Construction and expression of the recombinant plasmid pET32α-Sj26GST of Schistosoma japonicum in Escherichia coli BL21(DE3)
10.3760/cma.j.issn.1000-4955.2010.03.013
- VernacularTitle:日本血吸虫谷胱甘肽-S-转移酶重组质粒的构建及其在大肠埃希菌BL21(DE3)中的表达
- Author:
Wen-gui, LI
;
Bang-zhong, XIAO
;
Xing-jian, LUO
;
Ya-tang, CHEN
;
Cheng-guo, WU
- Publication Type:Journal Article
- Keywords:
Schistosoma japonicum;
Recombinant plasmid pET32α-Sj26GST;
Escherichia coli;
Vaccines,synthetic
- From:Chinese Journal of Endemiology
2010;29(3):287-291
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct and express the recombinant plasmid pET32α-Sj26GST of Schistosoma japonicum(sj)in Escherichia coli(E.coli)B121(DE3).Methods The total RNA was extracted from sj adult worms by ultrasound-breaking,Sj26GST antigen gene was amplified by RT-PCR from the total RNA,then cloned into prokaryotic expression plasmid pET32α(+) and transformed into E.coli B12(DE3)to construct pET32α-Sj26GST;BL21(pET32α-Sj26GST)WaS induced with isopropyl-β-D-thiogalactopyranosid(IPTG),and the expressed products were analyzed and identified by SDS-PAGE and Western blot.Results The 676 bp Sj26GST gene was successfully amplified by RT-PCR and cloned into pET32α(+)by restriction analysis and PCR identification,the recombinant plasmid pET32α-Sj26GST was successfully constructed;the relative molecular mass of the expressed recombinant protein was approximately 49×103 by SDS-PAGE,and the amount of the expressed protein was 24%of the total bacterial proteins;the fusion protein could be recognized by sera from rabbits infected with sj by Western blot.Conclusions The recombinant plasmid pET32α-Sj26GST is successfully constructed and highly expressed in E.coli in fused form with Trx-tag and His-tag,and the expressed fusion protein shows specific antigenicity.
