中国南方汉族人群HLA-DPA1和-DPB1基因多态性及其常见和确定的等位基因研究

A Study on Polymorphisms and Common and Well-Documented Alleles of HLA-DPA1 and-DPB1 in Southern Chinese Han Population

目的 研究中国南方汉族人群人类白细胞抗原（HLA）基因HLA-DPA1和-DPB1的多态性及常见和确定的（CWD）等位基因.方法 计算机随机数表法选择2010年2月至2011年9月,于深圳市血液中心采集的1 730份汉族健康志愿献血者全血样品为研究对象.采用HLA-DPA1及-DPB1基因第2外显子同步测序分型方法对血液样品的HLA-DPA1和-DPB1基因进行测序分型,对模棱两可的分型结果采用组特异性引物测序法鉴定.测序反应产物经乙醇醋酸钠/乙二胺四乙酸方法纯化后,采用ABI 3730测序仪进行测序,测序结果导入Assign 3.5测序分型软件,判定HLA-DPA1和DPB1等位基因型.结果 本组1 730例中国南方汉族无关个体中,共检出7种HLA-DPA1等位基因,其中基因频率＞0.1％的5种常见型等位基因依次为DPA1＊ 02∶ 02（54.65％）、DPA1＊ 01∶ 03（31.24％）、DPA1＊ 02∶ 01（10.26％）、DPA1 ＊04∶ 01（3.64％）及DPA1＊ 01∶04（0.12％）;另2种为罕见型的新等位基因,其检出次数均＜3次.共检出了33种HLA-DPB1等位基因,其中20种为常见型等位基因,其基因频率均＞0.1％;2种为确定型等位基因,其基因频率＜0.1％,但检出次数≥3次;其余11种为罕见型等位基因,其中含2个新等位基因.结论 本研究建立了HLA-DPA1和-DPB1基因第2外显子双向测序及组特异性引物鉴定模棱两可分型结果的分型技术,初步获得了中国南方汉族人群HLA-DPA1、-DPB1基因遗传多态性及CWD等位基因资料.

Objective To explore the polymorphisms of human leukocyte antigen （HLA）-DPA1 and-DPB1,and determine the common and well-documented （CWD） alleles in southern Chinese Han population.Methods From February 2010 to September 2011,a total of 1 730 cases of peripheral blood samples from unrelated healthy voluntary blood donors of Chinese Han population in Shenzhen blood center,were collected in this study by compater random number table.All these samples were subjected to HLA-DPA1 and-DPB1 sequencing-based typing at exon 2 in both directions.In-house group specific primers were designed and utilized to identify the ambiguous allele combinations.Products of sequencing reactions subjected to sequence cleaning by EtOH/ethylene diamine tetraacetic acid （EDTA） method.The purified products were run electrophoresis on the ABI 3730 DNA sequencer,then obtained sequences were imported into the Assign 3.5 software,and the HLA-DPA1 and-DPB1 allelic genotype were assigned.Results In 1 730 unrelated healthy southern Chinese Han individuals,a total of 7 HLA-DPA1 alleles were identified,including 5 common alleles of which the frequencies were more than 0.1%.Frequencies of each common allele were 54.65% of DPA1 ＊ 02 ∶ 02,31.24% of ＊ 01 ∶ 03,10.26% of ＊ 02 ∶ 01,3.64% of ＊ 04 ∶ 01,and 0.12% of ＊ 01 ∶ 04.Two rare novel alleles were observed in less than 3 times.Thirty three HLA-DPB1 alleles including 20 common alleles with a frequency more than 0.1%,and 2 well-documented alleles with a frequency less than 0.1% but were detected more than 3 times and 11 rare alleles of which two were novel alleles were determined.Conclusions Technique for simultaneous HLA-DPA1 and-DPB1 sequencebased typing at exon 2 and assay for determining ambiguous allele combinations using the in-house group specific primers have been established in this study.The information of allele frequencies and CWD alleles for HLA-DPA1 and-DPB1 in southern Chinese Han population were obtained.