Volume 15, Issue 9 (February 2013)
J Arak Uni Med Sci 2013, 15(9): 51-60 |
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Rezaee M, Honari H, Zand A M, Arefpour torabi M A. Recombinant expression and purification of Bacillus anthracis lethal factor domain 1 in Escherichia coli and production of polyclonal antibody against it in mice . J Arak Uni Med Sci 2013; 15 (9) :51-60
URL: http://jams.arakmu.ac.ir/article-1-1798-en.html
URL: http://jams.arakmu.ac.ir/article-1-1798-en.html
1- ihu
2- ihu ,honari.hosein@gmail.com
2- ihu ,
Abstract: (13438 Views)
Background: Anthrax is a common disease among human and livestock which is caused by Bacillus anthracis. Bacillus anthracis has two strong immunogenic proteins: Protective antigen (PA) and lethal factor domain I (LFD1) that have always been considered as vaccine candidates against Bacillus anthracis. The aim of this study is to express and purify the lethal factor domain I (LFD1) in Escherichia coli and produce polyclonal antibody against it in mice. Materials and Methods: In this experimental study, LFD1 gene was amplified with BamH I and Xho I restriction site by PCR. After isolation, the gene was cloned to the expression vector pET28a (+). This vector was transformed to E. coli-BL21 (DE3) PLysSto to express LFD1 gene. The expression of LFD1 gene was induced by IPTG. After protein purification by affinity chromatography, the produced antigen was injected into mice for four times. Then the produced polyclonal antibody in mice serum was evaluated. Results: The cloned LFD1 gene in pET28a (+) vector was confirmed by PCR, enzymatic analysis, and sequencing. The expressed and purified recombinant protein was confirmed by SDS-PAGE and Western blotting. Finally, the isolated polyclonal antibody from mice serum was evaluated and confirmed by ELISA test. Conclusion: Noticing the appropriate expression, easy purification of LFD1, and the titer of produced polyclonal antibody against LFD1 in mice due to its immunogenicity, it can be considered as a good vaccine candidate against anthrax.
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