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:: Volume 14, Issue 2 (May,Jun 2012) ::
J Shahrekord Univ Med Sci 2012, 14(2): 64-73 Back to browse issues page
Optimization of expression, extraction & purification of the N-terminal region of ipaD gene in Shigella dysenteriae by proteomics analysis
Mahdi Hesaraki , Mogtaba Saadati , Hosseyin Honari , Gholamreza Olad , Mohamad Heiat , Mokhtar Zare
1Biological Sciences Research Center, Imam Hossein University, Tehran, I.R. Iran; , E-mail:saadati_m@yahoo.com E-mail:saadati_m@yahoo.com
Abstract:   (10780 Views)
Background and aims: Shigella dysenteriae is one of the most important pathogens which in spite of many attempts vaccine preparation, extended researches are still in the way, Transport and surface expression of the invasion plasmid antigens (IpaD proteins) have essential role in the pathogenicity of Shigella spp. IpaD has been one of the most important proteins for Shigella vaccine candidate .Studies have shown that N– terminal region of this protein has a key role in the pathogen city and invasion. This study was done to evaluate the optimization of N-terminal region of Ipad in order to increase the production of recombinant protein. Methods: In this experimental labortary study, desired region of IpaD cloned in vector pET-28a (+). For confirming cloning procedure, standard tests were performed. The effect of IPTG concentration, temperature & induction times on the level of protein expression were evaluated by SDS-PAGE, qualitatively. The gels were evaluated with 2-D gel analysis software (Melanie 7). The recombinant protein was extracted by Urea & eventually purificated with affinity chromatography column. Results: SDS-PAGE analysis showed that approximately the same amount of recombinant protein is expressed at different times, but software analysis proved that the optimized condition for the expression of recombinant protein was in the final concentration of 0.7 mM of IPTG, 37˚C and 3 hours induction. Conclusion: According to the results every protein has its own expression after the homogenization process, and the temperature and the cells induction time length are more effective in the amount of protein production.
Keywords: Invasion plasmid, Recombinant protein, Proteomics, Shigella dysenteriae
Full-Text [PDF 412 kb]   (1352 Downloads)    
Type of Study: Research | Subject: general
Received: 2012/06/2 | Accepted: 2017/09/2 | Published: 2017/09/2
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Hesaraki M, Saadati M, Honari H, Olad G, Heiat M, Zare M. Optimization of expression, extraction & purification of the N-terminal region of ipaD gene in Shigella dysenteriae by proteomics analysis. J Shahrekord Univ Med Sci. 2012; 14 (2) :64-73
URL: http://journal.skums.ac.ir/article-1-1115-en.html


Volume 14, Issue 2 (May,Jun 2012) Back to browse issues page
مجله دانشگاه علوم پزشکی شهرکرد Journal of Shahrekord University of Medical Sciences
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