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目的扩增广西HIV_1感染者的HIV_1膜蛋白基因C2_V3 区核酸片段并提纯回收 ,进行核苷酸序列测定与亚型分析。方法用淋巴细胞分离液从HIV_1感染者的静脉血中分离PBMCs,然后用多组不同的内外侧引物进行Nested_PCR扩增HIV_1膜蛋白基因C2_V3 区核酸片段 ,后用琼脂糖电泳进行提纯 ,用荧光标记末端终止物循环测序试剂盒进行测序 ,反应物用自动DNA序列分析仪进行序列测定和分析。结果经琼脂糖凝胶电泳检测 ,获得26份DNA阳性产物 ,阳性率为86.67%。不同引物的阳性率相差较大 ,根据扩增结果可以推断广西HIV_1流行毒株的基因变异程度。14份HIV_1核苷酸序列测定与亚型分析结果 ,9份为B亚型 ,5份为E亚型。结论广西存在E和B亚型HIV_1毒株的流行。
OBJECTIVE: To amplify and recover the HIV-1 membrane protein gene C2_V3 region of HIV-1 infected persons in Guangxi by nucleotide sequencing and subtype analysis. Methods PBMCs were isolated from the venous blood of HIV-1 infected patients by lymphocyte separation. Nucleic acid fragments of HIV_1 membrane protein gene C2_V3 were amplified by Nested_PCR using multiple sets of internal and external primers. After purification by agarose electrophoresis, End Terminator Cycle Sequencing Kit was sequenced and the reagents were sequenced and analyzed using an automated DNA sequencer. Results After agarose gel electrophoresis, 26 DNA positive products were obtained, the positive rate was 86.67%. The positive rates of different primers are quite different. Based on the amplification results, it can be deduced that the degree of genetic variation in HIV-1-endemic strains in Guangxi is different. 14 HIV_1 nucleotide sequencing and subtype analysis results, 9 were B subtype, 5 were E subtype. Conclusion There is a prevalence of HIV-1 strains of E and B subtypes in Guangxi.