@2024 Afarand., IRAN
ISSN: 1735-7675 Kowsar Medical Journal 2011;16(1):1-6
ISSN: 1735-7675 Kowsar Medical Journal 2011;16(1):1-6
Cloning and expression of ipaC gene from Shigella dysenteriae
ARTICLE INFO
Article Type
Original ResearchAuthors
Mallaei F. (1 )Saadati M. (* )
Honari H. (1 )
Nazariyan Sh. (2)
Eghtedardoust M. (1 )
Heiat M. (3 )
Hesaraki M. (1 )
(* ) Department of Biological Sciences, Faculty of Sciences, Imam Hossein University, Tehran, Iran
(1 ) Department of Biological Sciences, Faculty of Sciences, Imam Hossein University, Tehran, Iran
(2) Department of Immunology, Faculty of Sciences, Shahed University, Tehran, Iran
(3 ) Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
Correspondence
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Fax:
f.mallaei@yahoo.com
Article History
Received:Accepted:
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ABSTRACT
Aims
Shigellosis is caused by Shigella species. Considering the high frequency of morbidity and mortality reports and antibiotics resistance, designing and producing a vaccine against this disease is one of the goals of World Health Organization. Invasion Plasmid Antigen especially IpaC and IpaB are the major Shigella virulence agents and are also vaccine candidates. The aim of this study was to express ipaC gene which is of the virulence factors in order to investigate its immunogenicity.
Materials & Methods In this study, ipaC gene was obtained from NCBI gene bank and primers were designed. After genome extraction from Shigella dysenteriae, it was used as template for PCR amplification. The amplified ipaC gene by PCR was cloned into pTZ57R and sub-cloned into the expression vector pET-28a(+). E. coli BL21(DE3)plysS was transformed by recombinant vector pET-28a(+)/ipaC and expression of the recombinant ipaC was investigated by IPTG induction and SDS-PAGE electrophoresis.
Results Cloning was confirmed by PCR, enzyme digestion and sequencing. In addition, the recombinant protein was expressed by IPTG induction. Protein expression was confirmed by Ni–NTA column, Western-Blotting analysis and SDS-PAGE electrophoresis.
Conclusion Cloning, sub-cloning and expression of ipaC gene are confirmed in the present study. Therefore, this recombinant protein can be used for production of a recombinant vaccine against Shigella dysenteriae in future studies, after immunogenicity assay.
Materials & Methods In this study, ipaC gene was obtained from NCBI gene bank and primers were designed. After genome extraction from Shigella dysenteriae, it was used as template for PCR amplification. The amplified ipaC gene by PCR was cloned into pTZ57R and sub-cloned into the expression vector pET-28a(+). E. coli BL21(DE3)plysS was transformed by recombinant vector pET-28a(+)/ipaC and expression of the recombinant ipaC was investigated by IPTG induction and SDS-PAGE electrophoresis.
Results Cloning was confirmed by PCR, enzyme digestion and sequencing. In addition, the recombinant protein was expressed by IPTG induction. Protein expression was confirmed by Ni–NTA column, Western-Blotting analysis and SDS-PAGE electrophoresis.
Conclusion Cloning, sub-cloning and expression of ipaC gene are confirmed in the present study. Therefore, this recombinant protein can be used for production of a recombinant vaccine against Shigella dysenteriae in future studies, after immunogenicity assay.
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