一个包含HCVIRES的高效真核双顺反子表达载体的构建  

Construction of an effective eukaryotic bi-cistronic expression vector containg HCV internal ribosome entry site

作　　者：欧山海[1] 吴婷[1] 何水珍[1] 伍小路[1] 郑舟[1] 张军[1] 夏宁邵[1] 

机构地区：[1]厦门大学福建省医学分子病毒学研究中心,厦门361005

出　　处：《中国人兽共患病杂志》2005年第9期783-787,832,共6页Chinese Journal of Zoonoses

基　　金：福建省科技重大专项(2004YZ01-01)

摘　　要：In this paper, a new eukaryotic bi-cistronic expression vector containing Hepatitis C Virus(HCV) internal ribosome entry site (IRES) expressing two foreign genes from one mRNA was constructed. The sequence starting from the 5’ untranslated region of 18nt to 32nt in HCV core coding region was cloned and then the encephalomyocarditis virus (ECMV) IRES sequence in the commercial vector pIRES was substituted to construct a new vector pCVIR. Green fluorescent protein (GFP) and Hepatitis B virus surface antigen (HBsAg) coding genes were inserted up-stream and down-stream of IRES sequence. The fluorescence intensity of GFP and HBsAg were determined by flow cytometry and ELISA respectively., thus, the expression efficiency of the two vectors, pCVIR and pIRES could be compared. The experimental results showed that the vector pCVIR could translate the GFP and HBsAg genes down-stream of its HCV IRES sequence more efficiently without impairing the expression of genes up-stream of IRES sequence than the vector pIRES.It is concluded that a new eukaryotic bi-cistronic expression vector containing HCV IRES was constructed successfully by the method described above.In this paper, a new eukaryotic bi-cistronic expression vector containing Hepatitis C Virus（HCV） internal ribosome entry site （IRES） expressing two foreign genes from one mRNA was constructod. The sequence starting from the 5＇ untranslated region of 18nt to 32nt in HCV core coding region was cloned and then the encephalomyocarditis virus （ECMV） IRES sequence in the commercial vector plRES was substituted to construct a new vector pCVIR. Green fluorescent protein （GFP） and Hepatitis B virus surface antigen （HBaAg） coding genes were inserted up-stream and down-stream of IRES sequence. The fluorescence intensity of GFP and HBaAg were determined by flow cytometry and ELISA respectively., thus, the expreasion efficiency of the two vectors, pCVIR and plRES could be compared. The experimental results showed that the vector pCVIR could translate the GFP and HBsAg genes down-stream of its HCV IRES sequence more efficiently without impairing the expression of genes up-stream of IRES sequence than the vector plRES. It is concluded that a new eukaryotic bi-cistronic expression vector containing HCV IRES was constructed suecessfially by the method described above.

关 键 词：丙型肝炎病毒 内部核糖体进入位点 真核表达载体 双顺反子 

分 类 号：R373.2[医药卫生—病原生物学]