Vaccine Research

fa Cloning and expression of porA gene as the first step of a vaccine candidate study against Neisseria meningitidis serogroup A infection Original article Original article Introduction: Neisseria meningitidis is a major causative agent of bacterial septicemia and meningitis in human. PorA is a major component of the outer membrane of N. meningitidis and functions as a cationic Porin. This study aimed to clone and determine the expression of PorA as the first step for producing a proper antigen in a vaccine study against N. meningitidis. Methods: An approximately 1200-bp fragment of porA gene was amplified by PCR using                  N. meningitidis serogroup A genomic DNA and then cloned into prokaryotic expression vector pET-28a. The resulting construct (pET28a-porA plasmid) was transformed into competent E.coli BL21 cells for expression of recombinant protein. The proper overexpression of the recombinant protein was verified by SDS-PAGE and Western Blotting. Results: Cloning of porA was confirmed by colony-PCR and enzymatic digestion. The nucleotide sequence homology of the cloned porA gene was 97% , compared to the reference gene (NCBI GenBank accession number AL157959.1). The prokaryotic expression system (pET28a-porA- BL21) could  produce our 45-kDa target recombinant protein, efficiently. Conclusion: The prokaryotic expression system and conditions used in this study provides an applicable method for producing recombinant PorA and possibly many other bacterial outer membrane proteins for future vaccine studies. Neisseria meningitidis, PorA, pET-28a. 60 63 http://vacres.pasteur.ac.ir/browse.php?a_code=A-10-1-30&slc_lang=fa&sid=1 P Afrough afroughparviz9@gmail.com No Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. Microbiology Research Center (MRC), Pasteur Institute of Iran, Tehran, Iran. Pasteur Institute of Iran. Tehran, Iran. A Behrouzi No Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. Microbiology Research Center (MRC), Pasteur Institute of Iran, Tehran, Iran. M Davari No Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. MA Malekan No Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. A Fateh afateh2@gmail.com No Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. Microbiology Research Center (MRC), Pasteur Institute of Iran, Tehran, Iran. F Vaziri farzam_vaziri@yahoo.com No Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. Microbiology Research Center (MRC), Pasteur Institute of Iran, Tehran, Iran. SD Siadat d.siadat@gmail.com Yes Mycobacteriology and Pulmonary Research Department, Pasteur Institute of Iran. Tehran, Iran. Microbiology Research Center (MRC), Pasteur Institute of Iran, Tehran, Iran.