Expression of recombinant Clostridium difficile toxin A and B in Bacillus megateriumReportar como inadecuado




Expression of recombinant Clostridium difficile toxin A and B in Bacillus megaterium - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

BMC Microbiology

, 8:192

First Online: 06 November 2008Received: 21 August 2008Accepted: 06 November 2008

Abstract

BackgroundMajor Clostridium difficile virulence factors are the exotoxins TcdA and TcdB. Due to the large size and poor stability of the proteins, the active recombinant TcdA and TcdB have been difficult to produce.

ResultsThe toxin genes tcdA and tcdB were amplified by PCR using chromosomal DNA from a toxigenic strain as a template, and cloned into a shuttle vector pHis1522. The sequences of both tcdA and tcdB genes in the vector have been verified by DNA sequencing. The constructs were transformed into B. megaterium protoplasts and the protein expression was controlled under a xylose promoter. The recombinant toxins rTcdA and rTcdB were purified from bacterial crude extracts. Approximately 5 – 10 mg of highly purified recombinant toxins were obtained from one liter of bacterial culture. The resulting rTcdA and rTcdB had similar molecular masses to the native toxins, and their biological activities were found to be similar to their native counterparts after an extensive examination.

ConclusionWe have generated the full length and active recombinant TcdA and TcdB in Bacillus megaterium.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2180-8-192 contains supplementary material, which is available to authorized users.

Guilin Yang, Boping Zhou contributed equally to this work.

Download fulltext PDF



Autor: Guilin Yang - Boping Zhou - Jufang Wang - Xiangyun He - Xingmin Sun - Weijia Nie - Saul Tzipori - Hanping Feng

Fuente: https://link.springer.com/article/10.1186/1471-2180-8-192







Documentos relacionados