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

Abstract

Coxiella burnetiiis an obligate and gram-negative bacteria causing query fever (Q fever) disease, despite the importance of Q fever, there is no universal vaccine against this disease. Therefore, application of the recombinant subunit vaccines which use Com1 and OmpH as immunogenic proteins can be useful in this regard. To perform the current project, Com1 and OmpH genes were amplified by polymerase chain reaction (PCR) method, then, the PCR products were purified by DNA precipitation technique. In order to clone, first, both genes along with the pET-22b(+) vector were digested by NcoI and XhoI enzymes and then, Com1 and OmpH genes were ligated in linear vectors by T4 DNA ligase. The recombinant vectors were transformed in BL21 (DE3) strain of Escherichia coli and expression was induced by 1 mM Isopropyl β-D-1-thiogalactopyranoside. Expression of Com1 and OmpH was investigated using 12% Sodium dodecyl sulfate polyacrylamide gel electrophoresis. Finally, both proteins were purified by Ni-NTA columns and consequently confirmed by western blotting. The results of assessing 1% agarose gel showed that PCR amplification, DNA precipitation, and digestion of both genes were successfully performed.Theresults of colony PCRs and sequencing revealed that Com1 and OmpH were correctly cloned in pET-22b(+) vector. Finally, the results of expression, purification, and western blotting of both proteins showed thatBL21 (DE3) strain of Escherichia colicould be able to express Com1 and OmpH proteins. Based on the collected data, it seems that Escherichia coli as an affordable and simple host can be applied to express Com1 and OmpH genes. It should be mentioned that products of the present project can be examined as recombinant subunit vaccines against Q fever.