Clone and express ctb-stx2 b fusion gene in Enterohemrrhagic escherichia coli O157:H7 Shigeai toxin 2B subunit and V cholera toxin B subunit and the detection of their immunogenicity
10.3321/j.issn:0254-6450.2008.04.016
- VernacularTitle:霍乱毒素B亚单位与志贺毒素2B亚单位融合表达及抗原性检测
- Author:
Zhen-Jun LI
1
;
Qiang-Zheng SUN
;
Huai-Qi JING
;
Jian-Guo XU
Author Information
1. 中国疾病预防控制中心传染病预防控制所
- Keywords:
Enterohemrrhagic escherichia coli O157:H7;
Shiga toxin 2B subunit;
Cholera toxin B subunit;
Immunogenicity
- From:
Chinese Journal of Epidemiology
2008;29(4):378-382
- CountryChina
- Language:Chinese
-
Abstract:
Objective To clone and express the fusion gene encoding Enterohemrrhagic escherichia coli O157:H7(EHEC O157:H7)Shigela toxin 2B subunit(Stx2B)and vibrio cholera toxin B subunit (CTB)as well as to detect the immunogenicity and GM1-binding ability of fusion protein.Methods To design a primer to amplify stx2b gene and ctb-stx2b fusion gene encoding Stx2B and CTB-Stx2B respectively and to clone the genes into express plasmid pET30a(+)C in order to construct pET30a-ctb-stx2b after T-A sequencing was varified,then to transform constructed plasmid into E.coliBL21(DE3)induced by IPTG and purified by a purify kit and to detect molecular weight and immunogenicity by SDSPAGE and Western-blot.Results The amplified ctb-stx2b fragments appeared to he 750 bp and gene sequence was identical to designed sequence.The prokaryotic expression system pET30a-ctb-stx2b/BL21 could express protein weight about Mr20×103and the expressed protein could react to CTB monoclone anti-body.The fusion protein CTB-Stx2B could bind GM1.Conclusion CTB-Stx2B had successfully been expressed in prokaryotic while the expressed protein had good immunogenicity and GM1-Binding ability.This study provided information on further EHEC O157:H7 vaccine research.
