[Construction of MyD88-Pseudomonas aeruginosa epitope DNA vaccine and its expression in eukaryotic cells].

ABSTRACT

AIM: To construct the MyD88-Pseudomonas aeruginosa epitope vaccine and study its expression in eukaryotic cells. METHODS: To design and synthesize an epigene containing three B cell epitopes of OprF and one foreign "promiscuous" T cell epitope by overlapping extension PCR. tPA signal encoding sequence was amplified by PCR and then it was inserted into the 5' terminus of the epigene to construct tPA-OprF. tPA-OprF and MyD88 were cloned into the expression vector pIRES and the recombinant plasmid pIRES-tPAOprF-MyD88 was constructed. The recombinant plasmid was transfected into COS-7 cells by electroporation. The expression protein of tPA-OprF and MyD88 was detected by Western blot. RESULTS: The recombinant plasmid pIRES-tPA-OprF-MyD88 was successfully constructed. Western blot analysis indicated the tPA-OprF fusion protein was expressed in supertanant of COS-7 cells and MyD88 protein in COS-7 cells. CONCLUSION: The recombinant plasmid pIRES-tPA-OprF-MyD88 has been successfully constructed and tPA-OprF and MyD88 protein can be highly expressed in transfected cells. It may be used as a potential candidate of preventive vaccine of pseudomonas aeruginosa.