Abstract: Objective To obtain the recombinant CFP10-ESAT6 fused protein of M.tuberculosis .Methods The gene coding CFP-10 and ESAT-6 protein was amplified by polymerase chain reaction (PCR).The gene was inserted into an expression vector pET-28a to get recombinant plasmid.The recombinant plasmid was transformed into E.coli BL21(DE3).The E.coli carrying recombinant plasmid were induced by IPTG.The expressed product was indentified by SDS-PAGE and purified by metal chelation chromatography,and its immunological characteristics were analyzed by Western-blotting and ELISA technology.Results The sequence of CFP-10 and ESAT-6 in recombinant plasmid was consistented to that of Genbank report. The recombinant CFP10-ESAT6 protein was highly expressed in the form of solution in E.coli ,which was confirmed by Western-blotting analysis with monoclonal antibody against 6×His·tag and TB patient serum .ELISA test Results indicated that the purified recombinant protein could distinguish sera from tuberculosis patients and those from healthy people.Conclusion The recombinant CFP10-ESAT6 protein was expressed and purified successfully in the form of solution in E.coli and showed specific immunogenicity.

Key words: Mycobacterium tuberculosis /antigen, ESAT-6, CFP-10, Gene expression, Immunological