Construction of recombinant plasmid pGEX-Sj32 of Schistosoma japonicum and expression in Escherichia coli BL21 / 国际检验医学杂志

ABSTRACT

Objective To construct the recombinant plasmid pGEX-Sj32 of Schistosoma japonicum(Sj )and to research its ex-pression in Escherichia coli (E.coli)BL21.Methods Sj32 gene was amplified by PCR from template of plasmid pET28α-Sj32 ex-tracted from recombinant bacterium BL21 (pET28α-Sj32 )stored by our laboratory,and then cloned into the vector pGEX-1λT to construct pGEX-Sj32.The recombinant plasmid pGEX-Sj32 was transformed into E.coli BL21(DE3).The recombinant strains were induced by isopropyl-β-d-thiogalactoside(IPTG),and the expressed products were identified by SDS-PAGE and Western blot.Re-sults Sj32 coding gene was successfully amplified by PCR and cloned into the vector pGEX-1λT,and the recombinant plasmid pGEX-Sj32 was constructed successfully.The molecular mass of the expressed recombinant protein was proximately 58 000 as de-tected by SDS-PAGE.The amount of the expressed protein was about 21% of the total bacterial protein.Western blot confirmed that the expressed protein could be recognized by the immune sera from rabbit infected with Schistosoma japonicum.Conclusion The recombinant plasmid pGEX-Sj32 is successfully constructed.The Sj32 protein was highly expressed in E.coli and the expressed recombinant protein possesses the specific antigenicity.