Genetic typing primer group for HLA (human leukocyte antigen) ultrahigh resolution level, kit and genetic typing method

Abstract

The invention relates to the technical field of biomedical detection, in particular to a genetic typing primer group, a kit and a genetic typing method for accurately realizing an ultrahigh resolution level of 11 sites of human leukocyte antigen (HLA) based on a nanopore sequencing technology. According to the genetic typing primer group for the HLA ultrahigh resolution level, the primer group is used for HLA gene multiple amplification, and the primer group comprises an HLA-A site primer pair, an HLA-B site primer pair, an HLA-C site primer pair, an HLA-DRB1 site primer pair, an HLA-DRB3 / 4 / 5 site primer pair, an HLA-DQA1 site primer pair, an HLA-DQB1 site primer pair, an HLA-DPA1 site primer pair and an HLA-DPB1 site primer pair. According to the primer group for multiple amplification of 11 genes for HLA typing, disclosed by the invention, the amplification of each gene can be simultaneously carried out on the loci of HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 of the HLA genes in a reaction system, so that the integrity, the amplification efficiency and the amplification uniformity of the gene amplification as well as the accuracy of subsequent HLA gene typing are improved.

Description

Technical Field

[0001] The present invention relates to the field of biomedical detection technology, and in particular to a genotyping primer set, a kit and a genotyping method for accurately achieving ultra-high resolution level of 11 human leukocyte antigen (HLA) sites based on nanopore sequencing technology. Background Art

[0002] The human leukocyte antigen (HLA) system is one of the most complex and highly polymorphic regions in the human genome. It plays a decisive role in antigen recognition, antigen presentation, immune response and regulation, and destruction of foreign antigen target cells. It is also the main material basis for causing immune rejection reactions. HLA class I and class II antigens on the surface of transplant cells are highly polymorphic and highly immunogenic cell surface markers with significant antigen presentation and immune activation potential. Therefore, whether the HLA matching between the donor and the recipient is the key to the success of allogeneic organ, tissue or cell transplantation. Many studies have shown that with higher resolution of the HLA typing system, the transplant survival rate is higher and the recurrence rate is lower.

[0003] Traditional HLA typing methods, serological methods, are difficult to accurately distinguish highly similar antigen subtypes. Molecular detection methods, such as PCR-sequence-specific oligonucleotide probe (PCR-SSOP), PCR-sequence-specific primer (PCR-SSP) typing, PCR-sequencing typing (PCR-SBT) and other methods have many disadvantages, such as limited resolution, lack of ability to detect new alleles, cumbersome operation, and susceptibility to contamination interference, which greatly restricts the demand for accurate HLA typing in clinical and scientific research application scenarios.

[0004] With the development of sequencing technology, the next generation sequencing technology (Next Generation Sequencing Technology, NGS) has been successfully used for HLA typing. Compared with PCR-based and first-generation sequencing technologies, the HLA typing method based on second-generation sequencing has significantly improved the resolution level of HLA typing. However, due to the short read length of NGS, ambiguous results will still appear for complex regions of the HLA gene.

[0005] In recent years, nanopore sequencing technology has emerged. It uses nanoscale protein pores to allow nucleic acid molecules to pass through one by one under the drive of an electric field, and monitors the changes in the current signal generated by the bases in real time to complete the sequencing work. This technology has an ultra-long sequencing read length, which is particularly suitable for the analysis of complex genomic regions, bringing new hope for overcoming the difficulty of HLA typing. Multiple studies have demonstrated that the HLA typing method based on nanopore sequencing can perform full-length sequencing of multiple HLA loci, greatly improving the resolution of some HLA loci. However, the current research only performs full-length amplification from 5'UTR to 3'UTR on HLA-A, HLA-B, HLA-C, HLA-DQA1, HLA-DQB1, HLA-DPB1 and HLA-DPA1 loci. For HLA-DRB1 and -DRB3 / 4 / 5 loci, due to the large similarity of some intervals and the different number and relative position of each monomer gene, it is difficult to design primers. At present, primers are basically designed in segments or amplified in multiple tubes. HLA is the most complex human genetic polymorphism system known to date. The HLA genetic region not only contains many genetic loci, but also a large number of alleles at each locus. China has a large population base, many ethnic groups, and a wide geographical distribution, with a large number of variations. However, in the existing websites and software related to HLA analysis, the overall proportion of Chinese population data is relatively low. Therefore, the traditional primer design method has the problem of poor inclusiveness, resulting in amplification failure, preference and missed detection in some special population groups. At the same time, there are already a number of long-PCR enzymes on the market, but these enzymes have low overall amplification efficiency, and one round of amplification takes about 8 hours. These combined factors have brought many inconveniences to the experiment, increased the risk of contamination, and increased the time and price costs of the experiment. Summary of the invention

[0006] The purpose of the present invention is to overcome the deficiencies of the prior art and provide a primer set and a genotyping method for HLA ultra-high resolution genotyping. The primer set for multiple amplification of 11 genes for HLA typing of the present invention can realize the simultaneous amplification of each gene of HLA gene HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 sites in one reaction system, thereby improving the integrity, efficiency and uniformity of gene amplification and the accuracy of subsequent HLA genotyping.

[0007] In order to solve the above technical problems, the present invention particularly adopts the following technical solutions:

[0008] In the first aspect of the present invention, a primer set for HLA ultra-high resolution genotyping is provided, and the primer set is used for HLA gene multiple amplification; the primer set includes primer pairs for HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3 / 4 / 5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 sites. In order to improve the inclusiveness of primers, scheme 1 adopts Ω primer design and nested primer design. This scheme covers the full-length exon sequence of 11 sites. The upstream and downstream primers of the primer pair and the related nucleotide sequences are selected from any one or more of the following groups 1 to 48:

[0009] The upstream and downstream primer sequences of the primer pairs for each site in Group 1 are as follows:

[0010] 1) The HLA-AF sequence is shown in SEQ ID NO.1, the HLA-A-R1 sequence is shown in SEQ ID NO.2, and the HLA-A-R2 sequence is shown in SEQ ID NO.3;

[0011] 2) The HLA-BF sequence is shown in SEQ ID NO.4, the HLA-B-R1 sequence is shown in SEQ ID NO.5, and the HLA-B-R2 sequence is shown in SEQ ID NO.6;

[0012] 3) The HLA-CF sequence is shown in SEQ ID NO.7, the HLA-C-R1 sequence is shown in SEQ ID NO.8, and the HLA-C-R2 sequence is shown in SEQ ID NO.9;

[0013] 4) The HLA-DPA-F1 sequence is shown in SEQ ID NO.10, the HLA-DPA-F2 sequence is shown in SEQ ID NO.11, and the HLA-DPA-R sequence is shown in SEQ ID NO.12;

[0014] 5) The HLA-DPB-F sequence is shown in SEQ ID NO.13, the HLA-DPB-R1 sequence is shown in SEQ ID NO.14, and the HLA-DPB-R2 sequence is shown in SEQ ID NO.15;

[0015] 6) The HLA-DQA-F sequence is shown in SEQ ID NO.16, the HLA-DQA-R1 sequence is shown in SEQ ID NO.17, and the HLA-DQA-R2 sequence is shown in SEQ ID NO.18;

[0016] 7) The HLA-DQB-F sequence is shown in SEQ ID NO.19, the HLA-DQB-R1 sequence is shown in SEQ ID NO.20, and the HLA-DQB-R2 sequence is shown in SEQ ID NO.21;

[0017] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO.22, the HLA-DRB1345-R1 sequence is shown in SEQ ID NO.23, and the HLA-DRB1345-R2 sequence is shown in SEQ ID NO.24;

[0018] The upstream and downstream primer sequences of the primer pairs for each site in Group 2 are as follows:

[0019] 1) The HLA-AF sequence is shown in SEQ ID NO. 25, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0020] 2) The HLA-BF sequence is shown in SEQ ID NO. 26, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0021] 3) The HLA-CF sequence is shown in SEQ ID NO. 27, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0022] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 28, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0023] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 29, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0024] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 30, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0025] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 31, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0026] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.32, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0027] The upstream and downstream primer sequences of the primer pairs for each site in Group 3 are as follows:

[0028] 1) The HLA-AF sequence is shown in SEQ ID NO. 33, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0029] 2) The HLA-BF sequence is shown in SEQ ID NO. 34, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0030] 3) The HLA-CF sequence is shown in SEQ ID NO. 35, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0031] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 36, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0032] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 37, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0033] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 38, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0034] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 39, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0035] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 40, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0036] The upstream and downstream primer sequences of the primer pairs for each site in Group 4 are as follows:

[0037] 1) The HLA-AF sequence is shown in SEQ ID NO. 41, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0038] 2) The HLA-BF sequence is shown in SEQ ID NO. 42, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0039] 3) The HLA-CF sequence is shown in SEQ ID NO. 43, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0040] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 44, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0041] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 45, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0042] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 46, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0043] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 47, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0044] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 48, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0045] The upstream and downstream primer sequences of the primer pairs for each site in Group 5 are as follows:

[0046] 1) The HLA-AF sequence is shown in SEQ ID NO. 49, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0047] 2) The HLA-BF sequence is shown in SEQ ID NO. 50, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0048] 3) The HLA-CF sequence is shown in SEQ ID NO.51, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0049] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 52, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0050] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 53, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0051] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 54, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0052] 7) The HLA-DQB-F sequence is shown in SEQ ID NO.55, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0053] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.56, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0054] The upstream and downstream primer sequences of the primer pairs for each site in Group 6 are as follows:

[0055] 1) The HLA-AF sequence is shown in SEQ ID NO.57, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0056] 2) The HLA-BF sequence is shown in SEQ ID NO. 58, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0057] 3) The HLA-CF sequence is shown in SEQ ID NO. 59, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0058] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 60, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0059] 5) The HLA-DPB-F sequence is shown in SEQ ID NO.61, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0060] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 62, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0061] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 63, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0062] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.64, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0063] The upstream and downstream primer sequences of the primer pairs for each site in Group 7 are as follows:

[0064] 1) The HLA-AF sequence is shown in SEQ ID NO.65, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0065] 2) The HLA-BF sequence is shown in SEQ ID NO. 66, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0066] 3) The HLA-CF sequence is shown in SEQ ID NO. 67, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0067] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 68, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0068] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 69, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0069] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 70, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0070] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 71, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0071] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.72, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0072] The upstream and downstream primer sequences of the primer pairs for each site in Group 8 are as follows:

[0073] 1) The HLA-AF sequence is shown in SEQ ID NO. 73, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0074] 2) The HLA-BF sequence is shown in SEQ ID NO. 74, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0075] 3) The HLA-CF sequence is shown in SEQ ID NO. 75, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0076] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 76, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0077] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 77, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0078] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 78, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0079] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 79, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0080] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 80, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0081] The upstream and downstream primer sequences of the primer pairs for each site in Group 9 are as follows:

[0082] 1) The HLA-AF sequence is shown in SEQ ID NO.81, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0083] 2) The HLA-BF sequence is shown in SEQ ID NO.82, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0084] 3) The HLA-CF sequence is shown in SEQ ID NO. 83, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0085] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 84, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0086] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 85, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0087] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 86, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0088] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 87, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0089] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.88, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0090] The upstream and downstream primer sequences of the primer pairs for each site in Group 10 are as follows:

[0091] 1) The HLA-AF sequence is shown in SEQ ID NO.89, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0092] 2) The HLA-BF sequence is shown in SEQ ID NO.90, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0093] 3) The HLA-CF sequence is shown in SEQ ID NO.91, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0094] 4) The HLA-DPA-R sequence is shown in SEQ ID NO.92, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0095] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 93, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0096] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 94, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0097] 7) The HLA-DQB-F sequence is shown in SEQ ID NO.95, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0098] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 96, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0099] The upstream and downstream primer sequences of the primer pairs for each site in Group 11 are as follows:

[0100] 1) The HLA-AF sequence is shown in SEQ ID NO.97, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0101] 2) The HLA-BF sequence is shown in SEQ ID NO.98, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0102] 3) The HLA-CF sequence is shown in SEQ ID NO. 99, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0103] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 100, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0104] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 101, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0105] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 102, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0106] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 103, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0107] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 104, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0108] The upstream and downstream primer sequences of the primer pairs for each site in Group 12 are as follows:

