بیان همزمان ژنهای mtb32c و hbha از مایکوباکتریوم توبرکلوزیس سویه H37Rv در سیتم پروکاریوت

Title

Co-Expression of hbha and mtb32C Genes from Mycobacterium tuberculosis H37Rv in a Prokaryotic System

English Abstract

Background and Objective: HBHA (heparin-binding hemagglutinin) is a surface adhesin that mediate in binding to host cells by its own unique, carboxyl-terminal region. This methylated region has specific motif that rich of lysine-, alanine-, and proline amino acids. More recently, has been shown that HBHA has potential activity in stimulating immune responses and is a promising new candidate for diagnostic and protective antigen against tuberculosis. Interferon-gamma release assay test (IGRAs) is a new method to identifying latent tuberculosis and in compared to old method, tuberculin skin test (TST), has several advantages. Therefore in this study recombinant heparin-binding haemagglutinin antigen as new antigen in IGRAs test was produced.Materials and Methods: In present work hbha and mtb32C genes were isolated from Mycobacterium tuberculosis H37Rv genome by PCR. PCR products and pet21+ vector were digested with specific restriction enzymes and then submitted to ligation procedure. E. coli BL21-CodonPlus (DE3) competent cells were transformed with recombinant Mtb32C-HBHA –pet21+ vector. Expression of recombinant protein (Mtb32C-HBHA) was confirmed with SDS-PAGE and Western blot methods.Results: Detection of about 500 bp gene in Colony-PCR method and sequencing of recombinant pet-Mtb32C-HBHA vector all confirmed the accuracy of cloning procedure. Also presence of a 36 KDa protein band was confirmed with Western blotting method. Conclusion:In this study, expression of Mtb32C-HBHA protein in prokaryotic system successfully was done and production of new recombinant protein confirmed by western blot technique and anti His tag antibodies. Other studies are needed to evaluate efficacy of recombinant Mtb32C-HBHA protein in diagnosis of latent tuberculosis.

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