Construction and expression of Mycobacterium tuberculosis PPE68/GST fusion protein expression plasmid

AIM: To construct the Mycobacterium tubereulosis (MTB) PPE68/GST fusion protein expression vector and induce its expression in E.coli JM109. METHODS: Rv3873 gene was amplified from MTB H37Rv by PCR and directionally cloned into expression plasmid pGEX-4T-1. Construct the recombinant expression plasmid pGEX-4T-1-Rv3873 and express it in E.coli JM109 under the induction of isopropyl-β-D-thiogalactopyranoside (IPTG). The fusion protein was purified by GST affinity chromatgraphy and confirmed by SDS-PAGE and Western Blot. RESULTS: The desired PPE68/GST fusion protein (M_r 63 000) was successfully expressed, accounting for 40% of the total bacteria. Western bolt revealed that the fusion protein could react with the polyvalent antiserum of tuberculosis patients. CONCLUSION: Prokaryotic expression plasmid pGEX-4T-1-Rv3873 was successfully cloned and expressed in E.coli JM109.