Volume 2, Issue 3 (11-2012)                   JABS 2012, 2(3): 135-141 | Back to browse issues page

‎ 20.1001.1.22285105.2012.2.3.3.1

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Fusion of Cholera toxin B subunit (ctxB) with Shigella dysenteriae type I toxin B subunit (stxB), Cloning and Expression that in E. coli
Hamid Madanchi * 1, Hossein Honari , Sadegh Safaei , Ali Sayadmanesh
1- , hamidmadanchi@yahoo.com
Abstract:   (20249 Views)
Background and Objective: Shiga toxin (STx) is the main virulence factor in Shigella Dysenteriae type I and is composed of an enzymatic subunit STxA monomer and a receptor-binding STxB homopentamer. Shigella toxin B subunit (STxB) is a non-toxic homopentameric protein responsible for toxin binding and internalization into target cells by interacting with glycolipid (Gb3). Cholera toxin B subunit (CTB) has been known as a mucosal adjuvant for vaccines and genetic fusions of CTB with several hetroantigens such as stxB and can increase humoral and mucosal immunity response.
Materials and Methods: In this study, after primer designing, the ctxB and stxB genes were amplified by PCR and cloned into the pGEM-T vector. The stxB gene with a nonfurin linker was fused to the ctB gene in the pGEM vector via the restriction enzyme method and thereafter the fused genes of ctB-stxB were subcloned in the pET28a(+) as an expression vector. The expressed chimeric protein was induced with IPTG and evaluated via the SDS.PAGE and Western blot techniques.
Result: The pET28a (+)/ctxB-stxB expression vector was confirmed by endonuclease digestion, PCR, and sequence analysis. The CTB-STB fusion protein was confirmed by the SDS-PAGE and Western-blot.
Conclusion: The CTB-STB recombinant protein can be used as a new and desirable mucosal vaccine for Shigella Dysenteriae type I.
Keywords: Adjuvant; CTB; Nonfurin linker; StxB; Shigella Dysenteriae type I
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Type of Study: Research | Subject: Microbiology
Received: 2013/01/31 | Accepted: 2013/09/14 | Published: 2013/09/14
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