همسانه سازی، بررسی بیان و تخلیص ژن کد کننده پروتئین نوترکیب متشکل از زیرواحدهای اتصالی (ETEC) از اشرشیا کلی انتروتوکسیژنیک CS و 2 CSI آنتیژنهای سطحی (Persian)

Cloning and the Evaluation of Gene Expression and Purification of Gene Encoding Recombinant Protein Containing Binding Subunit of Coli Surface Antigens CS1 and CS2 from Enterotoxigenic Escherichia coli. (English)

Background & Objective: Enterotoxigenic Escherichia coli (ETEC) is a major causative agent of diarrhea. Enterotoxins and the colonization factors (CFs) are major virulence factors in ETEC infections. The bacterium binds to the intestinal epithelial cell surface through colonization factors and produces enterotoxins that cause excessive fluid and electrolyte secretion in the lumen of the intestine, which ultimately leads to diarrhea. Due to the difficulty of treatment and the high prevalence of this disease, the design of the vaccine against this organism is one of the goals of the World Health Organization. The CooD-CotD protein, as adhesion tip subunits of CS1 and CS2, plays an important role in bacterial attachment to the intestinal epithelial cells. In this study, the expression and purification of chimeric protein CooD-CotD was carried out with the aim of investigating as candidate vaccine. Materials & Methods: In this study, codon optimization of cooD-cotD chimeric gene was performed by Gene Designer software. The gene was amplified by PCR and cloned into pJET1.2 / blunt then it was subcloned into pET28a to express chimeric protein. Recombinant protein was purified following expression, using Ni-NTA affinity chromatography and confirmed by western blotting analysis. Results: The presence of 82kDa in the SDS-PAGE gel showed that expression of CooD-CotD chimeric protein and confirmed by western blotting analysis. Yield of purified protein was 121 mg/ml. Conclusions: Expression and protein purification studies showed that this protein has expression in the homologous host. [ABSTRACT FROM AUTHOR]

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