丙型肝炎病毒C区和NS3区嵌合基因的克隆及表达

Cloning and Expression of HCV C-NS3 Chimeric Gene

目的获得丙型肝炎病毒(HCV)C-NS3嵌合抗原,以提高HCV酶联免疫吸附试验诊断试剂的质量。方法应用SOE-PCR(拼接重叠延伸PCR)的方法,构建了HCV的C区和NS3区的嵌合基因,克隆于表达载体pGEX-4T3,并转化于大肠杆菌BL21,经筛选,IPTG诱导HCV的C-NS3嵌合抗原的表达。经SDS-PAGE检测表达水平,Western blot检测抗原特异性。结果SOE-PCR构建的HCV C区和NS3区嵌合基因在BL21中获得表达。经鉴定,其相对分子质量约为75000,并具有高度特异性。结论已获得HCV C-NS3嵌合抗原,为提高HCV酶联免疫吸附试验诊断试剂的质量奠定了基础。

Objective To obtain the C-NS3 antigen of HCV and improve the quality of ELISA diagnostic kit for HCV. Methods Construct the C-NS3 chimeric gene of HCV by SOE-PCR,clone into expression vector pGEX-4T3 and transform to E. coli BL21 for expression under induction of IPTG. Determine the expression level by SDS-PAGE and the specificity of expressed protein by Western blot. Results HCV C-NS3 chimeric antigen, with a relative molecular weight of 75 000, was successfully expressed and showed good specificity. Conclusion HCV C-NS3 chimeric antigen was obtained, which laid a foundation of improving the quality of ELISA diagnostic kit for HCV.