丙型肝炎病毒核心蛋白基因的克隆及表达

Cloning and expression of hepatitis C core protein gene

目的在大肠杆菌中表达丙型肝炎病毒(HCV)核心蛋白基因片段。方法以HCV全长cDNA质粒为模板进行PCR扩增,获得366bp的核心蛋白基因片段。将其插入连接载体pMD18-T vector中,酶切后再与表达载体pBV220连接,构建重组表达载体pBV220/HCV-C。经鉴定后温度诱导表达,采用SDS-PAGE电泳及Western-blot检测核心蛋白基因的蛋白表达。结果经温度诱导后获得目的基因的非融合表达,SDS-PAGE电泳显示在14000 u处有一表达条带;Western-blot结果显示其具有HCV抗原特异性。结论丙型肝炎病毒核心蛋白基因成功表达,对临床诊断试剂和HCV基因疫苗的研制有一定的意义。

Objective To clone the fragment of hepatitis C virus （HCV） core gene and express it in E. coli. Methods The fragment of HCV core gene （approximate 366bp） was amplified by PCR and inserted into the pMD18-T vector. The cloned HCV core gene, which was confirmed by the digestion with EcoR I/Bam H I , was subcloned into the expression vector pBV220 to construct recombinant plasmid pBV220/HCV-C. The expressed gene product was identified by SDS-PAGE and Western blotting. Results The fragment of HCV core gene was expressed successfully after temperature induction and a protein of 14 000 u was resulted. Conclusion Expression of the HCV-C gene in E. coli was achieved, which may be helpful for further studies on characterizations of HCV-C gene.