Production of MPT-64 recombinant protein from virulent strain of .

Tuberculosis (TB) is a zoonotic infectious disease common to humans and animals which has been caused by a rod shaped, acid fast bacterium, called . The rapid and sensitive detection is a great challenge for TB diagnosis. The virulent strains of complex (MTBC) have 16 different regions of difference (RD) in their genome which encode some important antigens. The major protein of 64 (MPT-64) is one of the main immune-stimulating antigens which are encode by RD-2 region. The aim of the present study was cloning, expression and purification of MPT-64 as a protein antigen of in a prokaryotic system for the usage in the future diagnostic studies. In this experimental study, the gene with 687 bp has been proliferated from whole genome by polymerase chain reaction (PCR) method. The PCR product has been digested by HI and dIII restriction enzymes and cloned into pQE-30 plasmid. The recombinant protein has been expressed in the M15 with induction by isopropyl-β-D-thiogalactopyranoside (IPTG). The expressed protein was analyzed on SDS-PAGE, and purified with Nitrilotriacetic acid (Ni-NTA) column. Finally, its biological properties were confirmed in Western blotting method using specific antibodies. Data showed the successful cloning of gene (as a 687 bp segment) in expression vector. The MPT-64 recombinant protein was ideally expressed and purified as a 24 kDa protein. The result of this study indicated that MPT-64 recombinant protein (24 kDa) has been successfully expressed and purified in a prokaryotic system, so this protein could be used for differential diagnosis of pathogenic and non-pathogenic , in suspected BTB cases.