Author/Authors :
Zarif، Reza نويسنده , , Sankian، Mojtaba نويسنده , , Gholubi، Aida نويسنده Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad , , Farshadzadeh، Zahra نويسنده , , soleimanpour، Saman نويسنده , , Youssefi، Forough نويسنده Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad , , Khaje Karamoddini، Mehrangiz نويسنده Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad , , Ghazvini، Kiarash نويسنده , , Varasteh، Abdol Reza نويسنده Division of Immunobiochemistry, Immunology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad ,
Abstract :
Background: The 30 kDa major secretory protein of Mycobacterium tuberculosis (antigen 85B) is a primary vaccine candidate. This secreted
antigen induces a protective immune response and stimulates the production of IFN-? in animal models.
Objectives: The aim of this study was cloning and expression of Ag 85B of M. tuberculosis in Escherichia coli.
Materials and Methods: To produce recombinant Ag85B, the fbpB gene was amplified by PCR method. Then inserted into the pET101/D
vector and transported into E. coli strain TOPO10. Plasmid containing pET101/D: Ag85B was transformed into competence E.coli BL21 (DE3). The
transformed E.coli strain BL21 was effectively expressed recombinant Ag85B.
Results: The expressed fusion protein was found almost entirely in the insoluble form. Followed by sonication to disrupt the cells, Solution of
the cell debris was centrifuged and after use of Ni–NTA column and 6 molar urea and 6 M guanidine-HCl solutions recombinant protein was
purified.
Conclusions: These results could serve as a basis for further studies in endemic regions of tuberculosis on the usefulness of this gene and its
expression product in the development of subunit vaccine and DNA vaccine against tuberculosis.
