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[目的]克隆山东省流行的HIV-1优势亚型的代表毒株的包膜蛋白基因,建立用于山东省HIV-1分型的异源双链泳动分析体系。[方法]2003年11~12月将与山东省HIV-1 B′/C、B′和A/E各亚型共享序列最接近的毒株用套式聚合酶链反应(PCR)技术扩增包膜(env)基因,克隆到p GEM-Teasy载体中,构建成用于异源双链泳动分析(HMA)分型的山东标准亚型质粒,并摸索各反应和电泳的最佳条件。[结果]共建立HMA的分型标准质粒11株,建立以标准质粒为分型标准、包含3套可供选择的片段扩增策略的HMA体系,与序列分析比较各亚型代表株的检出率(敏感性)分别达到了B′94%、B/′C 90%、A/E 90%。[结论]建立的HMA分型体系经济、方便、易掌握、结果可靠,值得推广。
[Objective] To clone the envelope protein gene of the representative strains of the HIV-1 dominant subtype in Shandong Province and establish a heteroduplexation kinetic assay system for HIV-1 typing in Shandong Province. [Method] From November to December in 2003, the closest strains sharing the sequences of HIV-1 B ’/ C, B’ and A / E in Shandong Province were amplified by nested polymerase chain reaction (PCR) The envelope (env) gene was cloned into p GEM-Teasy vector and constructed into Shandong standard subtype plasmids for heterotypic HMA typing. The optimum conditions for each reaction and electrophoresis were also explored. [Result] A total of 11 HMA genotyping plasmids were established. HMA system based on the standard plasmids was established and included 3 sets of alternative fragment amplification strategies. Rates (sensitivity) reached B ’94%, B / C 90% and A / E 90%, respectively. [Conclusion] The established HMA typing system is economical, convenient and easy to grasp, with reliable results and worthy of promotion.