Objective: Leishmaniasis is usually treated by chemotherapy; however, toxicity, resistance and high-cost limits the use of the chemical drugs. Leishmania eukaryotic initiation factor (LeIF) protein acts the same as interleukin (IL)-12 and reduces the secretion of IL-4 in lymph node cells of mice infected with L major. The aim of this study was cloning of the gene encoding LeIF antigen into eukaryotic expression plasmid pEGFP-N1.
Methods: DNA was extracted from Iranian strain of the L major (MRHO/IR/75/ER) promastigotes. The full-length sequence of LeIF was amplified with Pfu DNA polymerase using a specific primer. The amplified LeIF was cloned into a pJET1.2/blunt vector. Then this fragment was digested with HindIII and EcoRI and was subcloned into the pEGFP-N1 vector. Confirmation of the cloning was done by colony polymerase chain reaction (PCR).
Results: Leishmania eukaryotic initiation factor gene was successfully cloned and subcloned into pJET1.2 and pEGFP-N1 plasmids, respectively. The results of colony PCR, restriction analysis and sequencing confirmed them.
Conclusion: We cloned LeIF gene which could be expressed in eukaryotic cells in vivo and could be used as a vaccine candidate against leishmaniasis in future studies.