[0109] 1) The HLA-AF sequence is shown in SEQ ID NO. 105, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0110] 2) The HLA-BF sequence is shown in SEQ ID NO. 106, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0111] 3) The HLA-CF sequence is shown in SEQ ID NO. 107, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0112] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 108, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0113] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 109, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0114] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 110, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0115] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 111, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0116] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.112, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0117] The upstream and downstream primer sequences of the primer pairs for each site in Group 13 are as follows:

[0118] 1) The HLA-AF sequence is shown in SEQ ID NO. 113, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0119] 2) The HLA-BF sequence is shown in SEQ ID NO. 114, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0120] 3) The HLA-CF sequence is shown in SEQ ID NO. 115, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0121] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 116, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0122] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 117, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0123] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 118, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0124] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 119, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0125] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 120, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0126] The upstream and downstream primer sequences of the primer pairs for each site in Group 14 are as follows:

[0127] 1) The HLA-AF sequence is shown in SEQ ID NO. 121, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0128] 2) The HLA-BF sequence is shown in SEQ ID NO. 122, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0129] 3) The HLA-CF sequence is shown in SEQ ID NO. 123, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0130] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 124, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0131] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 125, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0132] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 126, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0133] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 127, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0134] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 128, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0135] The upstream and downstream primer sequences of the primer pairs for each site in Group 15 are as follows:

[0136] 1) The HLA-AF sequence is shown in SEQ ID NO. 129, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0137] 2) The HLA-BF sequence is shown in SEQ ID NO. 130, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0138] 3) The HLA-CF sequence is shown in SEQ ID NO. 131, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0139] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 132, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0140] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 133, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0141] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 134, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0142] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 135, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0143] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 136, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0144] The upstream and downstream primer sequences of the primer pairs for each site in Group 16 are as follows:

[0145] 1) The HLA-AF sequence is shown in SEQ ID NO. 137, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0146] 2) The HLA-BF sequence is shown in SEQ ID NO. 138, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0147] 3) The HLA-CF sequence is shown in SEQ ID NO. 139, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0148] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 140, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0149] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 141, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0150] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 142, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0151] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 143, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0152] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 144, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0153] The upstream and downstream primer sequences of the primer pairs for each site in Group 17 are as follows:

[0154] 1) The HLA-AF sequence is shown in SEQ ID NO. 145, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0155] 2) The HLA-BF sequence is shown in SEQ ID NO. 146, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0156] 3) The HLA-CF sequence is shown in SEQ ID NO. 147, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0157] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 148, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0158] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 149, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0159] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 150, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0160] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 151, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0161] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.152, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0162] The upstream and downstream primer sequences of the primer pairs for each site in Group 18 are as follows:

[0163] 1) The HLA-AF sequence is shown in SEQ ID NO. 153, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0164] 2) The HLA-BF sequence is shown in SEQ ID NO. 154, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0165] 3) The HLA-CF sequence is shown in SEQ ID NO. 155, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0166] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 156, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0167] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 157, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0168] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 158, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0169] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 159, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0170] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 160, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0171] The upstream and downstream primer sequences of the primer pairs for each site in Group 19 are as follows:

[0172] 1) The HLA-AF sequence is shown in SEQ ID NO. 161, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0173] 2) The HLA-BF sequence is shown in SEQ ID NO. 162, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0174] 3) The HLA-CF sequence is shown in SEQ ID NO. 163, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0175] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 164, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0176] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 165, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0177] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 166, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0178] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 167, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0179] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 168, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0180] The upstream and downstream primer sequences of the primer pairs for each site in Group 20 are as follows:

[0181] 1) The HLA-AF sequence is shown in SEQ ID NO. 169, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0182] 2) The HLA-BF sequence is shown in SEQ ID NO. 170, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0183] 3) The HLA-CF sequence is shown in SEQ ID NO. 171, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0184] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 172, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0185] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 173, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0186] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 174, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0187] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 175, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0188] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 176, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0189] The upstream and downstream primer sequences of the primer pairs for each site in Group 21 are as follows:

[0190] 1) The HLA-AF sequence is shown in SEQ ID NO. 177, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0191] 2) The HLA-BF sequence is shown in SEQ ID NO. 178, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0192] 3) The HLA-CF sequence is shown in SEQ ID NO. 179, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0193] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 180, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0194] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 181, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0195] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 182, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0196] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 183, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0197] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 184, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0198] The upstream and downstream primer sequences of the primer pairs for each site in Group 22 are as follows:

[0199] 1) The HLA-AF sequence is shown in SEQ ID NO. 185, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0200] 2) The HLA-BF sequence is shown in SEQ ID NO. 186, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0201] 3) The HLA-CF sequence is shown in SEQ ID NO. 187, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0202] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 188, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0203] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 189, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0204] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 190, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0205] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 191, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0206] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 192, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0207] The upstream and downstream primer sequences of the primer pairs for each site in Group 23 are as follows:

[0208] 1) The HLA-AF sequence is shown in SEQ ID NO. 193, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0209] 2) The HLA-BF sequence is shown in SEQ ID NO. 194, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0210] 3) The HLA-CF sequence is shown in SEQ ID NO. 195, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0211] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 196, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0212] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 197, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0213] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 198, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0214] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 199, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0215] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 200, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0216] The upstream and downstream primer sequences of the primer pairs for each site in Group 24 are as follows:

[0217] 1) The HLA-AF sequence is shown in SEQ ID NO. 201, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0218] 2) The HLA-BF sequence is shown in SEQ ID NO. 202, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0219] 3) The HLA-CF sequence is shown in SEQ ID NO. 203, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0220] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 204, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0221] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 205, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0222] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 206, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0223] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 207, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0224] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 208, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0225] The upstream and downstream primer sequences of the primer pairs for each site in Group 25 are as follows:

[0226] 1) The HLA-AF sequence is shown in SEQ ID NO. 209, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0227] 2) The HLA-BF sequence is shown in SEQ ID NO. 210, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0228] 3) The HLA-CF sequence is shown in SEQ ID NO. 211, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0229] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 212, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0230] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 213, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0231] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 214, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0232] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 215, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0233] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 216, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0234] The upstream and downstream primer sequences of the primer pairs for each site in Group 26 are as follows:

[0235] 1) The HLA-AF sequence is shown in SEQ ID NO. 217, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0236] 2) The HLA-BF sequence is shown in SEQ ID NO. 218, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0237] 3) The HLA-CF sequence is shown in SEQ ID NO. 219, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0238] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 220, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0239] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 221, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0240] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 222, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0241] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 223, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0242] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 224, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0243] The upstream and downstream primer sequences of the primer pairs for each site in Group 27 are as follows:

[0244] 1) The HLA-AF sequence is shown in SEQ ID NO. 225, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0245] 2) The HLA-BF sequence is shown in SEQ ID NO. 226, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0246] 3) The HLA-CF sequence is shown in SEQ ID NO. 227, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0247] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 228, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0248] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 229, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0249] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 230, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0250] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 231, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0251] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 232, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0252] The upstream and downstream primer sequences of the primer pairs for each site in Group 28 are as follows:

[0253] 1) The HLA-AF sequence is shown in SEQ ID NO. 233, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0254] 2) The HLA-BF sequence is shown in SEQ ID NO. 234, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0255] 3) The HLA-CF sequence is shown in SEQ ID NO. 235, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0256] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 236, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0257] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 237, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0258] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 238, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0259] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 239, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0260] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 240, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0261] The upstream and downstream primer sequences of the primer pairs for each site in Group 29 are as follows:

[0262] 1) The HLA-AF sequence is shown in SEQ ID NO. 241, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0263] 2) The HLA-BF sequence is shown in SEQ ID NO. 242, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0264] 3) The HLA-CF sequence is shown in SEQ ID NO. 243, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0265] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 244, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0266] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 245, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0267] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 246, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0268] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 247, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0269] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 248, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0270] The upstream and downstream primer sequences of the primer pairs for each site in Group 30 are as follows:

[0271] 1) The HLA-AF sequence is shown in SEQ ID NO. 249, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0272] 2) The HLA-BF sequence is shown in SEQ ID NO. 250, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0273] 3) The HLA-CF sequence is shown in SEQ ID NO. 251, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0274] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 252, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0275] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 253, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0276] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 254, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0277] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 255, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0278] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 256, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0279] The upstream and downstream primer sequences of the primer pairs for each site in Group 31 are as follows:

[0280] 1) The HLA-AF sequence is shown in SEQ ID NO. 257, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0281] 2) The HLA-BF sequence is shown in SEQ ID NO. 258, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0282] 3) The HLA-CF sequence is shown in SEQ ID NO. 259, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0283] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 260, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0284] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 261, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0285] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 262, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0286] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 263, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0287] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 264, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0288] The upstream and downstream primer sequences of the primer pairs for each site in Group 32 are as follows:

[0289] 1) The HLA-AF sequence is shown in SEQ ID NO. 265, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0290] 2) The HLA-BF sequence is shown in SEQ ID NO. 266, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0291] 3) The HLA-CF sequence is shown in SEQ ID NO. 267, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0292] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 268, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0293] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 269, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0294] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 270, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0295] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 271, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0296] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 272, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0297] The upstream and downstream primer sequences of the primer pairs for each site in Group 33 are as follows:

[0298] 1) The HLA-AF sequence is shown in SEQ ID NO. 273, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0299] 2) The HLA-BF sequence is shown in SEQ ID NO. 274, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0300] 3) The HLA-CF sequence is shown in SEQ ID NO. 275, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0301] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 276, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0302] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 277, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0303] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 278, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0304] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 279, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0305] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 280, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0306] The upstream and downstream primer sequences of the primer pairs for each site in Group 34 are as follows:

[0307] 1) The HLA-AF sequence is shown in SEQ ID NO. 281, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0308] 2) The HLA-BF sequence is shown in SEQ ID NO. 282, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0309] 3) The HLA-CF sequence is shown in SEQ ID NO. 283, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0310] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 284, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0311] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 285, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0312] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 286, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0313] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 287, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0314] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 288, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0315] The upstream and downstream primer sequences of the primer pairs for each site in Group 35 are as follows:

[0316] 1) The HLA-AF sequence is shown in SEQ ID NO. 289, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0317] 2) The HLA-BF sequence is shown in SEQ ID NO. 290, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0318] 3) The HLA-CF sequence is shown in SEQ ID NO. 291, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0319] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 292, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0320] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 293, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0321] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 294, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0322] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 295, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0323] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 296, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0324] The upstream and downstream primer sequences of the primer pairs for each site in Group 36 are as follows:

[0325] 1) The HLA-AF sequence is shown in SEQ ID NO. 297, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0326] 2) The HLA-BF sequence is shown in SEQ ID NO. 298, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0327] 3) The HLA-CF sequence is shown in SEQ ID NO. 299, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0328] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 300, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0329] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 301, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0330] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 302, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0331] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 303, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0332] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 304, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0333] The upstream and downstream primer sequences of the primer pairs for each site in Group 37 are as follows:

[0334] 1) The HLA-AF sequence is shown in SEQ ID NO. 305, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0335] 2) The HLA-BF sequence is shown in SEQ ID NO. 306, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0336] 3) The HLA-CF sequence is shown in SEQ ID NO. 307, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0337] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 308, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0338] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 309, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0339] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 310, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0340] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 311, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0341] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.312, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0342] The upstream and downstream primer sequences of the primer pairs for each site in Group 38 are as follows:

[0343] 1) The HLA-AF sequence is shown in SEQ ID NO. 313, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0344] 2) The HLA-BF sequence is shown in SEQ ID NO. 314, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0345] 3) The HLA-CF sequence is shown in SEQ ID NO. 315, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0346] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 316, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0347] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 317, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0348] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 318, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0349] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 319, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0350] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 320, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0351] The upstream and downstream primer sequences of the primer pairs for each site in Group 39 are as follows:

[0352] 1) The HLA-AF sequence is shown in SEQ ID NO. 321, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0353] 2) The HLA-BF sequence is shown in SEQ ID NO. 322, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0354] 3) The HLA-CF sequence is shown in SEQ ID NO. 323, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0355] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 324, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0356] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 325, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0357] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 326, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0358] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 327, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0359] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 328, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0360] The upstream and downstream primer sequences of the primer pairs for each site in Group 40 are as follows:

