Subcloning and Expression of Leishmania major (MRHO/IR/75/ER) P4 Gene

Leishmania major p4 gene is localized in the endoplasmic reticulum at the intracellular amastigote stage. We expressed this gene for possible future vaccine preparation, drug target studies, and leishmaniasis serodiagnosis test. The Leishmania major (MRHO/IR/75/ER) p4 gene, which had been subcloned into the pQE-30 expression vector, was induced by IPTG. Recombinant protein was confirmed by SDS-PAGE followed by a double diffusion and western blot using an anti His-tag antibody or human antibody. Production of protein with approximately 35 kDa molecular weight in E. coli M15 transformed cells was confirmed using SDS-PAGE, which reacted with antibodies of leishmaniasis serum using double diffusion and western blot tests and either for anti His-tag antibody using western blot. We have expressed the Iranian L. major p4 gene successfully and are ready to continue the research for vaccine production. Positive results from the western blot and double diffusion test suggest the hypothesis of using this Ag in diagnostic tests. We used the pQE-30 plasmid as an expression vector, which has high quantity of expressed p4 protein.