异基因骨髓移植中PCR-SSP和RFLP对HLA-DRB、DPB1分型的应用

Application of HLA-DRB、DPB1 genotyping by PCR-SSP、RFLP for allogeneic bone marrow transplantation

目的:探讨序列特异性引物聚合酶链反应（PCR-SSP）和聚合酶链反应限制性片段长度多态性分析（PCR-RFLP）对HLA-DRB、DPB1分型在异基因骨髓移植中应用的可行性。方法:应用19对不同的DRB抗原所对应等位基因序列特异性的引物,PCR扩增供受者各20个样本DRB,产物直接琼脂糖凝胶电泳分析;同时用另一对引物PCR扩增DPB1第二外显子,产物10种限制性内切酶酶切分析。结果:各DRB带清晰可辨,直接确定为该型;DPB1的多态性片段编码转换后确定其基因型;4例找到供者。结论:它们是异基因骨髓植前快速、精确和可靠的基因分型手段。

Objective:To investigate the feasibility of clinical application of HLA-DRB,DPB1 genotyping by PCR-SSP,RFLP for alloge-neic bone marrow transplantation(Allo-BMT) .Methods: Corresponding to each of the DRB alleles, nineteen different pairs of sequence specific primers were used to PCR amplify them from twenty samples in both recipients and donors and their amplified products were directly analyzed on agarose gel. At the same time, a pair of primer was used to PCR amplify the second exon of DPB1 and the restriction fragment length polymorphisms were analyzed on PAGE after their PCR products were separately digested by ten endonucleases. Results: Each DRB was typed in according to its clear and visualized electrophoresis band; the DPB1 genotype was also determined by analysis of all codes representing polymorphism fragment band with a computer. Four couples of recipient and donor were matched. Conclusion:PCR-SSP and RFLP are rapid, accurate and reliable genotyping approaches for Allo-BMT.