[0361] 1) The HLA-AF sequence is shown in SEQ ID NO. 329, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0362] 2) The HLA-BF sequence is shown in SEQ ID NO. 330, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0363] 3) The HLA-CF sequence is shown in SEQ ID NO. 331, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0364] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 332, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0365] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 333, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0366] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 334, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0367] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 335, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0368] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 336, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0369] The upstream and downstream primer sequences of the primer pairs for each site in Group 41 are as follows:

[0370] 1) The HLA-AF sequence is shown in SEQ ID NO. 337, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0371] 2) The HLA-BF sequence is shown in SEQ ID NO. 338, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0372] 3) The HLA-CF sequence is shown in SEQ ID NO. 339, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0373] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 340, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0374] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 341, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0375] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 342, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0376] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 343, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0377] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 344, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0378] The upstream and downstream primer sequences of the primer pairs for each site in Group 42 are as follows:

[0379] 1) The HLA-AF sequence is shown in SEQ ID NO. 345, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0380] 2) The HLA-BF sequence is shown in SEQ ID NO. 346, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0381] 3) The HLA-CF sequence is shown in SEQ ID NO. 347, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0382] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 348, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0383] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 349, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0384] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 350, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0385] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 351, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0386] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.352, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0387] The upstream and downstream primer sequences of the primer pairs for each site in Group 43 are as follows:

[0388] 1) The HLA-AF sequence is shown in SEQ ID NO. 353, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0389] 2) The HLA-BF sequence is shown in SEQ ID NO. 354, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0390] 3) The HLA-CF sequence is shown in SEQ ID NO. 355, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0391] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 356, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0392] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 357, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0393] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 358, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0394] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 359, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0395] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 360, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0396] The upstream and downstream primer sequences of the primer pairs for each site in Group 44 are as follows:

[0397] 1) The HLA-AF sequence is shown in SEQ ID NO. 361, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0398] 2) The HLA-BF sequence is shown in SEQ ID NO. 362, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0399] 3) The HLA-CF sequence is shown in SEQ ID NO. 363, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0400] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 364, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0401] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 365, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0402] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 366, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0403] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 367, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0404] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 368, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0405] The upstream and downstream primer sequences of the primer pairs for each site in group 45 are as follows:

[0406] 1) The HLA-AF sequence is shown in SEQ ID NO. 369, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0407] 2) The HLA-BF sequence is shown in SEQ ID NO. 370, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0408] 3) The HLA-CF sequence is shown in SEQ ID NO. 371, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0409] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 372, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0410] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 373, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0411] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 374, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0412] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 375, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0413] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 376, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0414] The upstream and downstream primer sequences of the primer pairs for each site in Group 46 are as follows:

[0415] 1) The HLA-AF sequence is shown in SEQ ID NO. 377, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0416] 2) The HLA-BF sequence is shown in SEQ ID NO. 378, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0417] 3) The HLA-CF sequence is shown in SEQ ID NO. 379, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0418] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 380, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0419] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 381, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0420] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 382, ​​and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0421] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 383, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0422] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 384, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0423] The upstream and downstream primer sequences of the primer pairs for each site in group 47 are as follows:

[0424] 1) The HLA-AF sequence is shown in SEQ ID NO. 385, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0425] 2) The HLA-BF sequence is shown in SEQ ID NO. 386, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0426] 3) The HLA-CF sequence is shown in SEQ ID NO. 387, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0427] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 388, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0428] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 389, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0429] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 390, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0430] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 391, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0431] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 392, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in Group 1;

[0432] The upstream and downstream primer sequences of the primer pairs for each site in Group 48 are as follows:

[0433] 1) The HLA-AF sequence is shown in SEQ ID NO. 393, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0434] 2) The HLA-BF sequence is shown in SEQ ID NO. 394, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0435] 3) The HLA-CF sequence is shown in SEQ ID NO. 395, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0436] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 396, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0437] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 397, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0438] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 398, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0439] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 399, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0440] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO.400, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in Group 1.

[0441] In the second aspect of the present invention, another primer set for HLA ultra-high resolution genotyping is provided, and the primer set is also used for HLA gene multiple amplification; the primer set includes primer pairs for HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3 / 4 / 5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 sites. In order to improve the inclusiveness of primers, scheme 2 adopts the design of nested primers, which covers the full-length exon sequence of 11 sites. The upstream and downstream primers and their related nucleotide sequences are shown in the following table.

[0442] The site can be set into 2 sets of upstream and downstream primers, and the upstream and downstream primers and related nucleotide sequences are shown in the following table:

[0443]

[0444]

[0445] In the third aspect of the present invention, there is provided use of the primer set as described in the first aspect in preparing an HLA ultra-high resolution horizontal genotyping detection kit.

[0446] In a fourth aspect of the present invention, an HLA ultra-high resolution horizontal genotyping detection kit is provided, wherein the detection kit comprises the primer set and detection reagent as described in the first aspect.

[0447] Preferably, the detection reagent includes the HLA gene rapid amplification enzyme system Flash PCR Master MixFor Long Fragment.

[0448] In a fifth aspect of the present invention, there is provided a use of the primer set as described in the second aspect in preparing an HLA ultra-high resolution horizontal genotyping detection kit.

[0449] In a sixth aspect of the present invention, an HLA ultra-high resolution horizontal genotyping detection kit is provided, wherein the detection kit comprises the primer set and detection reagent as described in the second aspect.

[0450] Preferably, the detection reagent includes HLA gene rapid amplification enzyme system Flash PCR Master MixFor Long Fragment and Index synthesis related oligonucleotide sequences;

[0451] The Index synthesis-related oligonucleotide sequence is selected from any one or more groups of Index01-Index48. The upstream and downstream primers of the primer pairs of Index01-Index48 and the related nucleotide sequences are shown in the following table:

[0452]

[0453]

[0454]

[0455]

[0456] In the seventh aspect of the present invention, a full-length and rapid sequencing method for HLA typing based on nanopore sequencing is provided, the method comprising using the above-mentioned primer set or kit to amplify the HLA gene in the sample to be tested, and then sequencing the amplified product.

[0457] In an eighth aspect of the present invention, a method for HLA genotyping for non-diagnostic and non-therapeutic purposes is provided, the method comprising using the above-mentioned primer set or kit to amplify the HLA genes in the sample to be tested, constructing the amplified products into a sequencing library, and then sequencing and performing bioinformatics analysis on the sequencing data to obtain full-length ultra-high resolution typing information including 11 HLA genes in the sample to be tested.

[0458] Compared with the prior art, the present invention has the following beneficial effects:

[0459] The primer set for HLA gene multiplex amplification provided by the present invention performs full-length amplification and full-length direct sequencing on 11 gene loci, including HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3 / 4 / 5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1, thereby improving the efficiency of gene amplification and ensuring the integrity and uniformity of sequencing. The full-length direct amplification and direct measurement of genes directly obtain the allele type of each gene, eliminating the splicing process between different exons, thereby avoiding a large number of ambiguous results generated in the first-generation sequencing and second-generation sequencing and analysis processes, thereby improving the accuracy of subsequent HLA haplotype genotyping.

[0460] The HLA genotyping method provided by the present invention uses the primer set of the first aspect and the rapid amplification enzyme described in the second aspect to rapidly amplify 11 target genes in clinical samples, further uses a rapid library construction method to build a library for the amplified products, and uses nanopore sequencing technology to sequence the library. The obtained Reads have more complete full-length coverage of the 11 genes, and the number of reads detected for different genes is more balanced, which is more advantageous. Reliable full-length amplification and nanopore length sequencing provide strong guarantees for organ transplantation, bone marrow matching, etc.

[0461] Specifically, compared with the previous HLA detection methods, the technical advantages of the present invention are as follows:

[0462] 1) The present invention provides a primer design method based on omega primers, nested primers and degenerate bases. The primers or primer combinations designed by the method can maximize the population coverage of the detection method, so that the indiscriminate screening effect of the HLA population is better. The present invention designs a set of 11 gene multiple amplification primer combinations for HLA typing based on the above method. Among them, the primers in the scheme one, that is, a part of the embodiment, can be Ω primers, including 3p arms, 5p arms and a loop separating the two arms. The 3p arm is a promoter for the polymerase extension reaction, the 5p arm is an anchor for binding to the DNA template sequence, and the loop separating the two arms contains anchor sequence 1, anchor sequence 2 and a barcode sequence used for library splitting. During amplification, the 3p arm and the 5p arm need to specifically bind to the template at the same time. When there is a mismatch between the 3p arm and the template, the stability of the ring structure will be shaken, resulting in the separation of the 3p arm from the template, reducing the probability of the polymerase extending the primer. Compared with traditional primers, this technology significantly increases the specificity of the primer. The present invention designs the 5p arm to be longer and partially degenerates the bases, so that even if there is a mismatch in the template, it can still bind effectively. This feature effectively improves the tolerance of the primers and reduces the risk of PCR failure, missed detection and allele preference. The nested primers in the second scheme, i.e., the second embodiment, also have the same purpose. While ensuring full-length gene amplification, the nested primer design and partial degenerate bases greatly increase the detection of large populations, and will not cause imbalance in the output of reads between genes and allele preference due to the appearance of SNP sites with lower population frequency. This primer set can achieve simultaneous amplification of HLA gene HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 sites in one reaction system, thereby improving the simplicity of the experiment, as well as the integrity, efficiency and uniformity of gene amplification and the accuracy of subsequent HLA genotyping.

[0463] 2) The present invention adopts the multi-blocked long-fragment PCR amplification enzyme system of Kangwei Century, Flash PCR MasterMix For Long Fragment (commercial product of Kangwei, item number CW3397), which contains Super DNA Polymerase, dNTP, unique extension factor and high-performance optimized buffer system. Only primers and templates need to be added to perform PCR experiments, reducing the complicated sample addition process. Super DNA Polymerase has both 5′→3′ DNA polymerase activity and 3′→5′ exonuclease activity, and its fidelity is about 80 times that of Taq DNA polymerase, and the amplified product has a blunt end. The enzyme is a high-fidelity enzyme that has been multiply blocked. It effectively blocks the polymerase activity at 55°C and below, and quickly recovers the polymerase activity after incubation at 98°C for 30s. It is suitable for amplifying target fragments of 100bp to 40,000bp, and is particularly suitable for amplifying fragments ≥10,000bp. The amplification speed is as high as 10kb / s, which greatly saves PCR reaction time. The enzyme can complete the amplification of long fragments in a very short time, which can greatly shorten the detection time. It can complete 30 cycles of amplification of DNA fragments of 10,000 to 160,000 bp in about 40 minutes.

[0464] 3) Example 1 of the present invention uses a rapid library construction kit. Omega primers have their own barcodes, so the overall library construction process includes: long-fragment PCR, PCR product magnetic bead purification and quantification, PCR product mixing, end repair + A, sequencing adapter ligation and magnetic bead purification (see Figure 1 ). Common third-generation library construction methods: long-fragment PCR, PCR product magnetic bead purification and quantification, end repair + A, barcode ligation, library magnetic bead purification and quantification, library mixing, sequencing adapter ligation, magnetic bead purification and quantification (see Figure 1 ). Compared with the common third-generation library construction method, the rapid method reduces the steps of barcode connection, library magnetic bead purification and quantification, and directly mixes the samples as a mixed sample for library construction after the first round of long-fragment PCR, which greatly reduces the workload of subsequent end repair + A, library magnetic bead purification and quantification. Although the overall library construction process of Example 2 is more complicated than that of Example 1, the interference of low-frequency SNPs in HLA large population detection is well achieved through nested primer design, thus laying the foundation for ultra-high resolution analysis and detection of large-scale potential HLA donors.

[0465] In summary, compared with previous HLA detection methods, Examples 1 and 2 of this patent achieve the following advantages: ① Full-length direct detection of single-gene haplotypes; ② Ultra-fast amplification, library construction and sequencing; ③ Large population detection, overcoming the interference of low-frequency SNPs; ④ Ultra-high-resolution typing of 11 loci of Class I and II. BRIEF DESCRIPTION OF THE DRAWINGS

[0466] Figure 1 This is a schematic diagram of the database construction process.

