应用PCR-SSOP技术研究中国汉族、维吾尔族HLA-A基因座基因多态性

人类白细胞抗原（Human Leukocyte Antigen,HLA)基因复合物位于6p21.3，有220多个不同的功能基因，是人类基因组最复杂的遗传多态系统。HLA等位基因的变异在医学、法医学、人类学等领域具有重要的意义。自从1964年以来，HLA分型一直采用经典的微量淋巴细胞毒实验，但该方法是血清学水平的分，不能识别很多特异性的等位基因，而且高质量的抗体也不易获得。从20世纪90年代起，在国家自然科学基金的资助下，首先开展HLAⅡ类位点基因分研究及大规模群体多态性调查，所获得的中国主要民族基因数据已应用于多个领域。相比之下，HLAⅠ类基因数量更丰富，包含了A、B、C、E、F、G和假基因H、J、K、L等10个位点；基因分子结构更复杂，更具多态性。因此，HLAⅠ类DNA分型比HLAⅡ类分型及行多困难。直至目前中国人群HLA－A基因座基因多态性和分布频率的研究尚未充分进行。而任何DNA标记用于遗传分析、法医鉴定等领域之前，必须先进行群体调查，建立不同民族基因数据库，这是不可逾越的基础工作。鉴于此，采用灵敏而非同位素污染的PCR－SSOP基因分型技术，对165个汉族和162个维吾尔族个体的HLA－A基因座多态性进行调查。结果在汉族群体中发现22种等位基因，频率最高的是HLA－A＊1101（19．7％），其次是＊201（12．72％）；在维族群体中发现22种等位基因，频率最高的是＊2407（17．90％），等位基因＊0101、＊0201和＊3301的频率均大于10％；HLA－A＊0203、＊0205、＊0302、＊2403和＊3302仅在汉族群体中检出；HLA－A＊0205、＊0211、＊2301、＊2502、＊68012和＊6802仅在维族群体中检出。按照Hardy-Weinberg平衡定律检验，两个民族各等位基因型频率的预期值与实际观察值相吻合（P＞0．05），证明了所获得汉族、维吾尔族HLA－A位点基因频率具有可靠性；同时也表明各等位基因的遗传特征符合符合孟德尔规律。经计算机统计分析，汉族群体HLA－A基因座杂合度（Heterozygosity,H)、个体识别率（Discrimination Power,DP)和非父排排率（Proba-bility of Paternity Exclusion,EPP)分别为0．9029、0．9776和0．8592；维族群体H、DP和EEP分别为0．9063、0．9379和0．7885。和其他遗传标记（如VNTR、STR、SNP）的单一位点相比，HLA－A具有高度的杂合率、个体识别率和非父排除率。因此，HLA－A等位基因在法医个体识别、亲权鉴定、基因诊断、人类学等领域具有重要的应用价值。

Analysis of DNA Polymorphism at HLA-A Locus by PCR Amplification with Sequence Specific Oligonucleotide Probe in Chinese Han and Uygur Populations

To determine HLA A genetic polymorphism in Chinese populations and establish ethnic genetic database, 165 Han and 162 Uygur subjects were investigated with a non isotopic and sensitive method PCR SSOP. 22 alleles were identified in Han with the most frequent allele being HLA A*1101(19.7%), followed by *0201(12.72%). Also, 22 alleles were identified in Uygur with *2407(17.90%) being the most frequent one and the frequencies of following alleles: *0201,*0101,*3301were higher than 10%. HLA A *0203,*0205,*0210,*0302,*2403 and *3302 were only detected in Han; meanwhile *0205 ?*0211?*2301?*2502?*68012 and *6802 were only in Uygur. According to Hardy Weinberg equilibrium , each allele showed no significant ( P >0.05) deviation between the expected frequency and the observed one. Heterozygosity ( H ), discrimination power ( DP ) and probability of paternity exclusion ( EPP ) of HLA A locus from Han nationality were computed to be 0.9029,0.9776 and 0.8592;and those from Uygur as 0.9063 , 0.9379 and 0.7885. These results suggest that HLA A DNA polymorphism and the database of two Chinese populations have useful applications in processing forensic casework (as personal identification, paternity test), tracing population migration and genetic diagnosis.