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Cloning and expression of Brucella cyclic beta-1, 2 glucan transporter gene (cgt)

Bandehpour Mojgan نويسنده , Abdali Narges نويسنده , Sadeghi Farzaneh نويسنده , Parivar Kazem نويسنده , Sharifnia Zarrin نويسنده , Ryahi Hossein نويسنده , Haghighi Asl Ali نويسنده , Kazemi Bahram نويسنده Kavakeb P

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Background: 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. Materials and methods: 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 patientיs serum. Results: The PCR product was cloned in pTZ57R plasmid via T/A cloning method. Recombinant plasmid was digested by BamHI 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 patientיs serum and detected by HRP conjugated anti human antibody. Conclusion: 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.

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