Gene Cloning Expression and Purification of 16 KD Protein of Mycobactrium Tuberculosis

Objective By constructing expression vector, to express of 16KD protein of Mycobacterium Tuberculosis in E.coli and to gain a great deal purified 16KD protein.Methods Polymerase chain reaction (PCR) was employed to amplify the encoding gene of 16KD protein from Mycobacterium tuberculosis H37Rv. Then the amplified fragment was ligated to plasmid pGEX-4T-2. After the expression vector was transformed into E.coli DH5α, IPTG were added to induce the expression of the GST-16 fusion protein . the molecular mass and expression form of the fusion protein was analyzed by SDS-PAGE, and the activity of the protein was identified by Western blot. At last, the purified 16KD protein was gained by GST affinity chromatography.Results The expression vector was constructed successfully. There was about 42 % of fusion protein in total cellular proteins of the E.coli DH5α. Western blot demonstrated GST-16 fusion protein was able to combine with anti-TB antibody specially. The concentration of purified 16KD protein was 688 mg/L through ultraviolet spectrophotometry. Conclusion MTB 16KD protein is able to be high expressed in E.coli DH5, and it possess special immunological activities.