دوره 10، شماره 3 - ( 6-1401 )
جلد 10 شماره 3 صفحات 136-125 |
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Bozorgmehr F, Zeinoddini M, Rasouli Jazi H R. Evaluation of the anti-Salmonella Aptamer using Bioinformatic Tools and ELAA Method. Res Mol Med (RMM) 2022; 10 (3) :125-136
URL: http://rmm.mazums.ac.ir/article-1-472-fa.html
URL: http://rmm.mazums.ac.ir/article-1-472-fa.html
Evaluation of the anti-Salmonella Aptamer using Bioinformatic Tools and ELAA Method. Research in Molecular Medicine. 1401; 10 (3) :125-136
چکیده: (1550 مشاهده)
Background: Identifying and measuring food pathogens and environmental samples require accurate and high-sensitivity and specific methods. In addition, Salmonella is a zoonotic bacteria that can directly cause human disease. The aim of this work is to develop anti-Salmonella aptamer using bioinformatic tools and enzyme-linked aptamer assay (ELAA) for assessment of affinity and identifying Salmonella.
Methods: The aptamer sequence is chosen for attach to the surface protein of Salmonella (OmpA). In following, the second and the tertiary structures of aptamer are achieved, using the UNAFOLD and ROSETTA servers. The structure of OmpA was simulated in two methods: ab initio (using I-TASSER online server) and homology modeling (using MODELLER software and molecular dynamics simulation). In the next step, the binding of aptamer-protein was assessed via HDOCK software. Then, aptamer and biotin-labeled primers for amplification of specific sequence were synthesized. Finally, the performance of aptamer to detect Salmonella was assessed by streptavidin conjugated with horseradish peroxidase (SA-HRP) using the ELAA method.
Results: The results of this study showed that the aptamer, which binds to OmpA, is acceptable with a -283.3 docking score. The superiority of the selected aptamer to random aptamers was also reviewed and approved. After assuring the validation of an aptamer with bioinformatic analysis, the aptamer will be cloned in the pTZ57R plasmid, and then it is used as a template in PCR. After the optimization of PCR and ELAA, the performance of aptamer is successfully carried out for identifying Salmonella.
Conclusion: The designed aptasensor can be used for future investigations to detection of Salmonella.
Methods: The aptamer sequence is chosen for attach to the surface protein of Salmonella (OmpA). In following, the second and the tertiary structures of aptamer are achieved, using the UNAFOLD and ROSETTA servers. The structure of OmpA was simulated in two methods: ab initio (using I-TASSER online server) and homology modeling (using MODELLER software and molecular dynamics simulation). In the next step, the binding of aptamer-protein was assessed via HDOCK software. Then, aptamer and biotin-labeled primers for amplification of specific sequence were synthesized. Finally, the performance of aptamer to detect Salmonella was assessed by streptavidin conjugated with horseradish peroxidase (SA-HRP) using the ELAA method.
Results: The results of this study showed that the aptamer, which binds to OmpA, is acceptable with a -283.3 docking score. The superiority of the selected aptamer to random aptamers was also reviewed and approved. After assuring the validation of an aptamer with bioinformatic analysis, the aptamer will be cloned in the pTZ57R plasmid, and then it is used as a template in PCR. After the optimization of PCR and ELAA, the performance of aptamer is successfully carried out for identifying Salmonella.
Conclusion: The designed aptasensor can be used for future investigations to detection of Salmonella.
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