Cloning and expression of PMI1945 involved in iron acquisition as a promising vaccine candidate against Proteus mirabilis

• Main Authors: B Kavehei, M Habibi, S Sari, MR Asadi Karam
• Format: Article
• Language: English
• Published: Pasteur Institute of Iran 2018-06-01
• Series: Vaccine Research
• Subjects: proteus mirabilis; iron scavenger receptors; pmi1945; cloning; expression.
• Online Access: http://vacres.pasteur.ac.ir/article-1-130-en.html

Introduction: Proteus mirabilis is one of the common pathogens of urinary tract infections. Iron scavenger receptors from P. mirabilis are considered as important virulence factors of this strain and have the properties of an ideal vaccine candidate. In this study, the frequency of P. mirabilis iron receptor 1945 (PMI1945) was evaluated in the isolates and then its expression was conducted in pET28a-BL21. Methods: Amplification of PMI1945 was performed by PCR using P. mirabilis isolates genomic DNA. Cloning of PMI1945 gene was done in pET28a-BL21 system. After transformation, the expression of the cloned gene was induced by IPTG. The expression of this protein was then evaluated by SDS-PAGE and Western blot techniques. Results: The frequency of PMI1945 gene in the isolates was 76%. The cloning of PMI1945 gene into pET28a vector was confirmed by electrophoresis, PCR, enzyme digestion and sequencing. The sequencing of the cloned gene showed 100% identity with other sequences of PMI1945 gene in GenBank. SDS-PAGE and Western blot results showed that 77 kDa PMI1945 protein was successfully expressed in BL21 (DE3) host. Conclusion: Cloning and expression of PMI1945 was done as the first step for evaluation of a novel vaccine candidate against UTIs caused by P. mirabilis.