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Iranian Journal of Microbiology، جلد ۱۵، شماره ۵، صفحات ۶۷۴-۶۸۴
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عنوان انگلیسی Improvement the expression and purification of Loa22: a lipoprotein with OmpA domain from pathogenic Leptospira serovars
چکیده انگلیسی مقاله Background and Objectives: One of the highly conserved outer membrane proteins expressed only by pathogenic Leptospires is Loa22. The study aims is to achieve the optimum conditions for high expression of recombinant Loa22 (rLoa22) protein. Materials and Methods: Complete coding sequence of loa22 gene sub-cloned into a pET32a (+) expression vector. BL21 competent E. coli (pLysS) used as expression host for transformation. The recombinant clones selected on ampicillin plates and subjected to PCR by using pET T7 primers. Then expression conditions optimized by adjusting parameters such as culture media, induction time, temperature, and IPTG concentration. Results: SDS-PAGE analysis showed that high production of rLoa22 protein obtained when post induction incubation, IPTG concentration, and duration of induction were 37ºC, 0.1 M and 5 h in 2×TY medium respectively. The purification of rLoa22 protein under native conditions using Ni-NTA pull-down was optimum in one hour binding at 37°C, five times washing process and elution buffer with a pH 7.4 and a 0.3 M imidazole concentration. Conclusion: The findings of the study led to high production of pure Loa22 protein, which can form the basis for future investigation on the design of rapid diagnostic tests and more effective vaccine candidates for leptospirosis.
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نویسندگان مقاله | Mehdi Gharakhani
Department of Microbiology, Faculty of Basic Sciences, Lahijan Branch, Islamic Azad University, Lahijan, Iran


| Mohammad Faezi Ghasemi
Department of Microbiology, Faculty of Basic Sciences, Lahijan Branch, Islamic Azad University, Lahijan, Iran


| Pejvak Khaki
Department of Microbiology, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran


| Majid Esmaelizad
Department of Research and Development, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran


| Majid Tebianian
Department of Immunology, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran
نشانی اینترنتی https://ijm.tums.ac.ir/index.php/ijm/article/view/4201
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