Fusion and Prokaryotic Expression on the mpb83 and mpb63 Genes of Mycobacterium Bovis

Abstract:

Based on the (Gly4Ser)3 linker, the mpb83 and mpb63 gene were fused for raising the antigenicity of single antigen. The DNA fragments of mpb83 and mpb63 were fused by splicing by overlapping extension (SOE) polymerase chain reaction (PCR) ,and the fusion gene mpb83-63 were cloned into pMD18-T vector, and then we got the recombinant plasmid pMD-83-63. pMD-83-63 and pET28a (+) were digested by BamH I and EcoR I double enzymes. The purified mpb83-63 fusion gene was subcloned into the expression vector pET28a (+),and the prokaryotic expression vector pET-83-63 was constructed. Plasmid containing pET-83-63 was transformed into competence Escherichia coli BL21(DE3).The bacterium was induced by isopropyl-β-D-thioga-lactopyranoside (IPTG) and analyzed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE),approximately 38 kDa exogenous protein was observed on the SDS-PAGE. The protein was analyzed by using Western-blotting. The results indicated that the protein was of antigenic activity of Mycobacterium bovis. These results could serve as a basis for further studies on the usefulness of the fusion gene and its expression product in the development of DNA vaccine; living carrier vaccine; subunit vaccine and diagnostic reagents against bovine tuberculosis.

• Periodical: Advanced Materials Research (Volumes 791-793)
• Pages: 212-215
• DOI: https://doi.org/10.4028/www.scientific.net/AMR.791-793.212
• Online since: September 2013
• Authors: Jia Ning Guan, Chun Fang Wang, Jia Ming Lin, Hong Xia Ma, Shuang Hou, Xin Liu, Qiang Gou, Xiu Yun Jiang
• Keywords: Fusion Gene, Fusion Protein, mpb63 Gene, mpb83 Gene, Mycobacterium bovis, Prokaryotic Expression

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