Cloning and Prokaryotic Expression of Heat Shock Protein 65 Gene of Mycobacteria Intracellular

In order to research Mycobacteria intracellular and the foundation of heat shock protein65(Hsp65),the study clone the HSP65 gene of Mycobacteria intracellular and express it in E.coli.The method:The HSP65 gene is amplified from Mycobacteria intracellular ATCC13 950 genome DNA using PCR method.then inserted into PMD18-T vector and sequenced.The recombinant plasmid is called PMD18-T-HSP65.The PMD18-T-HSP65 is digested by the restriction endonucleases NdeⅠand BamHⅠand then ligated to the the prokaryotic expression vector PET15b which is digested by the same restriction endonucleases to product the recombinant prokaryotic expression vector PET15b-HSP65,then transform the BL21(DE3)/PolysS strain,and induce it with 1mmol/L IPTG.,The E.coli.containing PET15b-HSP65 could express a protein with Mr being 65 000 after analysis of SDS-PAGE.Conclusion:The study obtained the recombinant expressing plasmid and expressed successfully.