[0467] Figure 2 The results of HLA primer amplification of 3 clinical blood samples in Example 1.

[0468] Figure 3 This is the quality inspection result of the linker annealing electrophoresis in Example 2.

[0469] Figure 4 The results of HLA primer amplification of 3 clinical blood samples in Example 2 are shown. DETAILED DESCRIPTION

[0470] The technical scheme of the present invention will be clearly and completely described below in conjunction with embodiments, but the following embodiments are only used to illustrate the present invention and should not be regarded as limiting the scope of the present invention.

[0471] The primers in the examples of the present invention were synthesized by Sangon Biotech (Shanghai) Co., Ltd. The remaining components in the kit are all commercially available products from Jiangsu Kangwei Century Biotechnology Co., Ltd., and can also be replaced by other similar commercial products. The following method is used for genomic DNA extraction in the following examples:

[0472] Blood samples need to be extracted for gDNA using the Cowin Blood Genomic DNA Mini Kit (0.1-1mL, cw20877). If automated extraction is used, use the CWY129S-96 reagent. If the sample is DNA, use 0.5x Cowin Century DNA Purification Magnetic Beads (Cat. No.: CW3171M) for purification.

[0473] Embodiment 1

[0474] 1. Primer and barcode synthesis

[0475] Primer design and synthesis: HLA-related gene sequences were searched on NCBI (National Center of Biotechnology Information). According to the primer design principles of multiplex PCR, specific primer sequences and barcode sequences were designed as shown in Tables 1 and 2 below.

[0476] Table 1.48 barcode sequences

[0477]

[0478]

[0479] Table 2 Primer information

[0480]

[0481]

[0482]

[0483] The present invention provides 48 barcode sequences, which can be used to distinguish 48 different samples during sequencing.

[0484] The present invention provides 48 primer sets, and the primer pair sequences of each site in each group are different from the primer pair sequences of each site in the first group (see Table 2) only in that the barcode sequences in the Ω primers of each site are different, the Ω primers of each site in the second group contain the barcode_2 sequence, the Ω primers of each site in the third group contain the barcode_3 sequence, and so on.

[0485] The upstream and downstream primer sequences of the primer pairs for each site of Group 2 to Group 48 are as follows:

[0486] The upstream and downstream primer sequences of the primer pairs for each site in Group 2 are as follows:

[0487] 1) The HLA-AF sequence is shown in SEQ ID NO. 25, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0488] 2) The HLA-BF sequence is shown in SEQ ID NO. 26, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0489] 3) The HLA-CF sequence is shown in SEQ ID NO. 27, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0490] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 28, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0491] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 29, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0492] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 30, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0493] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 31, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0494] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.32, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0495] The upstream and downstream primer sequences of the primer pairs for each site in Group 3 are as follows:

[0496] 1) The HLA-AF sequence is shown in SEQ ID NO. 33, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0497] 2) The HLA-BF sequence is shown in SEQ ID NO. 34, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0498] 3) The HLA-CF sequence is shown in SEQ ID NO. 35, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0499] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 36, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0500] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 37, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0501] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 38, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0502] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 39, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0503] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 40, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0504] The upstream and downstream primer sequences of the primer pairs for each site in Group 4 are as follows:

[0505] 1) The HLA-AF sequence is shown in SEQ ID NO. 41, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0506] 2) The HLA-BF sequence is shown in SEQ ID NO. 42, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0507] 3) The HLA-CF sequence is shown in SEQ ID NO. 43, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0508] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 44, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0509] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 45, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0510] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 46, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0511] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 47, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0512] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 48, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0513] The upstream and downstream primer sequences of the primer pairs for each site in Group 5 are as follows:

[0514] 1) The HLA-AF sequence is shown in SEQ ID NO. 49, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0515] 2) The HLA-BF sequence is shown in SEQ ID NO. 50, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0516] 3) The HLA-CF sequence is shown in SEQ ID NO.51, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0517] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 52, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0518] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 53, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0519] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 54, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0520] 7) The HLA-DQB-F sequence is shown in SEQ ID NO.55, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0521] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.56, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0522] The upstream and downstream primer sequences of the primer pairs for each site in Group 6 are as follows:

[0523] 1) The HLA-AF sequence is shown in SEQ ID NO.57, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0524] 2) The HLA-BF sequence is shown in SEQ ID NO. 58, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0525] 3) The HLA-CF sequence is shown in SEQ ID NO. 59, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0526] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 60, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0527] 5) The HLA-DPB-F sequence is shown in SEQ ID NO.61, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0528] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 62, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0529] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 63, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0530] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.64, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0531] The upstream and downstream primer sequences of the primer pairs for each site in Group 7 are as follows:

[0532] 1) The HLA-AF sequence is shown in SEQ ID NO.65, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0533] 2) The HLA-BF sequence is shown in SEQ ID NO. 66, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0534] 3) The HLA-CF sequence is shown in SEQ ID NO. 67, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0535] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 68, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0536] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 69, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0537] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 70, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0538] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 71, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0539] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.72, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0540] The upstream and downstream primer sequences of the primer pairs for each site in Group 8 are as follows:

[0541] 1) The HLA-AF sequence is shown in SEQ ID NO. 73, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0542] 2) The HLA-BF sequence is shown in SEQ ID NO. 74, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0543] 3) The HLA-CF sequence is shown in SEQ ID NO. 75, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0544] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 76, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0545] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 77, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0546] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 78, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0547] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 79, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0548] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 80, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0549] The upstream and downstream primer sequences of the primer pairs for each site in Group 9 are as follows:

[0550] 1) The HLA-AF sequence is shown in SEQ ID NO.81, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0551] 2) The HLA-BF sequence is shown in SEQ ID NO.82, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0552] 3) The HLA-CF sequence is shown in SEQ ID NO. 83, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0553] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 84, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0554] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 85, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0555] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 86, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0556] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 87, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0557] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.88, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0558] The upstream and downstream primer sequences of the primer pairs for each site in Group 10 are as follows:

[0559] 1) The HLA-AF sequence is shown in SEQ ID NO.89, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0560] 2) The HLA-BF sequence is shown in SEQ ID NO.90, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0561] 3) The HLA-CF sequence is shown in SEQ ID NO.91, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0562] 4) The HLA-DPA-R sequence is shown in SEQ ID NO.92, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0563] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 93, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0564] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 94, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0565] 7) The HLA-DQB-F sequence is shown in SEQ ID NO.95, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0566] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.96, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0567] The upstream and downstream primer sequences of the primer pairs for each site in Group 11 are as follows:

[0568] 1) The HLA-AF sequence is shown in SEQ ID NO.97, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0569] 2) The HLA-BF sequence is shown in SEQ ID NO.98, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0570] 3) The HLA-CF sequence is shown in SEQ ID NO. 99, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0571] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 100, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0572] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 101, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0573] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 102, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0574] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 103, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0575] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 104, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0576] The upstream and downstream primer sequences of the primer pairs for each site in Group 12 are as follows:

[0577] 1) The HLA-AF sequence is shown in SEQ ID NO. 105, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0578] 2) The HLA-BF sequence is shown in SEQ ID NO. 106, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0579] 3) The HLA-CF sequence is shown in SEQ ID NO. 107, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0580] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 108, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0581] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 109, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0582] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 110, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0583] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 111, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0584] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.112, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0585] The upstream and downstream primer sequences of the primer pairs for each site in Group 13 are as follows:

[0586] 1) The HLA-AF sequence is shown in SEQ ID NO. 113, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0587] 2) The HLA-BF sequence is shown in SEQ ID NO. 114, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0588] 3) The HLA-CF sequence is shown in SEQ ID NO. 115, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0589] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 116, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0590] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 117, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0591] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 118, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0592] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 119, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0593] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 120, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0594] The upstream and downstream primer sequences of the primer pairs for each site in Group 14 are as follows:

[0595] 1) The HLA-AF sequence is shown in SEQ ID NO. 121, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0596] 2) The HLA-BF sequence is shown in SEQ ID NO. 122, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0597] 3) The HLA-CF sequence is shown in SEQ ID NO. 123, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0598] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 124, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0599] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 125, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0600] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 126, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0601] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 127, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0602] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 128, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0603] The upstream and downstream primer sequences of the primer pairs for each site in Group 15 are as follows:

[0604] 1) The HLA-AF sequence is shown in SEQ ID NO. 129, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0605] 2) The HLA-BF sequence is shown in SEQ ID NO. 130, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0606] 3) The HLA-CF sequence is shown in SEQ ID NO. 131, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0607] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 132, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0608] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 133, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0609] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 134, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0610] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 135, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0611] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 136, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0612] The upstream and downstream primer sequences of the primer pairs for each site in Group 16 are as follows:

[0613] 1) The HLA-AF sequence is shown in SEQ ID NO. 137, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0614] 2) The HLA-BF sequence is shown in SEQ ID NO. 138, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0615] 3) The HLA-CF sequence is shown in SEQ ID NO. 139, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0616] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 140, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0617] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 141, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0618] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 142, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0619] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 143, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0620] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 144, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0621] The upstream and downstream primer sequences of the primer pairs for each site in Group 17 are as follows:

[0622] 1) The HLA-AF sequence is shown in SEQ ID NO. 145, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0623] 2) The HLA-BF sequence is shown in SEQ ID NO. 146, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0624] 3) The HLA-CF sequence is shown in SEQ ID NO. 147, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0625] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 148, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0626] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 149, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0627] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 150, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0628] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 151, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0629] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.152, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0630] The upstream and downstream primer sequences of the primer pairs for each site in Group 18 are as follows:

[0631] 1) The HLA-AF sequence is shown in SEQ ID NO. 153, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0632] 2) The HLA-BF sequence is shown in SEQ ID NO. 154, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0633] 3) The HLA-CF sequence is shown in SEQ ID NO. 155, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0634] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 156, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0635] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 157, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0636] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 158, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0637] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 159, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0638] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 160, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0639] The upstream and downstream primer sequences of the primer pairs for each site in Group 19 are as follows:

[0640] 1) The HLA-AF sequence is shown in SEQ ID NO. 161, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0641] 2) The HLA-BF sequence is shown in SEQ ID NO. 162, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0642] 3) The HLA-CF sequence is shown in SEQ ID NO. 163, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0643] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 164, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0644] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 165, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0645] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 166, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0646] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 167, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0647] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 168, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0648] The upstream and downstream primer sequences of the primer pairs for each site in Group 20 are as follows:

[0649] 1) The HLA-AF sequence is shown in SEQ ID NO. 169, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0650] 2) The HLA-BF sequence is shown in SEQ ID NO. 170, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0651] 3) The HLA-CF sequence is shown in SEQ ID NO. 171, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0652] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 172, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0653] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 173, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0654] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 174, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0655] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 175, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0656] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 176, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0657] The upstream and downstream primer sequences of the primer pairs for each site in Group 21 are as follows:

[0658] 1) The HLA-AF sequence is shown in SEQ ID NO. 177, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0659] 2) The HLA-BF sequence is shown in SEQ ID NO. 178, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0660] 3) The HLA-CF sequence is shown in SEQ ID NO. 179, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0661] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 180, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0662] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 181, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0663] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 182, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0664] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 183, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0665] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 184, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0666] The upstream and downstream primer sequences of the primer pairs for each site in Group 22 are as follows:

[0667] 1) The HLA-AF sequence is shown in SEQ ID NO. 185, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0668] 2) The HLA-BF sequence is shown in SEQ ID NO. 186, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0669] 3) The HLA-CF sequence is shown in SEQ ID NO. 187, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0670] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 188, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0671] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 189, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0672] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 190, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0673] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 191, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0674] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 192, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0675] The upstream and downstream primer sequences of the primer pairs for each site in Group 23 are as follows:

[0676] 1) The HLA-AF sequence is shown in SEQ ID NO. 193, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0677] 2) The HLA-BF sequence is shown in SEQ ID NO. 194, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0678] 3) The HLA-CF sequence is shown in SEQ ID NO. 195, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0679] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 196, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0680] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 197, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0681] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 198, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0682] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 199, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0683] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 200, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0684] The upstream and downstream primer sequences of the primer pairs for each site in Group 24 are as follows:

