Volume 23, Issue 4 (2020)
mjms 2020, 23(4): 1-8 |
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Jafari Parsa F, Zare S, Mirhosseini S A, Amani J. Design of vaccine DNA containing genes encoding sicl Escherichia coli antigens Enterotoxigenic (ETEC) and Escherichia coli enterohemorrhagic (EHEC). mjms 2020; 23 (4) :1-8
URL: http://mjms.modares.ac.ir/article-30-45959-en.html
URL: http://mjms.modares.ac.ir/article-30-45959-en.html
1- Department of Genetics, Faculty of Basic Sciences, Islamic Azad University, Research Sciences Branch, Tehran, Iran.
2- Department of Genetics and Biotechnology, Pishva Faculty of Biological Sciences, Islamic Azad University, Varamin Branch, Iran.
3- Applied Microbiology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Sciences, Tehran- Iran.
4- Applied Microbiology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Sciences, Vanak Sq. Molasadra St. Tehran- Iran. , jafar.amani@gmail.com
2- Department of Genetics and Biotechnology, Pishva Faculty of Biological Sciences, Islamic Azad University, Varamin Branch, Iran.
3- Applied Microbiology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Sciences, Tehran- Iran.
4- Applied Microbiology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Sciences, Vanak Sq. Molasadra St. Tehran- Iran. , jafar.amani@gmail.com
Abstract: (811 Views)
Introduction: Enterotoxigenic Escherichia coli (ETEC) and Enterohemorrhagic Escherichia coli O157:H7 , the most common strains, are causes diarrhea that can kill hundreds of thousands of children annually. The development of an effective combination vaccine for enterohemorrhagic Escherichia coli and Enterotoxigenic Escherichia coli is very important.
Materials and Methods: In this study, the sicl gene was amplified and Subcloned as a DNA vaccine. The sequence of antigen encoding genes was evaluated from the genebank and their epitopes were evaluated to design of primer for the synthetic chimeric gene and was amplified by PCR. Subcloning of a multipartal chimeric gene in eukaryotic expression vector was performed to make a DNA vaccine and finally the protein was purified by nickel chromatography and evaluated by Western blotting.
Results: The immunoblotting results of the expression of SICL chimeric protein indicated the presence of a 76 kDa band in the form of insoluble particles after 12 hours induction. Purification of the recombinant protein using His tag sequencee and confirmation of the purified protein with the recombinant protein specific antibody demonstrated the accuracy of the protein expression.
Discussion & Conclusion: Protein expression, purification and verify by western blotting showed that this recombinant chimeric protein (SICL) can be expressed in eukaryotic host.
Materials and Methods: In this study, the sicl gene was amplified and Subcloned as a DNA vaccine. The sequence of antigen encoding genes was evaluated from the genebank and their epitopes were evaluated to design of primer for the synthetic chimeric gene and was amplified by PCR. Subcloning of a multipartal chimeric gene in eukaryotic expression vector was performed to make a DNA vaccine and finally the protein was purified by nickel chromatography and evaluated by Western blotting.
Results: The immunoblotting results of the expression of SICL chimeric protein indicated the presence of a 76 kDa band in the form of insoluble particles after 12 hours induction. Purification of the recombinant protein using His tag sequencee and confirmation of the purified protein with the recombinant protein specific antibody demonstrated the accuracy of the protein expression.
Discussion & Conclusion: Protein expression, purification and verify by western blotting showed that this recombinant chimeric protein (SICL) can be expressed in eukaryotic host.
Keywords: Vaccine DNA, Enterotoxigenic Escherichia coli, Enterohemorrhagic Escherichia coli, Chimeric protein
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