Cloning, expression and purification of early secretory antigenic target 6kDa protein (ESAT-6) of Mycobacterium tuberculosis
Journal Title: Jundishapur Journal of Microbiology - Year 2010, Vol 3, Issue 2
Abstract
Introduction and objective: The early secretory antigenic target 6kDa protein (ESAT-6) antigen from Mycobacterium tuberculosis is a dominant target for cell-mediated immunity in the early phase of tuberculosis (TB) in TB patients. ESAT-6 was found to distinguish TB patients from BCG-vaccinated donors. The aim of this study was cloning and expression of ESAT-6 of M. tuberculosis. Materials and methods: DNA was extracted from M. tuberculosis H37Rv. PCR was performed using gene-specific oligonucleotide primers and the PCR products were inserted into the pET102/D vector and transferred into Escherichia coli strain TOPO10. The recombinant plasmids transferred into E. coli strain BL21. Results: The transformed plasmid into E. coli strain BL21 was effectively expressed. The expressed fusion protein (23kDa on SDS-PAGE) was found almost entirely in the soluble form and the recombinant protein was purified by Ni-NTA column. Conclusion: We successfully cloned and expressed ESAT-6 protein of M. tuberculosis in E. coli. As a specific antigen, it can be useful for diagnosis of both active and latent tuberculosis with ELISA in future
Authors and Affiliations
Zahra Farshadzadeh, Mojtaba Sankian, Forough Yousefi, Aida Gholobi, Reza Zarif, Mahboobeh Naderi nasab, Tahereh Rashed, Abdolreza Varasteh
Keywords
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