[0685] 1) The HLA-AF sequence is shown in SEQ ID NO. 201, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0686] 2) The HLA-BF sequence is shown in SEQ ID NO. 202, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0687] 3) The HLA-CF sequence is shown in SEQ ID NO. 203, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0688] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 204, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0689] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 205, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0690] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 206, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0691] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 207, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0692] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 208, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0693] The upstream and downstream primer sequences of the primer pairs for each site in Group 25 are as follows:

[0694] 1) The HLA-AF sequence is shown in SEQ ID NO. 209, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0695] 2) The HLA-BF sequence is shown in SEQ ID NO. 210, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0696] 3) The HLA-CF sequence is shown in SEQ ID NO. 211, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0697] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 212, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0698] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 213, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0699] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 214, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0700] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 215, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0701] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 216, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0702] The upstream and downstream primer sequences of the primer pairs for each site in Group 26 are as follows:

[0703] 1) The HLA-AF sequence is shown in SEQ ID NO. 217, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0704] 2) The HLA-BF sequence is shown in SEQ ID NO. 218, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0705] 3) The HLA-CF sequence is shown in SEQ ID NO. 219, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0706] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 220, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0707] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 221, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0708] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 222, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0709] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 223, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0710] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 224, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0711] The upstream and downstream primer sequences of the primer pairs for each site in Group 27 are as follows:

[0712] 1) The HLA-AF sequence is shown in SEQ ID NO. 225, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0713] 2) The HLA-BF sequence is shown in SEQ ID NO. 226, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0714] 3) The HLA-CF sequence is shown in SEQ ID NO. 227, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0715] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 228, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0716] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 229, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0717] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 230, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0718] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 231, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0719] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 232, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0720] The upstream and downstream primer sequences of the primer pairs for each site in Group 28 are as follows:

[0721] 1) The HLA-AF sequence is shown in SEQ ID NO. 233, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0722] 2) The HLA-BF sequence is shown in SEQ ID NO. 234, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0723] 3) The HLA-CF sequence is shown in SEQ ID NO. 235, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0724] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 236, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0725] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 237, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0726] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 238, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0727] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 239, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0728] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 240, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0729] The upstream and downstream primer sequences of the primer pairs for each site in Group 29 are as follows:

[0730] 1) The HLA-AF sequence is shown in SEQ ID NO. 241, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0731] 2) The HLA-BF sequence is shown in SEQ ID NO. 242, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0732] 3) The HLA-CF sequence is shown in SEQ ID NO. 243, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0733] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 244, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0734] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 245, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0735] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 246, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0736] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 247, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0737] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 248, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0738] The upstream and downstream primer sequences of the primer pairs for each site in Group 30 are as follows:

[0739] 1) The HLA-AF sequence is shown in SEQ ID NO. 249, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0740] 2) The HLA-BF sequence is shown in SEQ ID NO. 250, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0741] 3) The HLA-CF sequence is shown in SEQ ID NO. 251, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0742] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 252, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0743] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 253, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0744] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 254, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0745] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 255, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0746] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 256, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0747] The upstream and downstream primer sequences of the primer pairs for each site in Group 31 are as follows:

[0748] 1) The HLA-AF sequence is shown in SEQ ID NO. 257, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0749] 2) The HLA-BF sequence is shown in SEQ ID NO. 258, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0750] 3) The HLA-CF sequence is shown in SEQ ID NO. 259, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0751] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 260, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0752] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 261, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0753] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 262, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0754] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 263, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0755] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 264, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0756] The upstream and downstream primer sequences of the primer pairs for each site in Group 32 are as follows:

[0757] 1) The HLA-AF sequence is shown in SEQ ID NO. 265, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0758] 2) The HLA-BF sequence is shown in SEQ ID NO. 266, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0759] 3) The HLA-CF sequence is shown in SEQ ID NO. 267, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0760] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 268, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0761] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 269, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0762] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 270, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0763] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 271, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0764] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 272, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0765] The upstream and downstream primer sequences of the primer pairs for each site in Group 33 are as follows:

[0766] 1) The HLA-AF sequence is shown in SEQ ID NO. 273, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0767] 2) The HLA-BF sequence is shown in SEQ ID NO. 274, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0768] 3) The HLA-CF sequence is shown in SEQ ID NO. 275, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0769] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 276, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0770] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 277, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0771] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 278, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0772] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 279, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0773] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 280, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0774] The upstream and downstream primer sequences of the primer pairs for each site in Group 34 are as follows:

[0775] 1) The HLA-AF sequence is shown in SEQ ID NO. 281, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0776] 2) The HLA-BF sequence is shown in SEQ ID NO. 282, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0777] 3) The HLA-CF sequence is shown in SEQ ID NO. 283, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0778] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 284, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0779] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 285, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0780] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 286, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0781] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 287, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0782] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 288, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0783] The upstream and downstream primer sequences of the primer pairs for each site in Group 35 are as follows:

[0784] 1) The HLA-AF sequence is shown in SEQ ID NO. 289, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0785] 2) The HLA-BF sequence is shown in SEQ ID NO. 290, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0786] 3) The HLA-CF sequence is shown in SEQ ID NO. 291, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0787] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 292, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0788] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 293, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0789] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 294, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0790] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 295, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0791] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 296, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0792] The upstream and downstream primer sequences of the primer pairs for each site in Group 36 are as follows:

[0793] 1) The HLA-AF sequence is shown in SEQ ID NO. 297, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0794] 2) The HLA-BF sequence is shown in SEQ ID NO. 298, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0795] 3) The HLA-CF sequence is shown in SEQ ID NO. 299, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0796] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 300, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0797] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 301, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0798] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 302, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0799] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 303, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0800] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 304, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1;

[0801] The upstream and downstream primer sequences of the primer pairs for each site in Group 37 are as follows:

[0802] 1) The HLA-AF sequence is shown in SEQ ID NO. 305, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0803] 2) The HLA-BF sequence is shown in SEQ ID NO. 306, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0804] 3) The HLA-CF sequence is shown in SEQ ID NO. 307, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0805] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 308, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0806] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 309, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0807] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 310, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0808] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 311, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0809] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.312, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0810] The upstream and downstream primer sequences of the primer pairs for each site in Group 38 are as follows:

[0811] 1) The HLA-AF sequence is shown in SEQ ID NO. 313, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0812] 2) The HLA-BF sequence is shown in SEQ ID NO. 314, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0813] 3) The HLA-CF sequence is shown in SEQ ID NO. 315, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0814] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 316, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0815] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 317, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0816] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 318, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0817] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 319, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0818] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 320, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0819] The upstream and downstream primer sequences of the primer pairs for each site in Group 39 are as follows:

[0820] 1) The HLA-AF sequence is shown in SEQ ID NO. 321, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0821] 2) The HLA-BF sequence is shown in SEQ ID NO. 322, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0822] 3) The HLA-CF sequence is shown in SEQ ID NO. 323, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0823] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 324, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0824] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 325, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0825] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 326, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0826] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 327, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0827] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 328, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0828] The upstream and downstream primer sequences of the primer pairs for each site in Group 40 are as follows:

[0829] 1) The HLA-AF sequence is shown in SEQ ID NO. 329, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0830] 2) The HLA-BF sequence is shown in SEQ ID NO. 330, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0831] 3) The HLA-CF sequence is shown in SEQ ID NO. 331, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0832] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 332, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0833] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 333, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0834] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 334, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0835] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 335, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0836] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 336, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0837] The upstream and downstream primer sequences of the primer pairs for each site in Group 41 are as follows:

[0838] 1) The HLA-AF sequence is shown in SEQ ID NO. 337, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0839] 2) The HLA-BF sequence is shown in SEQ ID NO. 338, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0840] 3) The HLA-CF sequence is shown in SEQ ID NO. 339, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0841] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 340, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0842] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 341, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0843] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 342, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0844] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 343, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0845] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 344, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0846] The upstream and downstream primer sequences of the primer pairs for each site in Group 42 are as follows:

[0847] 1) The HLA-AF sequence is shown in SEQ ID NO. 345, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0848] 2) The HLA-BF sequence is shown in SEQ ID NO. 346, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0849] 3) The HLA-CF sequence is shown in SEQ ID NO. 347, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0850] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 348, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0851] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 349, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0852] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 350, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0853] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 351, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0854] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.352, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0855] The upstream and downstream primer sequences of the primer pairs for each site in Group 43 are as follows:

[0856] 1) The HLA-AF sequence is shown in SEQ ID NO. 353, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0857] 2) The HLA-BF sequence is shown in SEQ ID NO. 354, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0858] 3) The HLA-CF sequence is shown in SEQ ID NO. 355, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0859] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 356, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0860] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 357, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0861] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 358, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0862] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 359, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0863] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 360, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0864] The upstream and downstream primer sequences of the primer pairs for each site in Group 44 are as follows:

[0865] 1) The HLA-AF sequence is shown in SEQ ID NO. 361, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0866] 2) The HLA-BF sequence is shown in SEQ ID NO. 362, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0867] 3) The HLA-CF sequence is shown in SEQ ID NO. 363, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0868] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 364, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0869] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 365, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0870] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 366, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0871] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 367, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0872] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 368, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0873] The upstream and downstream primer sequences of the primer pairs for each site in group 45 are as follows:

[0874] 1) The HLA-AF sequence is shown in SEQ ID NO. 369, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0875] 2) The HLA-BF sequence is shown in SEQ ID NO. 370, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0876] 3) The HLA-CF sequence is shown in SEQ ID NO. 371, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0877] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 372, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0878] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 373, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0879] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 374, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0880] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 375, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0881] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 376, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0882] The upstream and downstream primer sequences of the primer pairs for each site in Group 46 are as follows:

[0883] 1) The HLA-AF sequence is shown in SEQ ID NO. 377, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0884] 2) The HLA-BF sequence is shown in SEQ ID NO. 378, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0885] 3) The HLA-CF sequence is shown in SEQ ID NO. 379, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0886] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 380, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0887] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 381, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0888] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 382, ​​and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0889] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 383, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0890] 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 384, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1;

[0891] The upstream and downstream primer sequences of the primer pairs for each site in group 47 are as follows:

[0892] 1) The HLA-AF sequence is shown in SEQ ID NO. 385, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0893] 2) The HLA-BF sequence is shown in SEQ ID NO. 386, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0894] 3) The HLA-CF sequence is shown in SEQ ID NO. 387, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0895] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 388, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0896] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 389, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0897] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 390, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0898] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 391, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0899] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 392, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in Group 1;

[0900] The upstream and downstream primer sequences of the primer pairs for each site in Group 48 are as follows:

[0901] 1) The HLA-AF sequence is shown in SEQ ID NO. 393, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1;

[0902] 2) The HLA-BF sequence is shown in SEQ ID NO. 394, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1;

[0903] 3) The HLA-CF sequence is shown in SEQ ID NO. 395, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1;

[0904] 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 396, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1;

[0905] 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 397, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1;

[0906] 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 398, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1;

[0907] 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 399, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1;

[0908] 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO.400, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in Group 1.

[0909] The above primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd. After sequence synthesis, quality inspection and quantification were performed, and then diluted to the required concentration as needed.

[0910] The above primer set can be used to prepare an HLA ultra-high resolution horizontal genotyping detection kit, which also includes an HLA gene rapid amplification enzyme system Flash PCR Master Mix ForLong Fragment.

[0911] When testing 1 sample, you can select one of the 48 primer sets and perform the test according to the following method. When testing 2 samples, you can select 2 of the 48 primer sets and perform the test according to the following method. And so on. Up to 11 gene loci of 48 different samples can be tested simultaneously. If necessary, the number of barcode types can be further expanded to test more samples simultaneously in a single batch.

[0912] 2. Target gene amplification

[0913] 1. Primer dilution and mixing

[0914] The 24 HLA primers were diluted to 100uM with low-TE (10mM Tris, 1mM EDTA, pH 8.0) and the following ratios were used: HLA-AF:HLA-A-R1:HLA-A-R2:HLA-BF:HLA-B-R1:HLA-B-R2:HLA-CF:HLA-C-R1:HLA-C-R2:HLA-DPA-F1:HLA-DPA-F2:HLA-DPA-R:HLA-DPB-F:HLA-DPB-R1:HLA-DPB-R2: HLA-DQA-F:HLA-DQA-R1:HLA-DQA-R2:HLA-DQB-F:HLA-DQB-R1:HLA-DQB-R2:HLA-DRB1345-F:HLA-DR B1345-R1:HLA-DRB1345-R2=10:7:7:10:7:7:10:7:7:6:6:10:10:6:6:10:7:7:10:6:6:15:8:8 for mixing.

