Cloning and expression of Brucella cyclic beta-1, 2 glucan transporter gene [cgt]

ABSTRACT

Brucellosis is an important cosmopolitan infection disease caused by organisms belonging to the genus Brucella. The cgt gene [cyclic beta-1, 2 glucan transporter gene] is a virulent factor in Brucella genus. The present study was conducted with the aim of cloning and expression of Brucella cgt gene. Brucella melitensis cgt gene was amplified from extracted chromosomal DNA by PCR, then PCR product was cloned into pTZ57R and subcloned into pGEMEX-1 expression vector, then expressed in JM109 E. coli strain. Recombinant protein was confirmed by western blot analysis using patient's serum. The PCR product was cloned in pTZ57R plasmid via T/A cloning method. Recombinant plasmid was digested by Band HI and SacI restriction enzymes, the released band was purified and subcloned into pGEMEX-1 expression vector. Then, sample cells were lysed using lyses buffer and sonicated, then electrophoresed on SDS-PAGE. Protein bands were transferred on nitrocellulose membrane and reacted by patient's serum and detected by FIRP conjugated-anti human antibody. We cloned and expressed Brucella abortus cyclic beta-1, 2-glucan transporter gene [cgt] which is an important agent in brucellosis. Using cgt gene mutant may be an effective way for inhibiting or decreasing the pathogenicity of bacteria