丙型肝炎病毒非结构区NS3基因在大肠埃希菌中的可诱导性表达  

Inducible expression of non-structural protein 3 of hepatitis C virus in E. coli

作　　者：成军[1] 钟彦伟[1] 刘妍[1] 董菁[1] 杨继珍[1] 杨守纯[1] 

机构地区：[1]解放军第三○二医院传染病研究所基因治疗研究中心,北京100039

出　　处：《中华实验和临床病毒学杂志》2002年第1期85-87,共3页Chinese Journal of Experimental and Clinical Virology

摘　　要：目的　构建丙型肝炎病毒 (HCV)NS3基因的原核细胞表达载体 ,实现在大肠埃希菌中的可诱导性表达。方法　应用聚合酶链反应 (PCR)技术 ,以美国HCV H株全长cDNA质粒为模板 ,扩增获得NS3基因片段 ,克隆到原核表达载体pET 30C(+)中 ,构建原核表达载体pET NS3,转化BL2 1(DE3)宿主菌 ,以IPTG诱导 ,获得NS3蛋白的可诱导性表达 ,以HCVNS3的单链可变区抗体 (ScFv)证实表达的NS3蛋白的特异性。结果　以HCVNS3基因序列特异性引物 ,PCR扩增获得 1893bp的NS3DNA片段 ,插入pET 30C(+)表达载体。转化BL 2 1(DE3)受体菌 ,经培养、IPTG诱导 ,获得了重组HCVNS3蛋白的表达 ,以HCVNS3的ScFv证实了表达的重组蛋白HCVNS3的特异性。Objective To express recombinant non-structural protein 3 of hepatitis C virus (HCV) in E. coli. Methods The non-structural 3 (NS3) region DNA fragment of HCV was amplified by polymerase chain reaction (PCR) and inserted into inducible proeukaryotic expressive vector pET-30C(+)at Bam H Ⅰ/EcoR Ⅰ sites. The competent BL21 (DE3) E.coli was transformed, and then cultured and induced with IPTG. The expressed HCV NS3 protein was confirmed with ELISA and dot blot hybridization using HCV NS3-specific single chain Fv (ScFv) antibody. Results 1 893 bp DNA fragment of HCV NS3 coding region was amplified by PCR technique. HCV NS3 expressive vector pET-NS3 was constructed. After transformation with pET-NS3 and induction with IPTG, recombinant HCV NS3 protein was expressed and confirmed by specific ELISA and dot blot hybridization. Conclusion The recombinant HCV NS3 can be expressed in E. coli.

关 键 词：丙型肝炎病毒 NS3 基因表达 聚合酶链反应 大肠埃希菌 

分 类 号：R373[医药卫生—病原生物学]