Construction of clone and expression vector of gene of vibrio parahaemolyticus

  Objective   To construct clone and expression vector of tlh gene of vibrio parahaemolyticus and examine its expression in DE ₃ for preparaing monoclonal antibody and dignosing reagent and studying function of tlh.

  Methods   Using VP14-90 as template, the tlh gene was amplified by modified PCR with a pair of specific primer which contained the restrictive sites of Eco RV and HindⅢ, and then subcloned into plasmid pGEM-T and expression plasmid pET32a.The recombinant plasmids were identified by restiction endonuclease enzyme analysis and PCR.The recombinant protein expression was induced by IPTG.

  Results   The amplified frament was about 1.3kb by showed on the agarose gel electrophoresis and had 99% homeologous with the tlh gene in Genbank.The recombinant protein was about 63 KD.

  Conclusion   The PCR method amplified the tlh gene.A clone vector and a expression vector containing tlh gene was successfully constructed, and the cell DE ₃ transformed with the expression vector was capable of expression tlh gene.