Construction of fusion vector of core gene and NS4B gene of HCV and its expression in E.coli
FANG Yu-zhu 1, ZHU Yan 1, YE Sen 2, YAO Dong-ming 1
(1. Department of Clinical Laboratory, the Affiliated People′s Hospital of Jiangsu University, Zhenjiang Jiangsu 212002； 2. NORMAN Biological Technology， Nanjing Jiangsu 210031, China)
Abstract：Objective: To construct a fusion gene clone of hepatitis C virus(HCV) core protein and NS4B and to check it expression in E.coli. Methods: cDNA fragments of HCV core gene and NS4B gene were separately amplified from the anti-HCV positive sera by PCR, followed by connecting them to form an intact cDNA and inserted into the expression vector pET28a.Positive recombinant plasmid named core-NS4B-pET28a was screened and confirmed by colony PCR and DNA sequencing. Afterwards it was transformed into E.coli BL21 to express recombinant fusion protein, which was analyzed by SDS-PAGE and Western blotting. Results: Agarose gel electrophoresis showed that core gene and NS4B gene as well as the corresponding cDNA consisting of them were obtained successfully. Colony PCR and DNA sequencing showed that the plasmid core-NS4B-pET28a was constructed successfully. SDS-PAGE clearly demonstrated the successful expression of the fusion protein in E.coli BL21. Western blotting showed that the protein had reactivity with anti-HCV positive sera. Conclusion: The fusion expression vector for core gene and NS4B gene of HCV was successfully constructed and the protein was expressed successfully in E.coli. ［Key words］hepatitis C virus； core protein； NS4B； construct； expression
房玉珠1, 朱燕1, 叶森2, 姚冬明1. 丙型肝炎病毒核心蛋白与NS4B融合蛋白的构建和表达[J]. 江苏大学学报:医学版, 2017, 27(06): 470-473.
FANG Yu-Zhu-1, Zhu-Yan-1, Ye-Sen-2, Yao-Dong-Ming-1. Construction of fusion vector of core gene and NS4B gene of HCV and its expression in E.coli. Journal of Jiangsu University(Medicine Edition), 2017, 27(06): 470-473.
2017, Vol. 27 
