GENERATION OF A GENE CONSTRUCT TO SIPA GENE DELETION OF SALMONELLA TYPHIMURIUM

Salmonella Typhimurium is a negative-gram, non-spore, free capsule, moving
bacteria with Trish Perry flagella. Salmonella is the most common cause of food poisoning. The ability
to enter and survive in host cells is the condition for pathogenic Salmonella species. Proteins of invasive
Salmonella are transferred to the host cells by bacteria. This study was performed for generation of a
gene construct to SipA gene deletion of Salmonella Typhimurium and its cloning in E.Coli bacteria.
Materials & Methods: This laboratory study was conducted in biotechnology research center of Islamic
Azad University Shahrekord Branch from September 2017 to May 2018. In this study, 5' and 3' sequence
of SipA gene was amplified by the specific primers and PCR method. Then, each of these sequences was
cloned by the T/A cloning method in pGEM-Teasy vector and then was transformed into E.Coli bacteria.
Using the PCR method, the part related to each region was amplified and confirmed. The final
confirmation of the produced construct was performed by the Xbal and Xhol enzymes.
Results: The results indicated the successful cloning of the target gene in E.Coli and generation of a
gene construct with a length of the 1520 bp. Also, pET32 vector with a length of 5900 bp was the best
vector for the admission construct.
Conclusion: Based on the results, it seems that by inserting a gene, the damaged gene can be deleted
and it can be used in the future research as a gene vaccine against Salmonellosis. Also, the target gene
can be deleted with electroporation method and transferred into Salmonella Typhimurium, and due to
the similarities between upstream and downstream gene sequences of sipA and Kan genes, homologous
recombination between these two genes can occur and pathogenic genes will be removed