Cloning and expression of Escherichia coli O157∶H7 Stx2B subunit / 医学研究生学报

ABSTRACT

Objective: To clone,identify and express the Shigalike toxin 2B(Stx2B) subunit gene of EHEC O157∶H7.Methods: A pair of primers were designed based on the Stx2B subunit gene sequence of EHEC O157∶H7.The Stx2B gene was amplified from the EHEC O157∶H7 chromosome by PCR and cloned into the pMD18-T vector.Thereafter,the gene was cut from the pMD18-T vector and cloned into the prokaryotic expression plasmid pET-28a vector.Then the recombinant plasmids were transformed into E. coli BL21(DE3) and the transformed host strain induced by IPTG.The expression protein was detected by SDS-PAGE and Western-blot analyses.Results: The Stx2B gene was successfully cloned into pMD18-T and pET-28a vectors,and the expressed protein identified by SDS-PAGE and Western blot.The molecular mass(Mr) of the expressed product was about 7 500,and the expression rate about 40%. Conclusion: The Stx2B gene was successfully cloned and effectively expressed in the prokaryotic expression system.