2006•Biotechnology(Faisalabad)Requires access
Gene Cloning and Prokaryotic Expression of Human Chemokine Receptor CXCR4
Deli Liu
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Abstract
Objective:To construct prokaryotic expression vector and express human CXCR4 protein in E.coli.Methods:The total RNA was extrated from normal human PBMC and used as a template for RT-PCR.A 1059bp fragment containing the whole length gene coding region of protein CXCR4 was obtained and cloned into pMD18-T vector.The recombinant which had analysied by restriction endonucleases and clony PCR was sequenced to confirm its idenity.The right recombinant was ligated with vector pET-28a(+)and transformed into E.coli BL21(DE3).Results:After being induced by IPTG at 30℃ for 4h,a fusion protein His-CXCR4 with an apparent MW of 43ku was obtained in 12%SDS-PAGE gel and accounted for 25% of the total cellular protein.Conclusion:Human CXCR4 fusion protein was obtained successfully.