HIV-1基因亚型分型:　异源双链泳动分析法

Genetic Subtyping of HIV 1 Strains by Heteroduplex Mobility Assay

应用套式ＰＣＲ（ｎｅｓｔｅｄＰＣＲ）对２４份ＨＩＶ感染者外周血单核细胞（ＰＢＭＣ）样品进行扩增，从样品中获得了ＨＩＶ１膜蛋白（ｅｎｖ）基因的核酸片段，然后与已知各标准亚型的对照质粒的相同片段ＰＣＲ产物进行杂交，使样品ＤＮＡ与质粒ＤＮＡ同源部位配对形成异源双链体，根据其在聚丙烯酰胺凝胶电泳中的泳动率可确定其亚型。并将这种异源双链泳动分析法（ｈｅｔｅｒｏｄｕｐｌｅｘｍｏｂｉｌｉｔｙａｓａｙ，ＨＭＡ）的结果与ＤＮＡ序列测定结果比较。实验证明，ＨＭＡ是一种快速、简便。

DNA fragments of HIV 1 env gene were amplified by nested PCR from uncultured peripheral blood mononuclear cells(PBMCs) obtained from 24 HIV 1 infected individuals. The PCR products were separated by melting and annealing with denatured PCR product prepared from reference plasimd of representative subtypes. Heteroduplex were then formed between the single stranded DNA from the two sources and were analysed on polyacrylamide gels. The results from heteroduplex mobility assay(HMA) were compared with HIV 1 subtype results determined by DNA sequencing. With advantages of high speed,low cost and high specificity,HMA is a reliable screening method for HIV 1 subtyping.