Construction of the Mycobacterium shuttle expression plasmid with Mycobacterium tuberculosis heat shock protein 70 promoter / 重庆医科大学学报

ABSTRACT

Objective:To reconstruct Mycobacterium shuttle expression plasmid with Mycobacterium tuberculosis HSP70 promoter based on shuttle plasmid pJEM11.Methods:The cDNA fragment of 229bp promoter and its correlative sequence was amplified by PCR.Multiple clone sites was added to the end of amplified fragment,and then it was cloned into the pJEM11 to construct the plasmid pJCH02.The pJCH02 was identified by digestion with restriction endonuclease and sequence analysis.The lhp-esat6 fusion gene was cloned into the pJCH02,then its expression in BCG was confirmed by SDS-PAGE.Results:The sequence of HSP70 promoter and its correlative sequence was in accordance with the predicted sequence.The lhp-esat6 fusion gene was expressed in BCG.Conclusion:The shuttle expression plasmid pJCH02 is constructed successfully.The study provides the basis for BCG polyvaccine.