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目的 构建含HIV 1中国流行株B亚型核心蛋白Gag基因的真核表达质粒 ,并在体外进行表达和鉴定。方法 将Gag基因插入到核酸疫苗载体质粒pVAX1中 ,构建真核表达质粒 pVAXGAG。用脂质体法 ,将重组质粒转染入Hela细胞后进行表达产物的检测。结果间接免疫荧光检测显示 ,转染重组质粒的细胞表面有绿色荧光。Westernblot和DotELISA分析显示 ,重组质粒转染细胞的裂解物中存在表达的Gag蛋白。 结论构建的核酸疫苗可在体外进行表达 ,且表达的蛋白具有特异性
Objective To construct an eukaryotic expression plasmid containing the Gag gene of subtype B of HIV-1 Chinese strain and to express and identify it in vitro. Methods The Gag gene was inserted into the DNA vaccine plasmid pVAX1 to construct the eukaryotic expression plasmid pVAXGAG. The recombinant plasmids were transfected into Hela cells by lipofectamine and the expression products were detected. Results Indirect immunofluorescence assay showed that the surface of transfected plasmids had green fluorescence. Western blot and Dot ELISA analysis revealed the presence of expressed Gag protein in the lysates of the recombinant plasmids transfected cells. Conclusion The constructed DNA vaccine can be expressed in vitro and the expressed protein is specific