Journal of Pure and Applied MicrobiologyVol. 6 No. 1

Cloning and Expression of Brucella abortus Omp19 an Immunogenic Minor Outer Membrane Protein

Fatemeh Farahi1,2*, Esmaeil Asli¹, Ashraf Mohabati Mobarez², Nima Khoramabadi² and Hanieh Aghababa²

¹Department of Microbiology, Islamic Azad University, Karaj Branch, Karaj, Iran. ²Department of Bacteriology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.

Received on 12 May 2011 and accepted on 15 June 2011

 

ABSTRACT

Brucellae species are important bacterial zoonotic pathogens which cause brucellosis, an infection with a worldwide distribution. Prevention and diagnosis of infected cases are challenging researchers. Surface proteins are valuable antigens as they are most exposed to the host immune system. Many of outer membrane proteins are expected to be capable of eliciting immune response; one of them is an 18kDa immunogenic protein (Omp19). The aim of present study is producing recombinant Omp19 in a way to prevent brucellosis. Using Gene Runner software to design primers. Omp19 amplified by PCR method. After purification, pJET1.2 and pET28a (+) were used as cloning and expression vectors respectively. Escherichia coli DH5a had the role of unexpression host with usual aim of cloning. BamHI and HindIII were applied as restriction enzymes and E. coli BL21 (DE3) as expression host. Recombinant Omp19 devoid of other Brucella antigens will allow determination of their potential protective activity against brucellosis. Vaccination against animal brucellosis is usually performed by using living attenuated Brucella strains. These vaccines have numerous disadvantages due to basic infections in both animals and mankind. They also elicit antibodies against the Lipopolysaccharide (LPS) which interfere in the differential diagnosis between vaccinated and infected animals. So, accessing to the method or technique to improve new vaccines is an aim in brucellosis investigation. Production of recombinant 18kDa outer membrane lipoprotein of Brucella abortus (Omp19) gained a new vision for preventional researches. Sequencing results of the cloned plasmid vector confirmed the cloning procedure. Gene fragment was subsequently subcloned in expression system pET28a(+). Expression of recombinant protein was induced by adding 1mM IPTG to the growing culture of OD 0.6. Cloning and expression of recombinant 18kDa outer membrane lipoprotein of Brucella abortus (Omp19) allow the evaluation of the potential protective activity and the possibility for the development of subunit vaccines in further investigations.

Keywords : Brucellosis, Brucella abortus, Omp19, Cloning, Expression.