title: Genetic transformation and regeneration of rubber tree (Hevea brasiliensis Muell. Arg) transgenic plants with a constitutive version of an anti-oxidative stress superoxide dismutase gene.
abstract: Agrobacterium tumefaciens-mediated genetic transformation and the regeneration of transgenic plants was achieved in Hevea brasiliensis. Immature anther-derived calli were used to develop transgenic plants. These calli were co-cultured with A. tumefaciens harboring a plasmid vector containing the H. brasiliensis superoxide dismutase gene (HbSOD) under the control of the CaMV 35S promoter. The beta-glucuronidase gene (uidA) was used for screening and the neomycin phosphotransferase gene (nptII) was used for selection of the transformed calli. Factors such as co-cultivation time, co-cultivation media and kanamycin concentration were assessed to establish optimal conditions for the selection of transformed callus lines. Transformed calli surviving on medium containing 300 mg l(-1) kanamycin showed a strong GUS-positive reaction. Somatic embryos were then regenerated from these transgenic calli on MS2 medium containing 2.0 mg l(-1) spermine and 0.1 mg l(-1) abscisic acid. Mature embryos were germinated and developed into plantlets on MS4 medium supplemented with 0.2 mg l(-1) gibberellic acid, 0.2 mg l(-1) kinetin (KIN) and 0.1 mg l(-1) indole-3-acetic acid. A transformation frequency of 4% was achieved. The morphology of the transgenic plants was similar to that of untransformed plants. Histochemical GUS assay revealed the expression of the uidA gene in embryos as well as leaves of transgenic plants. The presence of the uidA, nptII and HbSOD genes in the Hevea genome was confirmed by polymerase chain reaction amplification and genomic Southern blot hybridization analyses.
authors.affiliation: Biotechnology Division, Rubber Research Institute of India, Ministry of Commerce and Industry, Kottayam-686 009, Kerala, India.
authors.forename: R
authors.initials: R
authors.lastname: Jayashree
authors.forename: K
authors.initials: K
authors.lastname: Rekha
authors.forename: P
authors.initials: P
authors.lastname: Venkatachalam
authors.forename: S L
authors.initials: SL
authors.lastname: Uratsu
authors.forename: A M
authors.initials: AM
authors.lastname: Dandekar
authors.forename: P
authors.initials: P
authors.lastname: Kumari Jayasree
authors.forename: R G
authors.initials: RG
authors.lastname: Kala
authors.forename: P
authors.initials: P
authors.lastname: Priya
authors.forename: S
authors.initials: S
authors.lastname: Sushma Kumari
authors.forename: S
authors.initials: S
authors.lastname: Sobha
authors.forename: M P
authors.initials: MP
authors.lastname: Ashokan
authors.forename: M R
authors.initials: MR
authors.lastname: Sethuraj
authors.forename: A
authors.initials: A
authors.lastname: Thulaseedharan
mesh_terms: D001483:Base Sequence; D017931:DNA Primers; D005822:Genetic Vectors; D028482:Hevea / Q000196:embryology / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D007612:Kanamycin / Q000494:pharmacology; D018384:Oxidative Stress; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D012038:Regeneration; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013482:Superoxide Dismutase / Q000235:genetics*; D014170:Transformation, Genetic / Q000187:drug effects; D014197:Trees / Q000201:enzymology / Q000235:genetics / Q000502:physiology
chemical_list: D017931:DNA Primers; D012333:RNA, Messenger; D007612:Kanamycin; D013482:Superoxide Dismutase