title: Cloning of a Recombinant Plasmid Encoding Thiol-Specific Antioxidant Antigen (TSA) Gene of Leishmania majorand Expression in the Chinese Hamster Ovary Cell Line.
abstract: BACKGROUND
  TSA (thiol-specific antioxidant antigen) is the immune-dominant antigen of Leishmania major and is considered to be the most promising candidate molecule for a recombinant or DNA vaccine against leishmaniasis. The aim of the present work was to express a plasmid containing the TSA gene in eukaryotic cells.
  
  
  METHODS
  Genomic DNA was extracted, and the TSA gene was amplified by polymerase chain reaction (PCR). The PCR product was cloned into the pTZ57R/T vector, followed by subcloning into the eukaryotic expression vector pcDNA3 (EcoRI and HindIII sites). The recombinant plasmid was characterised by restriction digest and PCR. Eukaryotic Chinese hamster ovary cells were transfected with the plasmid containing the TSA gene. Expression of the L. major TSA gene was confirmed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting.
  
  
  RESULTS
  The plasmid containing the TSA gene was successfully expressed, as demonstrated by a band of 22.1 kDa on Western blots.
  
  
  CONCLUSIONS
  The plasmid containing the TSA gene can be expressed in a eukaryotic cell line. Thus, the recombinant plasmid may potentially be used as a DNA vaccine in animal models.
authors.affiliation: Department of Parasitology, Faculty of Medical Sciences, Tarbiat Modares University, PO Box 14115-111, Tehran, Iran.
authors.forename: Ghaffarifar
authors.initials: G
authors.lastname: Fatemeh
authors.forename: Tabatabaie
authors.initials: T
authors.lastname: Fatemeh
authors.forename: Sharifi
authors.initials: S
authors.lastname: Zohreh
authors.forename: Dalimiasl
authors.initials: D
authors.lastname: Abdolhosein
authors.forename: Hassan
authors.initials: H
authors.lastname: Mohammad Zahir
authors.forename: Mahdavi
authors.initials: M
authors.lastname: Mehdi