[0915] 2. Amplification system

[0916] Table 3. Group 1 reaction system

[0917] Reagents Volume(ul) FlashPCRMasterMixForLongFragment(CW3397M) 12.5 Primers 1 RNase-FreeWater(CW0612M) 11.5-x DNA (30ng) x Total volume 25

[0918] Prepare the long-fragment amplification system in a PCR tube according to Table 3 on an ice box, add 30 ng of DNA template, pipette to mix, and centrifuge.

[0919] 3. Amplification Procedure

[0920] Place the PCR tube into the PCR instrument and perform the reaction according to the conditions in Table 4.

[0921] Table 4. Long fragment amplification program

[0922]

[0923] 3. Purification, Quantification and Fragment Analysis of PCR Amplification Products

[0924] 1. PCR Product Purification

[0925] (1) Preparation of reagents and consumables: Take out the Kangwei Century DNA purification magnetic beads (Cat. No.: CW3171M) stored at 4°C and equilibrate them at room temperature for about 30 minutes; prepare 80% alcohol and prepare it at 440uL / reaction.

[0926] (2) After the magnetic beads have reached room temperature, use a vortex shaker to mix the beads at medium speed for 1 minute. Add 12.5 μl (0.5X) of magnetic bead suspension to the PCR tube. Mix by pipetting at half scale for about 10 times until the beads are mixed.

[0927] (3) Incubate at room temperature for 5 minutes.

[0928] (4) After incubation, place the PCR tube on a magnetic rack for about 3 minutes. When the solution becomes clear, discard the supernatant completely, taking care not to let the pipette tip touch the magnetic beads.

[0929] (5) Keep the PCR tubes on the magnetic rack, add 200 μl of freshly prepared 80% alcohol to each tube, and let it stand at room temperature for about 30 seconds.

[0930] (6) Wait until the solution becomes clear and remove the supernatant, taking care not to let the tip of the gun touch the magnetic beads.

[0931] (7) Repeat steps (5) and (6) once for a second wash.

[0932] (8) Keep the PCR tube fixed on the magnetic rack, open the tube cap, and dry it at room temperature for 3 to 5 minutes until the magnetic beads are no longer reflective and cracked.

[0933] (9) Remove the PCR tube from the magnetic rack, add 35uL of deionized water to each well, pipette to mix, incubate at room temperature for about 10 minutes and centrifuge briefly.

[0934] (10) Place the PCR tube on a magnetic rack. When the solution becomes clear, transfer about 30uL of the purified amplification product to a new PCR tube. Be careful not to let the tip of the pipette touch the magnetic beads.

[0935] 2. PCR product quantification

[0936] Quantitative use of PCR products dsDNA BRAssay Kits (Cat. No.: Q32853), Qubit TM Test tubes (Cat. No.: Q32856) and Qubit TM 4 Fluorometer (Cat. No.: Q33238) was used for quantification according to the standard procedure in the instruction manual, and the product concentration was recorded;

[0937] 3. PCR product fragment analysis

[0938] The fragment analysis of PCR products was performed on the Agilent 4200 TapeStation capillary electrophoresis analyzer and supporting reagents.

[0939] IV. Construction of the third generation HLA library

[0940] 1. PCR product mixing

[0941] Mix the PCR amplification products in equal amounts according to their mass in a 0.2 ml PCR tube. The total amount of amplicon samples input is 2000 ng (for example, for 20 samples, 100 ng / sample template can be input). If the number of samples is less than 2000 ng, all of them should be input, mixed gently, centrifuged briefly, and the reaction solution collected at the bottom of the tube to ensure that there are no bubbles in the reaction solution.

[0942] 2. End repair

[0943] (1) ERAT Mix (CW3045) and 10×ERAT Buffer (CW3045) are the enzyme and buffer for end repair and +A, respectively. Prepare the end repair system on an ice box according to Table 5 and dispense into PCR tubes. Then add ≥2000ng of PCR product into the PCR tubes, vortex gently to mix, and then centrifuge briefly to collect the reaction solution at the bottom of the tube. Make sure there are no bubbles in the reaction solution.

[0944] Table 5. End repair reaction system

[0945] Components Volume (μL) Pooling of PCR products X Nuclease-free water 21.5-X 10×ERAT Buffer (CW3045) 2.5 ERATMix(CW3045) 1 Total volume 25

[0946] (2) Place the above PCR tube into a PCR instrument and perform the reaction according to Table 6.

[0947] Table 6. End repair reaction conditions

[0948]

[0949] (3) After the reaction is completed, proceed directly to the next step.

[0950] 3. Sequencing adapter ligation

[0951] (1) Prepare the mixture according to Table 7 in a 0.2 mL PCR tube, mix gently, and then centrifuge briefly to collect the reaction solution at the bottom of the tube, making sure there are no bubbles in the reaction solution.

[0952] Table 7. Connector ligation reaction solution system

[0953] Components Volume (μl) End repair products 25 Hydration 5.5 LA connector 4 T4 DNA ligase Buffer (CW2701) 12 T4 DNA ligase, 15 U / μL (CW2701) 3.5 Total volume 50

[0954] (2) Place the PCR tube containing the reaction mixture from the previous step on a PCR instrument and react according to the conditions in Table 8:

[0955] Table 8. Adapter ligation reaction conditions

[0956]

[0957] 4. Magnetic Bead Purification

[0958] (1) Equilibrate the magnetic beads to room temperature in advance, add the reaction system to 25 μL (0.5X) of Conway DNA purification magnetic beads (1.5 mL centrifuge tube), and gently mix by flicking or pipetting; incubate at room temperature for 5 min;

[0959] (2) Transfer the test tube to the magnetic rack and let it stand for 2-5 minutes until the liquid becomes clear. Carefully discard the supernatant (be careful not to touch the magnetic beads).

[0960] (3) Remove the EP tube from the magnetic rack, slowly add 125 μL SFB (short fragment buffer, SFB) along the side where the magnetic beads are adsorbed, rotate the EP tube to gently resuspend the magnetic beads and then put it back on the magnetic rack (the magnetic beads are not attached to the wall to form aggregates), and after the liquid becomes clear, aspirate the clear liquid and be careful not to touch the magnetic beads);

[0961] (4) Repeat the previous step (using SFB for secondary cleaning and impurity removal);

[0962] (5) Remove the EP tube and centrifuge it using a mini centrifuge. Use a pipette to carefully discard the remaining liquid. Open the tube cover and air dry it for 30 seconds (do not allow it to dry out and crack).

[0963] (6) Add 35 μL of Elution Buffer (EB) to elute nucleic acids, flick the tube to mix, and incubate at 37°C for 10 min (tap the tube every two minutes);

[0964] (7) Place the EP tube back into the magnetic rack until the magnetic beads are attracted to one side of the magnetic rack or the mixed solution becomes clear. Transfer 32 μL of the supernatant into a clean PCR tube.

[0965] 5. Library Quantification

[0966] (1) Take 1 μL DNA library for Qubit fluorescence quantification and record the concentration.

[0967] (2) Take 150 ng of library for experiment.

[0968] 6. Sequencing

[0969] The sequencing platform was the PromethlON 2Solo nanopore sequencer from Oxford Nanopore Technologies (ONT).

[0970] 1. Chip loading and sequencing

[0971] (1) Reagent preparation: Take out Sequencing Buffer (SB), Library Beads (LIB) or Library Solution (LIS), Flow Cell Tether (FCT), and Flow Cell Flush (FCF) from the refrigerator, thaw them, vortex mix, and place on ice.

[0972] (2) Use a 1.5 ml centrifuge tube to prepare the reagents (priming mix) according to the system in Table 9:

[0973] Table 9. Primingmix system configuration

[0974]

[0975] (3) Mix the priming mix by pipetting and add 800 μl of priming mix through the flow cell priming port (before adding liquid, adjust the 1 ml pipette to 200 μl, insert the pipette tip into the flow cell priming port, and aspirate back to 220-230 μl);

[0976] (4) Let stand for 5 minutes;

[0977] (5) Use a 1.5 ml centrifuge tube to prepare the library for loading system according to Table 10:

[0978] Table 10. Library for loading system configuration

[0979] Reagents volume Sequencing Buffer (SB) 37.5μl LibraryBeads(LIB) (mix by pipetting before aspiration) 25.5μl DNA library Xμl (about 15fmol, can be higher if the sample size is small) EB 12-X Total 75μl

[0980] (6) Add 200 μl of priming mix through the flow cell priming port (before adding liquid, adjust the 1 ml pipette to 200 μl, insert the pipette tip into the flow cell priming port, and aspirate back to 220-230 μl);

[0981] (7) Add 75 μl of library for loading reagent through the SpotON sample port (mix well before adding and drip at a constant rate);

[0982] (8) Cover the SpotON sample port and the flow cell priming port.

[0983] (9) Start sequencing (pay attention to the proofreading kit number).

[0984] 2. Clean the chip

[0985] (1) Take out Wash Mix (WMX), Wash Diluent (DIL), and Storage Buffer (S) and thaw them at room temperature. Wash Mix (WMX) reagent can be mixed by inversion, but cannot be vortexed. Wash Diluent (DIL) and Storage Buffer (S) reagents can be mixed by vortexing.

[0986] (2) Prepare Flow Cell Wash Mix according to Table 11.

[0987] Table 11. Flow Cell Wash Mix

[0988]

[0989]

[0990] (3) Mix well with a pipette and place on ice;

[0991] (4) Open the flow cell priming port (before adding liquid, adjust the 1 ml pipette to 200 μl, insert the pipette tip into the flow cell priming port, and aspirate back to 220-230 μl), and add 400 μl Flow Cell Wash Mix at a constant speed;

[0992] (5) Leave at room temperature for 30 minutes;

[0993] (6) Open the flow cell priming port (before adding liquid, adjust 200 μl with a 1 ml pipette, insert the pipette tip into the flow cell priming port, and aspirate back to 220-230 μl), and add 500 μl Storage Buffer(S) at a constant speed;

[0994] (7) Cover the priming port and test the number of remaining available holes;

[0995] (8) sucking waste liquid from the waste liquid outlet and discarding it;

[0996] (9) Mark the chip with the expiration date and the number of remaining available wells, and store it at 4°C.

[0997] 7. Data Analysis

[0998] The data from sequencing was in POD5 / FAST5 file format, which was converted into fastq data format using the dorado analysis module and split according to the barcode information to obtain sequencing data corresponding to each sample. Then, the sequencing data was analyzed using CWBIO-LINKAGE software from Kangwei Century, and the typing results of HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3 / 4 / 5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 loci were finally determined.

[0999] 8. Results

[1000] 1. The results of the 11-site PCR amplification product fragment analysis of HLA in 3 clinical blood samples are shown in Figure 2 The amplified fragments were all larger than 4000 bp. Overall, the amplified fragments were mainly concentrated between 5000 bp and 10000 bp. The fragment sizes were in line with expectations, and the overall uniformity of the bands was relatively good.

[1001] 2. The full-length HLA gene sequencing method based on nanopore technology established in the embodiment was used to test 30 national reference samples. Ultra-high-resolution typing results were obtained for 11 sites of the 30 national reference samples (see Table 12), and the consistency rate with the national reference standard typing results was 100%.

[1002] Table 12. Test results of HLA national reference products

[1003]

[1004]

[1005]

[1006]

[1007]

[1008]

[1009]

[1010]

[1011] 3. 28 clinical blood samples were tested using the embodiment, and high-resolution HLA typing results were obtained using the patented method (see Table 13), among which a new allele HLA-A*24:NEW was identified in the sample HLA-S2.

[1012] Table 13. HLA analysis results of 28 clinical blood samples

[1013]

[1014]

[1015]

[1016] Embodiment 2

[1017] Primer design and synthesis The HLA-related gene sequences were searched on NCBI (National Center of Biotechnology Information). According to the primer design principles of multiplex PCR, specific primer sequences and index sequences were designed (Tables 14 and 15 below).

