HIV-1表面糖蛋白gp120 DNA疫苗质粒的构建与表达

目的 构建含HIV-1表面糖蛋白基因的真核表达质粒,并在Hela细胞内表达。方法 在核酸疫苗载体质粒 pVAX1中插入gp120基因,构建真核表达质粒pVAXGP。在体外用脂质体法将重组质粒转染Hela细胞,并用间接免疫荧光试验、免疫印迹试验和Dot-ELISA对表达产物进行检测。结果 间接免疫荧光试验结果显示,转染重组质粒的细胞表面有绿色荧光。免疫印迹试验和Dot-ELISA结果均显示,重组质粒转染细胞的裂解物中存在表达的gp120蛋白。结论 已成功地构建了真核表达质粒,且表达的蛋白具有良好的反应原性和特异性。

Construction of Recombinant Plasmid and Expression of Surface Glycoprotein gp120 of HIV-1

Objective To construct an eukaryolic expression plasmid and express surface glycoprotein gp120 gene of HIV-1 in Hela cell. Methods An eukaryotic expression plasmid pVAXGP was constructed by inserting gp120 gene into DNA vaccine vector plasmid pVAX1, and transfected to Hela cell in vitro by lipo-some-mediated method. The expressed product was detected by indirect immunofluorescence assay(IFA), Western blot and Dot-ELISA. Results Indirect IFA showed green fluorescence on the surface of transfected cells.Both Western blot and Dot-ELISA proved that the lysate of transfected cells contained gp120 protein, Conclusion An eukaryotic expression plasmid for gp120 gene of HIV-1 was successfully constructed, and the expressed product showed good reactogenicity and specificity.