Cloning and Expression of Com1 and OmpH Genes of Coxiella burnetii in Periplasmic Compartment of Escherichia coli with Purpose of Recombinant Subunit Vaccine Production

Document Type: Original Articles

Authors

1 Department of Microbiology, Faculty of Veterinary Medicine, Lorestan University, Khorramabad, Iran.

2 Department of Animal Science, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran.

3 Department of animal science, Faculty of agriculture, Lorestan university, Khorramabad, Iran

Abstract

Coxiella burnetii is a obligate and gram negative bacteria causing Q fever disease, despite, importance of Q fever, there is no universal vaccine against this disease. Therefore, applying recombinant subunit vaccines which use Com1 and OmpH as immunogenic proteins can be useful. To perform this project, first genes of Com1 and OmpH were amplified by PCR, then, PCR products were purified by DNA precipitation technique. In order to clone, first, both genes along with pET-22b (+) vector were digested by NcoI and XhoI enzymes and then, Com1 and OmpH were ligated in linear vectors by T4 DNA ligase. Recombinant vectors were transformed in BL21 (DE3) strain of Escherichia coli and expression was induced by 1mM Isopropyl β-D-1-thiogalactopyranoside. Expression of Com1 and OmpH was investigated on 12% Sodium dodecyl sulfate polyacrylamide gel electrophoresis, finally, both proteins were purified by Ni-NTA columns and then confirmed by western blotting. Assessment of results on 1% agarose gel shown, PCR amplification, DNA precipitation and digestion of both genes were successfully done. Results of colony PCRs and sequencing revealed that Com1 and OmpH were correctly cloned in pET-22b(+). Finally, the results of expression, purification and western blotting of both proteins shown, BL21 (DE3) strain of Escherichia coli could be able to express Com1 and OmpH proteins. In view of results, it seems, Escherichia coli as affordable and simple host can be applied to express Com1 and OmpH genes. It should be mentioned, products of this project can be examined as recombinant subunit vaccines against Q fever.

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