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Expression of Multi-Epitope Gene Structure Based on Bacterial Genes of Salmonella Typhimurium and Escherichia Coli in BALB/c Mice
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Saeid Mikaeel1    , Abbas Doosti2 , Ali Sharifzadeh3 |
1- PhD Candidate, Department of Biology, Faculty of Basic Sciences, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran.
2- .Professor, Biotechnology Research Center, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran. , bio.gene84@gmail.com
3- Assistant Professor, Department of Micribiology, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran. |
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Abstract: (429 Views) |
Background and Aim: Chitosan nanoparticles are becoming a popular alternative to deliver nucleic acids to tissues for gene transfer. The present study was carried out to design a combined multi-epitope gene construct based on the pathogenic genes of Salmonella typhimurium and Escherichia coli strain O157:H7 and to transfer it to BALB/c mice with the help of chitosan nanoparticles.
Materials and Methods: In this experimental study, pcDNA3.1(+) recombinant vector containing a multi-epitope gene construct was designed and synthesized. E. coli strain TOP10F was transformed with the mentioned vector. Using the plasmid extraction kit, the plasmid was extracted from the transformed bacteria and confirmed. In the next step, this recombinant vector was combined with chitosan nanoparticles, and the characteristics of the complex after production were investigated. Plasmid-containing complexes with or without nanoparticles were injected into the thigh muscle of animals at time intervals of 0, 7, and 14 days. The animal tissue was collected, and the expression of gene structure and cytokines (IFN-γ and IL-10) was analyzed by the reverse transcription method. The spleen tissue of the animals was also examined histopathologically.
Results: The correctness of the synthesis and extraction of the recombinant vector was successfully confirmed. Chitosan nanoparticles and recombinant vector were combined in the ratio (1:1). In animals treated with the recombinant vector, compared to the control group, the expression of the gene construct was successfully confirmed. Examination of cytokines showed that the cellular immune response increased in the challenge groups compared to the control group. However, the structure of the spleen tissue in the group that was transferred with the recombinant vector carrying the gene construct along with chitosan nanoparticles was healthy and normal.
Conclusion: The recombinant vector, after combining with chitosan nanoparticles, was correctly transferred and expressed in the animal tissue, and there was no damage to the spleen pulp. In the future, this multi-epitope gene structure can be used as a vaccine against Salmonella Typhimurium and Escherichia coli.
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| Keywords: Chitosan nanoparticles, Gene transfer, Recombinant vector, Combined multi-epitope gene construct. |
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Type of Study: Original Research |
Subject:
Biotechnology
Received: 2024/02/24 | Accepted: 2024/07/14
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