title: Cloning and Constructing a Plasmid Encoding Leishmania Eukaryotic Initiation Factor Gene of Leishmania major Fused with Green Fluorescent Protein Gene as a Vaccine Candidate.
abstract: OBJECTIVE
  Leishmaniasis is usually treated with chemotherapy; however, toxicity, resistance and high-cost limit use of the chemical drugs.Leishmaniaeukaryotic 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 withLeishmaniamajor. 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
  Leishmaniaeukaryotic 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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Medical Parasitology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: N
authors.initials: N
authors.lastname: Maspi
authors.affiliation: Department of Medical Parasitology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran, E-mail: ghafarif@modares.ac.ir.
authors.forename: F
authors.initials: F
authors.lastname: Ghaffarifar
authors.affiliation: Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Z
authors.initials: Z
authors.lastname: Sharifi
authors.affiliation: Department of Medical Parasitology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: A
authors.initials: A
authors.lastname: Dalimi
keywords: Cloning; DNA vaccine; Leishmania eukaryotic initiation factor; Leishmania major