Now NGS technology is increasingly used in HLA genotyping forhigh resolution[18–27],it has high throughput and can bedecreased the rate of the ambiguous allele combinations. Here weused a commercial NGS method for HLA genotyping. However, someambiguities typing was not discriminated in the method, includingHLA-A*02:06:01:01,02:07:01/A*02:01:01:01,02:474;A*02:03:01,02:06:01:01/A*02:01:01:01,02:355;B*35:01:23,46:01:01/B*35:67,46:33; B*35:01:01:05,51:01:01:01/B*53:01:13,B*78:02:01; B*51:01:01:01,55:02:01/B*51:157,56:05:02;C*05:01:01:02,08:01:01/C*08:02:01:02,08:10,DQB1*06:02:01:01,06:09:01:01/DQB1*06:84,06:88 ambiguous allele combinations, andDRB1*15:02:01G (DRB1*15:02:01/15:140/ 15:149),DRB1*09:01:02/09:31,DRB1*04:07:01/04:92, DRB1*15:11/15:15:01,DQB1*02:02:01:01/ 02:29,DQB1*03:01:01:12/03:01:41/03:276N, DQB1*04:01:01/04:05DQB1*03:01:01:12/03:01:41,DQB1*06:01:01/06:01:15 and DQB1*05:01:01:03/05:01:24. All of them was finally assigned according toguideline of common alleles and well documented alleles (CWD) inChina ,butHLA-DRB1*12:01:01G (DRB1*12:01:01/12:10) was not discriminated.This phenomenon may be due to no detection for the sequences ofexon 1 and intron 1 of HLA-DRB1, -DRB3/4/5, -DQB1, -DPB1 loci orphase break leading to two variants could not be phased. Sequencingfor the full length of HLA loci and extend read length each timemay be solved this ambiguities typing. In additional, the resultsof two samples have errors by the NGS method. TheHLA-DRB1*12:02:01,16:09:01 of a subject was confirmed to be asDRB*12:02:01,15:01:01 by PCR sequence-based typing, andB*40:130:02,48:01:01 of NGS was confirm to be B*40:01:02,48:04:01.HLA-DRB1*16:09:01 were well document allele and HLA-B*40:130:02 wasrare alleles in the Chinese population, therefore it was suggestedthat this NGS method need to improve and some low/rare frequencyallele combination need to be checked using other technology.
The study of HLA allele and haplotype frequencies in humanpopulations provides an important source of information foranthropological investigation, organ and haematopoietic stem celltransplantation purposes as well as disease associationstudies[1–2]. The distribution of HLA alleles andhaplotypes is ethnically and geographically restricted. Now manystudies have been reported the data of HLA alleles and haplotypesat low resolution or two fields resolution, but the data for HLAalleles and haplotypes at three fields resolution was rare in theChinese populations. In this study, the distribution of HLA-A, -B,-C, -DRB1, -DQB1 and -DRB3/4/5 loci in Zhejiang Han population atthree fields resolution level was reported. Our data showed thatthe characteristic of HLA allele in the Zhejiang Han population wasin close proximity to that of the Chinese southern Han populationat low and two fields resolution level, and consistent with ourprevious data[13–15]. However, comparison with the data at twofields resolution in our previous report, some HLA alleles werefurther divided. For example, previously HLA-A*11:01 can be dividedinto A*11:01:01(25.8%) and A*11:01:04(0.03%), and HLA-A*24:02 wascomposed of A*24:02:01(16.7%), A*24:02:31(0.03%) andA*24:02:40(0.03%) at three fields resolution level. Similarly,HLA-C*07:02 can be further divided into C*07:02:01(19.03%) andC*07:02:06 (0.03%).
Estimation of HLA haplotype in different ethnic groups isextremely useful for searching HLA-matched donors in unrelatedhaematopoietic stem cell transplantation. The most prevalenthaplotypes in the study were generally consistent with our previousreport. TheHLA-DRB3, -DRB4, -DRB5 allele is associated with HLA-DRB1 allele,now the AFs data of HLA-DRB3, -DRB4, -DRB5 was rare at highresolution level in the Chinese population.HLA-DRB4*01:03:01(25.72%) was the common alleles for HLA-DRB4 locusin our study. However, HLA-DRB4*01:01 (34.93%) and DRB4*01:03:01(34.03%) were the most common in the nativeAmericans.For the HLA-DRB3 locus, the DRB3*02:02:01 was common in theZhejiang Han population, but DRB3*01:01 was common in the nativeAmericans. The data of our study was supported that the view of thedistribution of HLA alleles was various in the differentethnicity.
In conclusion, we firstly reported the characteristics of HLAallele and haplotype of HLA-A, -B, -C, -DRB1, -DQB1 and -DRB3/4/5loci at three fields resolution level in the Zhejiang province ofChina. The data may help to searching unrelated bone marrow donorsfor patients. Furthermore, our study revealed that the technologyof NGS for HLA typing was high-throughput and precise, mostlyalleles can be genotyped at three fields resolution with fewerambiguous typing.