Expression of blf1 Gene of Burkholderia pseudomallei in Escherichia coli and Assessment of Antibody Titer in Mouse

Abstract

Background and Objectives: Burkholderia pseudomallei is the cause of the melioidosis disease in human, and is transmitted to patients through oral route, inhalation, or skin scratch. This agent is classified as a category B in the classification of biological agents. BLF1 specifically deaminates Gln339 from eIF4A. In this study, the expression of blf1 gene of B. pseudomallei in E. coli and antibody production in mouse, were assessed.   Methods: In this experimental study, blf1 gene of B. pseudomallei, was expressed in E. coli. The synthetic gene in pUC57 plasmid was purchased from Nedaye Fan COR. pUC57 plasmid containing blf1 gene with BamHI and SalI restriction enzyme sites, was subcloned in pET28a(+) expression vector and transformed into E. coli BL21(DE3). blf1 gene expression was induced by IPTG.   Results: In this study, blf1 gene in cloned pET28a(+) expression vector, was approved by PCR and enzymatic analysis. Also, the produced recombinant protein was confirmed by SDS-PAGE and Western blotting. Then, antibody produced from the mice serum, was isolated and confirmed by ELISA test.   Conclusion: Given that BLF1 protein has the ability to stop protein synthesis and the produced antibody was confirmed by ELISA, the BLF1 recombinant protein can be used to treat cancer and as a vaccine candidate against B. pseudomallei.  

Keywords

• burkholderia pseudomallei
• blf1 toxin
• eif4a
• melioidosis
• subcloning.