GAO Ya-li, et al. Prokaryotic expression,identification and purification of HCV F protein[J]. Journal of Chongqing Medical University, 2009,34(4):385-388.
GAO Ya-li, et al. Prokaryotic expression,identification and purification of HCV F protein[J]. Journal of Chongqing Medical University, 2009,34(4):385-388.DOI：
Objective:To construct a prokaryotic expression plasmid containing HCVF gene
and purify the recombinant fusion protein in E.coli system. Methods: F gene of Hepatitis C virus was amplified by PCR method from plasmid H/FL(containing full length cDNA sequence of HCV 1a subtype)
cloned into pET32a(+)vector
and then transformed into E.coli JM109. After identified by restriction diges- tion and DNA sequencing
recombinant plasmid was transformed into E.coli BL21 and induced with IPTG. The fusion protein trxA- F was further confirmed by Western blot analysis and purified by affinity chromatography method. Results: Restriction digestion and PCR screening showed that HCV F gene was cloned into pET32a(+)successfully. After BL21 was transformed with recombinant vector pET32a (+)- HCVF and induced with 0.5mmol/L IPTG
a 35.4kDprotein band was found bySDS- PAGE. And this recombinant protein showed a highly specific and strong reaction with anti- His monoclonal antibody by Western blot analysis. Conclusion: The prokaryotic expression plasmid pET32a(+)- HCVF was successfully constructed
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