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:: Volume 20, Number 3 (Iranian South Medical Journal 2017) ::
Iran South Med J 2017, 20(3): 245-256 Back to browse issues page
Cloning and Study of Expression of Helicobacter Pylori FlaAGene in Eukaryotic System
Motaram Sadeghi1, Abbas Doosti *2
1- Biotechnology Research Center, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran
2- Biotechnology Research Center, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran , abbasdoosti@yahoo.com
Abstract:   (612 Views)

Background: Helicobacter pylori is the most common bacterium causing chronic infections worldwide.Expression of flagella and bacterial motility are very important incolonization and virulence. FlaA geneis one of theflagellin-encoding genes that play the key role in the colonization and bacterial motility and it has a significant impact in immunization. The aim of this study wasto design, construction and the evaluation of the Helicobacter pylori flaA gene expression in eukaryotic cells.

MaterialandMethods: In this experimental study, genomic DNA was purified from the Helicobacter pylori standard strain and flaA gene was amplified and isolated by PCR method with use of the specific primers. Then, this gene was cloned into pTZ vector by T/A cloning technique. In order to flaA gene expression and generation of final construct, the flaA gene was removed from pTZ plasmid and sub-cloned into the pcDNA3.1 (-) expression vector. The pcDNA3.1 (-)-flaA construct was transformed into CHO cells by electroporation, and flaA eukaryotic gene expression was studied on SDS-PAGE.

Results: The results showed that flaA gene PCR product was cloned into pTZ vector and amplified in Escherichia coli TOP10F strain. Also the enzymatic digestion and sequencing showed that the pcDNA3.1 (-)-flaA was performed. Finally, the evaluation of the Helicobacter pyloriflaA gene expression in CHO cells showed that the generated gene construct can expressed the flaA product in eukaryotic system, successfully.

Conclusion: Given that the pcDNA3.1 (-)-flaA as a final construct is able to express the flaA protein of the Helicobacterpylori in animal cells. Flagellin protein is one of the important antigens of the bacteriumSo we can properly say that gene pcDNA3.1(-)-flaA consteuct is an appropriate candidate for use in the field of gene vaccines against Helicobacter pylori and in future researches can be used to check the immunization in laboratory animals. 

Keywords: Helicobacter pylori, flaA gene, Cloning, Electroporation
Full-Text [PDF 792 kb]   (180 Downloads)    
Type of Study: Original | Subject: Biochemistry. Cell Biology and Genetics
Received: 2017/07/5 | Accepted: 2017/07/5 | Published: 2017/07/5
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Sadeghi M, Doosti A. Cloning and Study of Expression of Helicobacter Pylori FlaAGene in Eukaryotic System . Iran South Med J. 2017; 20 (3) :245-256
URL: http://ismj.bpums.ac.ir/article-1-876-en.html
Volume 20, Number 3 (Iranian South Medical Journal 2017) Back to browse issues page
دانشگاه علوم پزشکی بوشهر، طب جنوب ISMJ

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