Expression and identification of recombinant Clostridium difficile toxin B using Bacillus megaterium system
10.3760/cma.j.issn.1000-6680.2011.01.001
- VernacularTitle:艰难梭状杆菌肠毒素B在巨大芽孢杆菌中的表达和鉴定
- Author:
Guilin YANG
;
Weilong LIU
;
Hongyan YAO
;
Boping ZHOU
;
Hanping FENG
- Publication Type:Journal Article
- Keywords:
Clostridium difficile;
Bacillus;
Entero toxins;
Gene expression;
Plasmids;
Recombinant proteins
- From:
Chinese Journal of Infectious Diseases
2011;29(1):1-5
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express and purify recombinant and biologically active Clostridium difficile toxin B (rTcdB). Methods The genes of TcdB were amplified by polymerase chain reaction (PCR) using chromosomal DNA from a toxigenic strain, and cloned into a shuttle vector pHis1522.The sequences of TcdB genes in the vector were verified by DNA sequencing. The construction was transformed into Bacillus megaterium protoplasts and the protein expression was driven by a xylose promoter. The purified protein was tested for biological activity. Results rTcdB was successfully purified from bacterial crude extracts. Approximately 5-10 mg of highly purified recombinant toxin was obtained from one liter of bacterial culture. The expressed rTcdB had molecular mass similar to the native toxin, and its biological activity was proved to be similar to its native counterpart after an extensive examination. Conclusion rTcdB with biological activities is successfully expressed in Bacillus megaterium.