[1018] Table 14 Specific primer sequences

[1019]

[1020]

[1021] Table 15 Index synthesis related oligonucleotide sequences

[1022]

[1023]

[1024]

[1025]

[1026]

[1027]

[1028]

[1029]

[1030] The above primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd. After the sequence synthesis, quality inspection and quantification were performed, and then diluted to the required concentration as needed. The present invention provides 48 Index sequences, which can be used to distinguish 48 different samples during sequencing.

[1031] The primer set in Table 14 above can be used to prepare an HLA ultra-high resolution horizontal genotyping detection kit, which also includes an HLA gene rapid amplification enzyme system Flash PCR Master Mix ForLong Fragment and the Index synthesis-related oligonucleotide sequences in Table 15.

[1032] When testing 1 sample, you can select one of the 48 indexes for labeling and perform the test as follows. When testing 2 samples, you can select 2 of the 48 indexes for labeling and perform the test as follows. Similarly, up to 48 different samples can be labeled at the same time. If necessary, you can continue to expand the number of index types to test more samples in a single batch.

[1033] 1. Preparation of double-stranded nucleic acid for index sample label connection

[1034] 1. Preparation of 100uM stock solution

[1035] 1) Centrifuge the EP tube containing the oligonucleotide powder at 12000rpm for 2min;

[1036] 2) Add the corresponding volume of enzyme-free water to the dry powder according to the order feedback information and dilute it to 100uM stock solution.

[1037] 3) Take 1ul of the mother solution for nanodrop quantification and calculate the actual molar concentration. If it is <100uM or >130uM, subsequent electrophoresis and library quality inspection are required at the same time.

[1038] 4) Record A260 / 280 and A260 / 230. If A260 / 280 < 1.6, A260 / 280 > 2.2, subsequent electrophoresis and library quality inspection are required at the same time.

[1039] 2. Preparation of 10uM working solution

[1040] 1) Prepare the working solution according to the following table.

[1041] Table 16 Final concentration 10uM preparation system

[1042] Reagents 25ul system addition amount (ul) 50ul system addition amount (ul) Final concentration 4xT4DANLligasebuffer 6.25 12.5 1X indexF(100uM) 2.5 5 10uM indexR(100uM) 2.5 5 10uM T4PNK(10u / ul) 1.0 2.0 ddH2O 12.75 25.5 Total 25 50

[1043] 2) Set up the annealing program

[1044] Annealing procedure: 37℃ / 30min,

[1045] 95℃ / 2min、0.1℃ / s to 90℃,

[1046] 90℃ / 2min, 0.1℃ / s to 85℃,

[1047] ...,

[1048] 35℃ / 2min、0.1℃ / s to 30℃,

[1049] 30℃ / 2min, 0.1℃ / s to 25℃,

[1050] 25℃Hold.

[1051] 3) Annealing adapter quality inspection - 2% agarose gel electrophoresis detection

[1052] Reagents: 6×SuperStain Loading Buffer (CW2636); SuperDNAMarker (CW2583)

[1053] Sample volume 1ul, 6×SuperStain Loading Buffer volume 1ul, ddH2O volume 4ul, a total of 6ul, spot sample;

[1054] Marker 5ul, 6×SuperStain Loading Buffer 1ul, a total of 6ul, spot sample;

[1055] Qualified standard: If the connector annealing is qualified, there is only one main band. If there is a miscellaneous band, it is a failure and cannot be used. It is necessary to check the sample purity or re-anneal. Figure 3 .

[1056] 2. Target gene amplification

[1057] 1. Primer dilution and mixing

[1058] The 24 HLA primers were diluted to 100uM with low-TE (10mM Tris, 1mM EDTA, pH 8.0) and the following ratios were used: HLA-A_F_1:HLA-A_R_1:HLA-B_F_1:HLA-B_R_1:HLA-C_F_1:HLA-C_R_1:HLA-DPA_F_1:HLA-DPA_R_1:HLA-DPB_F_1:HLA-DPB_R_1:HLA-DQA_F_1:HLA-DQA_R_1:HLA-DQB_F_1:HLA-DQB_R_1:HLA-DRB1345_F_1:HLA-DRB1345_R_1:HLA-A_F_2:HLA-A_R_2:HLA-B_F_2:H LA-B_R_2:HLA-C_F_2:HLA-C_R_2:HLA-DPA_F_2:HLA-DPA_R_2:HLA-DPB_F_2:HLA-DPB_R_2:HLA-DQA_F_2:HLA-DQA_R_2:HLA-DQB_F_2:HLA-DQB_R_2:HLA-DRB1345_F_2:HLA-DRB1345_R_2=1:1:1:1:2:2::1:1:2:2:1:1:2:2:4:4:2:2:1:1:2:2:1:1:3:3:2:2:2:2:2:4 are mixed, and then diluted 5 times and PCR process is carried out according to the following amplification system

[1059] Amplification system

[1060] Table 17. Group 1 reaction system

[1061]

[1062]

[1063] Prepare the long-fragment amplification system in a PCR tube according to Table 3 on an ice box, add 30 ng of DNA template, pipette to mix, and centrifuge.

[1064] Amplification Procedure

[1065] Place the PCR tube into the PCR instrument and perform the reaction according to the conditions in Table 4.

[1066] Table 18. Long fragment amplification program

[1067]

[1068] 2. Purification of PCR Amplification Products

[1069] ① Preparation of reagents and consumables: Take out the Comwe Century DNA purification magnetic beads (Cat. No.: CW3171M) stored at 4°C and equilibrate at room temperature for about 30 minutes; prepare 80% alcohol at 440uL / reaction.

[1070] ②After the magnetic beads are balanced to room temperature, use a vortex shaker to mix the magnetic beads at medium speed for 1 minute. Add 12.5u1 (0.5X) magnetic bead suspension to the PCR tube. Mix by pipetting at half scale for about 10 times until the magnetic beads are mixed;

[1071] ③Incubate at room temperature for 5 minutes.

[1072] ④After incubation, place the PCR tube on the magnetic rack for about 3 minutes. When the solution becomes clear, discard the supernatant completely, taking care not to let the gun tip touch the magnetic beads.

[1073] ⑤ Keep the PCR tubes on the magnetic rack, add 200u1 of freshly prepared 80% alcohol to each tube, and let it stand at room temperature for about 30 seconds.

[1074] ⑥ When the solution becomes clear, remove the supernatant, taking care not to let the gun tip touch the magnetic beads.

[1075] ⑦ Repeat steps (5) and (6) once for a second wash.

[1076] ⑧ Keep the PCR tube fixed on the magnetic rack, open the tube cover, and dry at room temperature for 3 to 5 minutes until the magnetic beads are not reflective or cracked. ⑨ Remove the PCR tube from the magnetic rack, add 35uL of deionized water to each well, pipette to mix, incubate at room temperature for about 10 minutes, and centrifuge briefly.

[1077] ⑩ Place the PCR tube on the magnetic rack, wait until the solution becomes clear, transfer about 30uL of the purified amplification product to a new PCR tube, and be careful not to let the gun tip touch the magnetic beads.

[1078] The purified PCR product was used to quantify nucleic acid using qubit2.0 and placed on ice for later use.

[1079] 3. End Repair

[1080] 1. Prepare the final reaction system, see Table 4 for details.

[1081] Table 19 Preparation of end repair reaction solution

[1082] Components Volume (μl) - 1 reaction PCR product samples X 10×ERAT Buffer (CW3045) 2.5 ERATMix(CW3045) 1 Nuclease-free water 21.5-X Total volume 25

[1083] Place the PCR tube containing the reaction mixture from the previous step on a PCR instrument and react according to the following conditions (Table 5):

[1084] Table 20 End repair reaction conditions

[1085]

[1086] Four index connections

[1087] 1. Connection system configuration

[1088] Add the corresponding index to each sample and record it (Cwbar0001-1343), prepare the mixed system in a 0.2mL PCR tube, mix gently, and then centrifuge briefly to collect the reaction solution at the bottom of the tube, making sure there are no bubbles in the reaction solution. The reaction system is as follows (when preparing a mix or adding a single enzyme, the enzyme should be added last):

[1089] Table 21 Preparation of index ligation reaction solution

[1090] Components Volume (μl) - 1 reaction End-repaired DNA 14.25 Nativeindex 2.5 4XT4 DNA ligase Buffer (CW2701) 6.25 T4 DNA ligase (CW2701) 2 Total volume 25

[1091] Note: Samples and indexes need to correspond one to one. When adding indexes, contamination needs to be prevented.

[1092] Place the PCR tube containing the reaction mixture from the previous step on the PCR instrument and react according to the following conditions:

[1093] Table 22 Index ligation reaction conditions

[1094] temperature time Hot cover (off) 23℃ / Room temperature 15min

[1095] Note: After this step, the next step should be carried out immediately.

[1096] 2. Magnetic Bead Purification

[1097] 1) Take out the magnetic beads in advance, mix them well, and equilibrate them at room temperature for 30 minutes. According to the amount of subsequent mixed samples, take the corresponding volume of magnetic beads in advance and divide them into 8 strips for standby use (the magnetic beads can be divided into 8 strips according to the amount of experimental samples).

[1098] 2) Sample mixing: Take an equal amount of the reaction product from the previous step and add it to the eight pre-packed magnetic beads. Then mix the eight strips of 0.5X magnetic beads with samples into the same tube. At the same time, terminate the ligation reaction, mix well, and incubate at room temperature for 5 minutes.

[1099] 3) Take 1 mL of the mixed sample from the incubated magnetic beads into a new EP tube for purification. If the total volume is less than 1 mL, purify all of it. Transfer the EP tube to the magnetic rack and let it stand for about 2-5 minutes. When the liquid becomes clear, carefully discard the supernatant (be careful not to touch the magnetic beads).

[1100] 4) Keep the magnetic adsorption state, add 1mL 80% ethanol to the tube (cover the magnetic beads), do not blow the magnetic beads directly, after a while, carefully discard the supernatant (be careful not to touch the magnetic beads);

[1101] 5) Repeat the previous step (using 80% ethanol to clean the sample twice);

[1102] 6) Remove the EP tube, spin it off instantly, and carefully discard the remaining liquid with a pipette; open the test tube cover and dry the magnetic beads for about 3-5 minutes until there is no reflective surface on the magnetic beads (the surface of the magnetic beads should not be dry and cracked);

[1103] 7) Add 33 μl of nuclease-free water to elute the nucleic acid, and place in a constant temperature shaking metal bath, 300 rpm, 37°C and incubate for 5 min. ;

[1104] 8) Place the tube back on the magnetic rack until the magnetic beads are attracted to one side of the magnetic rack or the mixed solution becomes clear; transfer 32 μl of the supernatant to a clean PCR tube;

[1105] 9) Take 1 μL of the purified product for Qubit quantification.

[1106] 10) Take 300 ng of the purified product for the next step of connecting the sequencing adapter experiment.

[1107] 3. Sequencing adapter ligation

[1108] 3.1 System configuration

[1109] Prepare the mixed system in a 0.2mL PCR tube, mix gently, and then centrifuge briefly to collect the reaction solution at the bottom of the tube, making sure there are no bubbles in the reaction solution. The reaction system is as follows:

[1110] Table 23 Preparation of adapter ligation reaction solution

[1111] Components Volume (μl) Purified product from the previous step 30 Sequencing Adapter(NA) 2.5 4XT4 DNA ligase Buffer (CW2701) 12.5 T4 DNA ligase, 15 U / μL (CW2701) 5 EB Fill volume to 50

[1112] 3.2 Reaction conditions Place the PCR tube containing the reaction mixture from the previous step on a PCR instrument and react according to the following conditions:

[1113] Table 24 Connector ligation reaction solution conditions

[1114]

[1115]

[1116] 3.3 Magnetic bead purification

[1117] 1) Take out the magnetic beads in advance, mix them well, and equilibrate them at room temperature for 30 minutes. Add the sample to the magnetic beads (1.5mL centrifuge tube, 0.5X magnetic bead purification), gently mix by flicking or pipetting, and incubate at 37°C for 5 minutes.

