VOO
DRB1
0101
030101
|
VOO
DRB3
01
01
|
VOO
DQA1
0101 (1.1)
0501 (4.1)
|
VOO
DQB1
0201/2
0501
|
VOO
DPA1
010301
010301
|
VOO
DPB1
020102
0401
|
VOO
HLA-A
0101
0301
|
VOO
HLA-B
0801
5601
|
VOO
HLA-C
0102
0701/06/16
|
AMALA
DRB1
1402
1402
|
AMALA
DRB3
0101
0101
|
AMALA
DQA1
0501 (4.1)
0501 (4.1)
|
AMALA
DQB1
0301
0301
|
AMALA
DPA1
010301
010301
|
AMALA
DPB1
0402
0402
|
AMALA
HLA-A
021701
021701
|
AMALA
HLA-B
1501
1501
|
AMALA
HLA-C
0303
0303
|
E4181324
DRB1
150201
150201
|
E4181324
DRB5
0102
0102
|
E4181324
DQA1
0103 (1.3)
0103 (1.3)
|
E4181324
DQB1
060101
060101
|
E4181324
DPA1
010301
010301
|
E4181324
DPB1
020102
0401
|
E4181324
HLA-A
0101
0101
|
E4181324
HLA-B
520101
520101
|
E4181324
HLA-C
1202
1202
|
SAVC
DRB1
0401
0401
|
SAVC
DRB4
0101
0101
|
SAVC
DQA1
0301 (3)
0301 (3)
|
SAVC
DQB1
0302
0302
|
SAVC
DPA1
020101
020101
|
SAVC
DPB1
1001
1001
|
SAVC
HLA-A
0301
0301
|
SAVC
HLA-B
0702
0702
|
SAVC
HLA-C
0702
0702
|
LADA
DRB1
090102
1201/6
|
LADA
DRB3
02
0
|
LADA
DRB4
01/02
0
|
LADA
DQA1
0101 (1.1)
0301 (3)
|
LADA
DQB1
0201/2
0501
|
LADA
DPA1
010301
020101
|
LADA
DPB1
0301
1701
|
LADA
HLA-A
0201
8001
|
LADA
HLA-B
0702
5703
|
LADA
HLA-C
0702/3
0802
|
DBUG
DRB1
0701
1105
|
DBUG
DRB3
0202
0
|
DBUG
DRB4
0101
0
|
DBUG
DQA1
0101 (1.1)
0201 (2)
|
DBUG
DQB1
030302
0602
|
DBUG
DPA1
010301
020202
|
DBUG
DPB1
040101
0501
|
DBUG
HLA-A
1101
2601
|
DBUG
HLA-B
0705/6
55
|
AMAI/AMAL
DRB1
1503
1503
|
AMAI/AMAL
DRB5
0101
0101
|
AMAI/AMAL
DQA1
0102 (1.2)
0102 (1.2)
|
AMAI/AMAL
DQB1
0602
0602
|
AMAI/AMAL
DPA1
0301
0301
|
AMAI/AMAL
DPB1
0402
0402
|
AMAI/AMAL
HLA-A
6802
6802
|
AMAI/AMAL
HLA-B
5301
5301
|
AMAI/AMAL
HLA-C
0401
0401
|
CRK
DRB1
0701
0701
|
CRK
DQA1
0201 (2)
0201 (2)
|
CRK
DQB1
0201
0201
|
CRK
DPA1
020101
020202
|
CRK
DPB1
010101
110101
|
CRK
HLA-A
2902/4
2902/4
|
CRK
HLA-B
4403
4403
|
CRK
HLA-C
1601
1601
|
H0301
DRB1
1302
1302
|
H0301
DQA1
0102 (1.2)
0102 (1.2)
|
H0301
DQB1
0609
0609
|
H0301
DPA1
020101
020101
|
H0301
DPB1
0501
0501
|
H0301
HLA-A
0301
0301
|
H0301
HLA-B
1402
1402
|
H0301
HLA-C
0802
0802
|
OOS
DRB1
0101
0101
|
OOS
DQA1
0101 (1.1)
0101 (1.1)
|
OOS
DQB1
0501
0501
|
OOS
DPA1
010301
010301
|
OOS
DPB1
020102
020102
|
OOS
HLA-A
2601
2601/11N
|
OOS
HLA-B
5601
5601
|
OOS
HLA-C
0102
0102
|
SSTO
DRB1
0403
0403
|
SSTO
DQA1
0301 (3)
0301 (3)
|
SSTO
DQB1
0305
0305
|
SSTO
DPA1
010301
010301
|
SSTO
DPB1
0401
0401
|
SSTO
HLA-A
3201
3201
|
SSTO
HLA-B
4402
4402
|
SSTO
HLA-C
0501
0501
|
BIN40/BIN-40
DRB1
0404
0404
|
BIN40/BIN-40
DRB4
01/02
01/02
|
BIN40/BIN-40
DQA1
0301 (3)
0301 (3)
|
BIN40/BIN-40
DQB1
0302
0302
|
BIN40/BIN-40
DPA1
010301
010301
|
BIN40/BIN-40
DPB1
0301
0601
|
BIN40/BIN-40
HLA-A
02
310102
|
BIN40/BIN-40
HLA-B
1401
4001
|
BIN40/BIN-40
HLA-C
03
0802
|
APD
DRB1
1301
1301
|
APD
DRB3
02
02
|
APD
DQA1
0103 (1.3)
0103 (1.3)
|
APD
DQB1
0603
0603
|
APD
DPA1
010301
010301
|
APD
DPB1
0402
0402
|
APD
HLA-A
0101
0101
|
APD
HLA-B
4001
4001
|
APD
HLA-C
0602
0602
|
HAR
DRB1
0301
0301
|
HAR
DRB3
0101
0101
|
HAR
DQA1
0501 (4.1)
0501 (4.1)
|
HAR
DQB1
0201
0201
|
HAR
DPA1
010301
010301
|
HAR
DPB1
040101
0401
|
HAR
HLA-A
0101
0101
|
HAR
HLA-B
0801
0801/5
|
HAR
HLA-C
0701/6
0701/5
|
|
Patent Application
20100086914
