Stable transformation of Pinus radiata embryogenic tissue by Agrobacterium tumefaciens

Cerda, F; Aquea F.; Gebauer M.; Medina, C; Arce Johnson P.

Abstract

Embryogenic cultures from immature zygotic embryos of Pinus radiata seeds were established on semisolid proliferation medium with 2,4-D and BAP. Growing embryogenic masses containing embryonal cells and suspensor cells were subcultured on this media every 2 weeks. After 10 weeks, embryogenic masses (1.5 cm diameter) were transferred to a maturation medium containing ABA. Fully developed somatic embryos were obtained in this medium after 12 weeks. Embryogenic masses were genetically transformed using Agrobacterium tumefaciens. The pB1121 vector containing ?-glucuronidase (uidA) and the neomycin phosphotransferase (nptll) genes was introduced into this tissue. After co-cultivation with Agrobacterium, the embryogenic tissues were transferred to a selection media containing geneticin and carbenicillin. After 1 month of selection, histochemical assays showed extensive GUS positive activity zones in the transformed embryogenic tissues. Under light microscope, blue crystals were seen inside the embryogenic and suspensor cells, and also completely blue somatic embryos were obtained. The uidA gene was also detected by PCR analysis in genomic DNA isolated from transformed embryogenic tissues. These results indicate stable transformation of P. radiata somatic embryogenic tissues using Agrobacterium-mediated transformation.

Más información

Título según WOS: Stable transformation of Pinus radiata embryogenic tissue by Agrobacterium tumefaciens
Título según SCOPUS: Stable transformation of Pinus radiata embryogenic tissue by Agrobacterium tumefaciens
Título de la Revista: PLANT CELL TISSUE AND ORGAN CULTURE
Volumen: 70
Número: 3
Editorial: Springer
Fecha de publicación: 2002
Página de inicio: 251
Página final: 257
Idioma: English
URL: http://link.springer.com/10.1023/A:1016508031151
DOI:

10.1023/A:1016508031151

Notas: ISI, SCOPUS