丙型肝炎病毒NS3基因真核表达载体的构建及表达  

Construction of eukaryotic plasmid including HCV NS3 gene and expression of NS3 protein in cells of HeLa

作　　者：孙菊[1] 楚雍烈[2] 安润[2] 付慧[1] 姜凤良[1] 柴长斌[1] 胡志芳[1] 

机构地区：[1]西安医学院病原生物与免疫学教研室,西安710021 [2]西安交通大学医学院

出　　处：《陕西医学杂志》2009年第7期799-801,共3页Shaanxi Medical Journal

摘　　要：目的:构建丙型肝炎病毒(HCV)非结构基因NS3的真核表达载体,并分析其在HeLa细胞中的表达。方法:以HCV全长cDNA质粒为模板进行PCR扩增,获得全长NS3基因,将其插入克隆载体pMD18-T中,鉴定后与真核表达载体pcDNA3.1(+)重组,用脂质体介导法转染HeLa细胞,间接免疫荧光法及Western blot鉴定转染后HCVNS3蛋白的表达。结果:PCR扩增的NS3序列正确,构建的真核表达载体成功转染HeLa细胞,表达的NS3蛋白相对分子量为70kD。结论:成功构建了真核表达载体pcDNA3.1(+)/NS3,并在HeLa细胞中获得表达。Objects： To construct eukaryotic plasmid which includes HCV NS3 gene , and induce the expression of NS3 protein in cells of HeLa. Methods ： HCV NS3 gene was amplified with PCR and inserted into the pMD 18-T vector. We recombined the NS3 segment with expression plasmid pcDNA3.1（＋） to construct eukaryotic plasrfiid pcDNA3. 1（＋） / NS3. Then the plasmid pcDNA3, 1（＋） / NS3 was transfected into cells of HeLa with Lipofectamine 2000 transfection reagent. The expression of NS3 protein was detected by Immunofluorescent staining method and Western Blot. Results ： The cloned NS3 segment had correct length and sequence. The cells of HeLa were successfully transfected and expressed NS3 protein. Conclusion ： Eukaryotic expression plasmid pcDNA3, 1（＋） /NS3 including HCV NS3 gene is successfully constructed and expressed in cells of HeLa.

关 键 词：肝炎病毒/免疫学 @非结构蛋白3 @Hela细胞 基因表达 

分 类 号：R382.31[医药卫生—医学寄生虫学] R373.21[医药卫生—基础医学]