Cloning and Expression of Secreted Protein Ag85B of Mycobacterium Tuberculosis

Purpose To clone and express the ag85b ,one gene of the mycobacterium secreted proteins,and to establish a basis for diagnasis of tuberculosis. Methods We used genomic DNA of M.tuberculosis H37Rv strain as template,amplified the ag85b gene with PCR constructed the recombinant plasmid with ag85b and vector plasmid pET24b,transformed the recombinant plasmid pET24b ag85b into E.coli DH5α.Then purified the plasmid DNA,tested it by restriction endonucleases digest,transformed the plasmid into expressive strain E.coli JM109(DE3) again,induced the transformants with IPTG,split bacteria,centrifuged the split fliud and the supernatant was used for electrophoresis. Results SDS?PAGE demonstrated that the transformants expressed Ag85B,the protein molecular weight was 30 000. Conclusions The target gene has been cloned into host bacterium strain.Expression of Ag85B makes a basis of diagnosis of tuberculosis.