[1118] 2) Transfer the centrifuge tube to the magnetic rack and let it stand for about 2-5 minutes. When the liquid becomes clear, carefully discard the supernatant (be careful not to touch the magnetic beads);

[1119] 3) Remove the EP tube from the magnetic stand, add 125 μl SFB (short fragment buffer (SFB), remove the centrifuge tube from the magnetic stand, flick to mix the magnetic beads, put it back on the magnetic stand, and after the liquid becomes clear, aspirate the clear liquid (be careful not to touch the magnetic beads);

[1120] 4) Repeat the previous step (use SFB for secondary cleaning and impurity removal);

[1121] 5) Remove the centrifuge tube from the magnetic rack, spin it up briefly, carefully discard the remaining liquid with a pipette, open the tube cap, and air dry for 30 seconds;

[1122] 6) Add 33 μl Elution Buffer (EB) to elute nucleic acid, flick to mix, and incubate at 37°C, 300 rpm for 8 min;

[1123] 7) Place the EP tube back on the magnetic stand until the magnetic beads are attracted to one side of the magnetic stand or the mixed solution becomes clear, then transfer 32 μl of the supernatant to a clean PCR tube.

[1124] 8) Take 1 μL of purified product for Qubit quantification (if the input amount is 300 ng, the Qubit concentration is ≥ 2.5 ng / ul).

[1125] 9) Take 20 ng of purified product for experiment.

[1126] Note: Do not clean the chip immediately after sequencing is completed. If the bioinformatics feedback indicates that the data is insufficient and unsuccessful, you can continue to click Start Sequencing to perform a supplementary test.

[1127] 3.4 Library quality control

[1128] After purification, the library was subjected to electrophoresis quality inspection. The results are shown in Figure 4 , the library size and distribution were consistent with the expected product size distribution, and the library continued to proceed with sequencing adapter ligation and upstream sequencing.

[1129] 5. Sequencing

[1130] Refer to the process of Example 1. The blood samples and reference samples tested in Example 2 are the same as those in Example 1, and the typing results are completely consistent.

[1131] The above are only embodiments of the present invention, and are not intended to limit the patent scope of the present invention. Any equivalent transformations made using the contents of the present invention specification, or directly or indirectly applied in other related technical fields, are also included in the patent protection scope of the present invention.

Claims

1. A primer set for HLA ultra-high resolution genotyping, the primer set being used for HLA gene multiplex amplification, characterized in that: The primer set includes primer pairs for HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3 / 4 / 5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 loci, and the upstream and downstream primers of the primer pair and the related nucleotide sequences are selected from any one or more of the following groups 1 to 48: The upstream and downstream primer sequences of the primer pairs for each site in Group 1 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO.1, the HLA-A-R1 sequence is shown in SEQ ID NO.2, and the HLA-A-R2 sequence is shown in SEQ ID NO.3; 2) The HLA-BF sequence is shown in SEQ ID NO.4, the HLA-B-R1 sequence is shown in SEQ ID NO.5, and the HLA-B-R2 sequence is shown in SEQ ID NO.6; 3) The HLA-CF sequence is shown in SEQ ID NO.7, the HLA-C-R1 sequence is shown in SEQ ID NO.8, and the HLA-C-R2 sequence is shown in SEQ ID NO.9; 4) The HLA-DPA-F1 sequence is shown in SEQ ID NO.10, the HLA-DPA-F2 sequence is shown in SEQ ID NO.11, and the HLA-DPA-R sequence is shown in SEQ ID NO.12; 5) The HLA-DPB-F sequence is shown in SEQ ID NO.13, the HLA-DPB-R1 sequence is shown in SEQ ID NO.14, and the HLA-DPB-R2 sequence is shown in SEQ ID NO.15; 6) The HLA-DQA-F sequence is shown in SEQ ID NO.16, the HLA-DQA-R1 sequence is shown in SEQ ID NO.17, and the HLA-DQA-R2 sequence is shown in SEQ ID NO.18; 7) The HLA-DQB-F sequence is shown in SEQ ID NO.19, the HLA-DQB-R1 sequence is shown in SEQ ID NO.20, and the HLA-DQB-R2 sequence is shown in SEQ ID NO.21; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO.22, the HLA-DRB1345-R1 sequence is shown in SEQ ID NO.23, and the HLA-DRB1345-R2 sequence is shown in SEQ ID NO.24; The upstream and downstream primer sequences of the primer pairs for each site in Group 2 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 25, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 26, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 27, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 28, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 29, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 30, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 31, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.32, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 3 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 33, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 34, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 35, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 36, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 37, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 38, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 39, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 40, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 4 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 41, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 42, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 43, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 44, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 45, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 46, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 47, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 48, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 5 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 49, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 50, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO.51, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 52, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 53, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 54, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO.55, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.56, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 6 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO.57, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 58, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 59, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 60, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO.61, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 62, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 63, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.64, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 7 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO.65, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 66, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 67, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 68, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 69, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 70, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 71, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.72, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 8 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 73, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 74, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 75, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 76, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 77, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 78, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 79, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 80, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 9 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO.81, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO.82, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 83, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 84, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 85, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 86, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 87, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.88, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 10 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO.89, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO.90, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO.91, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO.92, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 93, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 94, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO.95, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 96, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 11 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO.97, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO.98, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 99, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 100, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 101, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 102, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 103, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 104, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in Group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 12 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 105, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 106, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 107, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 108, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 109, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 110, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 111, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.112, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 13 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 113, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 114, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 115, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 116, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 117, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 118, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 119, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 120, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 14 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 121, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 122, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 123, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 124, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 125, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 126, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 127, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 128, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 15 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 129, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 130, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 131, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 132, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 133, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 134, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 135, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 136, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 16 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 137, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 138, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 139, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 140, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 141, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 142, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 143, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 144, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 17 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 145, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 146, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 147, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 148, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 149, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 150, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 151, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.152, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 18 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 153, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 154, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 155, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 156, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 157, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 158, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 159, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 160, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 19 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 161, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 162, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 163, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 164, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 165, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 166, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 167, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 168, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 20 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 169, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 170, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 171, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 172, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 173, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 174, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 175, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 176, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 21 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 177, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 178, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 179, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 180, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 181, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 182, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 183, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 184, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 22 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 185, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 186, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 187, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 188, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 189, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 190, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 191, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 192, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 23 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 193, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 194, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 195, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 196, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 197, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 198, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 199, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 200, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 24 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 201, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 202, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 203, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 204, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 205, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 206, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 207, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 208, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 25 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 209, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 210, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 211, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 212, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 213, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 214, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 215, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 216, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 26 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 217, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 218, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 219, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 220, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 221, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 222, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 223, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 224, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 27 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 225, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 226, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 227, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 228, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 229, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 230, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 231, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 232, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 28 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 233, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 234, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 235, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 236, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 237, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 238, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 239, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 240, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 29 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 241, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 242, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 243, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 244, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 245, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 246, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 247, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 248, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 30 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 249, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 250, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 251, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 252, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 253, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 254, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 255, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 256, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 31 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 257, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 258, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 259, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 260, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 261, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 262, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 263, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 264, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 32 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 265, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 266, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 267, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 268, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 269, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 270, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 271, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 272, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 33 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 273, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 274, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 275, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 276, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 277, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 278, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 279, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 280, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 34 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 281, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 282, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 283, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 284, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 285, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 286, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 287, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 288, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 35 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 289, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 290, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 291, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 292, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 293, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 294, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 295, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 296, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 36 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 297, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 298, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 299, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 300, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 301, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 302, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 303, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO. 304, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 37 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 305, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 306, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 307, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 308, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 309, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 310, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 311, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.312, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 38 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 313, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 314, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 315, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 316, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 317, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 318, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 319, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 320, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 39 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 321, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 322, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 323, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 324, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 325, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 326, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 327, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 328, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 40 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 329, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 330, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 331, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 332, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 333, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 334, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 335, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 336, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 41 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 337, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 338, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 339, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 340, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 341, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 342, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 343, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 344, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 42 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 345, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 346, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 347, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 348, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 349, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 350, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 351, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO.352, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 43 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 353, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 354, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 355, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 356, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 357, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 358, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 359, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 360, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 44 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 361, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 362, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 363, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 364, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 365, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 366, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 367, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 368, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in group 45 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 369, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 370, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 371, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 372, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 373, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 374, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 375, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 376, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 46 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 377, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 378, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 379, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 380, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 381, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 382, ​​and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 383, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 384, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 47 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 385, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 386, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 387, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 388, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 389, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 390, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 391, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The sequence of HLA-DRB1345-F is shown in SEQ ID NO. 392, and the sequences of HLA-DRB1345-R1 and HLA-DRB1345-R2 are the same as those in group 1; The upstream and downstream primer sequences of the primer pairs for each site in Group 48 are as follows: 1) The HLA-AF sequence is shown in SEQ ID NO. 393, and the HLA-A-R1 and HLA-A-R2 sequences are the same as those in group 1; 2) The HLA-BF sequence is shown in SEQ ID NO. 394, and the HLA-B-R1 and HLA-B-R2 sequences are the same as those in group 1; 3) The HLA-CF sequence is shown in SEQ ID NO. 395, and the HLA-C-R1 and HLA-C-R2 sequences are the same as those in group 1; 4) The HLA-DPA-R sequence is shown in SEQ ID NO. 396, and the HLA-DPA-F1 and HLA-DPA-F2 sequences are the same as those in group 1; 5) The HLA-DPB-F sequence is shown in SEQ ID NO. 397, and the HLA-DPB-R1 and HLA-DPB-R2 sequences are the same as those in group 1; 6) The HLA-DQA-F sequence is shown in SEQ ID NO. 398, and the HLA-DQA-R1 and HLA-DQA-R2 sequences are the same as those in group 1; 7) The HLA-DQB-F sequence is shown in SEQ ID NO. 399, and the HLA-DQB-R1 and HLA-DQB-R2 sequences are the same as those in group 1; 8) The HLA-DRB1345-F sequence is shown in SEQ ID NO.400, and the HLA-DRB1345-R1 and HLA-DRB1345-R2 sequences are the same as those in Group 1.

2. A primer set for HLA ultra-high resolution genotyping, the primer set being used for HLA gene multiplex amplification, characterized in that: The primer set includes primer pairs for HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3 / 4 / 5, HLA-DQA1, HLA-DQB1, HLA-DPA1 and HLA-DPB1 loci. The loci can be set into two sets of upstream and downstream primers. The upstream and downstream primers and related nucleotide sequences are shown in the following table:

3. Use of the primer set as claimed in claim 1 in preparing an HLA ultra-high resolution horizontal genotyping detection kit.

4. An HLA ultra-high resolution genotyping detection kit, characterized in that: The detection kit comprises the primer set and detection reagent as claimed in claim 1.

5. The detection kit according to claim 4, characterized in that The detection reagent includes the HLA gene rapid amplification enzyme system Flash PCR Master Mix For Long Fragment.

6. Use of the primer set as claimed in claim 2 in preparing an HLA ultra-high resolution horizontal genotyping detection kit.

7. An HLA ultra-high resolution genotyping detection kit, characterized in that: The detection kit comprises the primer set and detection reagent as claimed in claim 1.

8. The detection kit according to claim 7, characterized in that The detection reagent includes HLA gene rapid amplification enzyme system Flash PCR Master Mix For Long Fragment and Index synthesis related oligonucleotide sequences; The Index synthesis-related oligonucleotide sequence is selected from any one or more groups of Index01-Index48. The upstream and downstream primers of the primer pairs of Index01-Index48 and the related nucleotide sequences are shown in the following table:

9. A full-length and rapid sequencing method for HLA typing based on nanopore sequencing, characterized in that: The method uses any one of the primer sets described in claims 1-2 or any one of the kits described in claims 3-8 to amplify the HLA gene in the sample to be tested, and then sequences the amplified product.

10. A method for HLA genotyping for non-diagnostic and non-therapeutic purposes, characterized in that: The method comprises amplifying the HLA gene in the sample to be tested using any one of the primer sets described in claims 1-2 or any one of the kits described in claims 3-8, constructing the amplified products into a sequencing library, and then sequencing and performing bioinformatics analysis on the sequencing data to obtain full-length ultra-high-resolution typing information including 11 HLA genes in the sample to be tested.

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