title: Cloning, Transformation and Expression of Human Interferon α2b Gene in Tobacco Plant (Nicotiana tabacum cv. xanthi).
abstract: BACKGROUND
  Molecular farming is the production of important recombinant proteins in transgenic organisms on an agricultural scale. Interferons are proteins with antiviral and antitumor activities and can be used for viral infections and cancers treatments.
  
  
  OBJECTIVE
  This study reports the transformation of INF α2b gene in tobacco plant for the first time in Iran.
  
  
  METHODS
  Interferon α2b gene was amplified by PCR using specific primers containing appropriate restriction enzymes, plant highly expression sequence and Histidine tag sequence. Target sequence was cloned in plant expression vector pCAMBIA1304 and the construct named pCAMINFα. pCAMINFα was transferred to E. coli strain DH5α and plated on LB agar medium containing kanamycin 50 mgl-1. The colonies were confirmed by colony PCR and sequencing. The construct was transferred into Agrobacterium tumefaciens by freeze-thaw method and transformed colonies were confirmed by colony PCR. Tobacco plants (cultivar xanthi) were inoculated with A. tumefaciens strain LBA4404 by leaf disc method. Inoculated explants were cultured on MSII (MS + BAP 1mgl-1 + NAA 0.1 mgl-1) at 28°C and darkness for 48 hours. Then explants were transferred to selection medium containing cephotaxime (250 mgl-1) and hygromycin (15 mgl-1) in a 16/8 (day/night) h photoperiod in growth room with an irradiance of 5000 lux. Transgenic plants were regenerated and transferred to perlite. Genomic DNA was extracted from regenerated plants by Dellaporta method at 5-leaf step and transgenic lines were confirmed by PCR with specific primers. Expression of Interferon α2b gene was confirmed by dot blotting.
  
  
  CONCLUSIONS
  Since no report of interferon alpha production in plants in Iran has been expressed yet, this research could create a field of producing this drug in tobacco, in Iran.
authors.affiliation: Department of Agricultural Biotechnology, University of Agriculture and Natural Resources, Ahwaz, IR Iran ; Department of Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, IR Iran.
authors.forename: Shahrzad
authors.initials: S
authors.lastname: Ahangarzadeh
authors.affiliation: Department of Agricultural Biotechnology, University of Agriculture and Natural Resources, Ahwaz, IR Iran.
authors.forename: Mohammad Hosein
authors.initials: MH
authors.lastname: Daneshvar
authors.affiliation: Department of Agronomy and Plant Breeding, Shahid Chamran University, Ahwaz, IR Iran.
authors.forename: Hamid
authors.initials: H
authors.lastname: Rajabi-Memari
authors.affiliation: Department of Genetics, Shahid Chamran University, Ahwaz, IR Iran.
authors.forename: Hamid
authors.initials: H
authors.lastname: Galehdari
authors.affiliation: Department of Agricultural Biotechnology, University of Agriculture and Natural Resources, Ahwaz, IR Iran.
authors.forename: Khalil
authors.initials: K
authors.lastname: Alamisaied
keywords: Agrobacterium; DNA Transformation Competence; Interferons; Molecular Farming; Tobacco