HLA－DRB1基因位点多态性的PCR－RFLP分析

设计并建立一套适合国内应用的改良ＰＣＲ－ＲＦＬＲ方法，分５组特异性扩增ＤＮＡ样品，随后进行酶切定型分析，准确检测了编码ＤＲ抗原特异性的ＨＬＡ－ＤＲＢ１基因位点的多态性，该法采用分组扩增，不发生与其它ＤＲＢ位点等位基因的交叉扩增，不仅适合纯合子的区分而且可以清楚准确地检测杂合子样品，已报道过的ＤＲＢ１位点编码的特异性组合都可以通过这个方法得到准确分析。所使用的Ⅱ类限制性内切酶均价格便宜、易购。

Analysis of the Polymorphism at HLA-DRB1 Gene Locus by PCR-RFLP

A modified PCR-RFLP method was designed and established to analyse the polymorphism of HLA-DRB1 gene locus that codes the HLA-DR antigenic specificities. This technique involves two steps:1) Amplification of the second exons of DRB1 gene with five pairs of group-specific primers. In this step, DRB1 04 can be determined. 2) Endonucleases digest the amplified DNA to assign the genetic types of the other specificities (DRB1 01, DRB1 15, DRB1 16,DRB1 03, DRB1 11, DRB1 12, DRB1 13, DRB1 14, DRB1 07, DRB1 08, DRB1 09,DRB110) and four alleles(DRB1 0301, DRB1 1303, DRB1 1403, DRB1 08031). In the modified PCR-RFLP analysis, five pairs of group-specific primers have been chosen to avoid cross-amplification with other DRB alleles. Endonucleases used in this study are rather common and cheap.Particularly, this method can assign HLA-DRB1 genetic specificities in hetetozygous as well as homozygous individuals unequivocally. Individuals carrying any combinations of the HLA-DR antigenic specificities published so far can be typed by this method.