Cloning, expression and purification of early secretory antigenic target 6kDa protein (ESAT-6) of Mycobacterium tuberculosis

Zahra Farshadzadeh; Mojtaba Sankian; Forough Yousefi; Aida Gholobi; Reza Zarif; Mahboobeh Naderi nasab; Tahereh Rashed; Abdolreza Varasteh

Jundishapur Journal of Microbiology

Ahvaz Jundishapur University of Medical Sciences

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Cloning, expression and purification of early secretory antigenic target 6kDa protein (ESAT-6) of Mycobacterium tuberculosis

Author(s):

Zahra Farshadzadeh^1,*,

Mojtaba Sankian²,

Forough Yousefi²,

Aida Gholobi²,

Reza Zarif²,

Mahboobeh Naderi nasab²,

Tahereh Rashed²,

Abdolreza Varasteh²

1Infectious and Tropical Diseases Research Center, Ahvaz Jundishapur University of Medical Sciences, Zahra.farshadzadeh@gmail.com, Iran

2Booali Immunology Research Center Mashhad University of Medical Sciences, Iran

Jundishapur Journal of Microbiology:Vol. 3, issue 2; 53-60

Article type:Research Article

Received:Aug 01, 2009

Accepted:Feb 01, 2010

How to Cite:Zahra FarshadzadehMojtaba SankianForough YousefiAida GholobiReza ZarifMahboobeh Naderi nasabTahereh RashedAbdolreza Varastehet al.Cloning, expression and purification of early secretory antigenic target 6kDa protein (ESAT-6) of Mycobacterium tuberculosis.Jundishapur J Microbiol.3(2):53-60.

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.

Keywords

ESAT6 antigen

Cloning

Mycobacterium tuberculosis

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