EVALUATION OF A COTYLEDONARY NODE REGENERATION SYSTEM FOR AGROBACTERIUM-MEDIATED TRANSFORMATION OF PEA (PISUM-SATIVUM L)

A rapid regeneration system was used for studies of Agrobacterium-medi ated transformation in Pisum sativum L. Cotyledonary node explants wer e inoculated with Agrobacterium tumefaciens strains containing binary vectors carrying genes for nopaline synthase (NOS), beta-glucuronidase (GUS), and neomycin phosphotransferase (NPTII) and placed on selectio n medium containing either 75 or 150 mg/liter kanamycin. A GUS encodin g gene (uidA) containing an intron was used to monitor gene expression from 6 to 21 days postinoculation. GUS activity could be observed 6 d ays after inoculation in the area of the explant in which regeneration occurred. Regenerating tissue containing transformed cells was observ ed in explants on selection medium 21 days postinoculation. Using this system, a single transgenic plant was obtained. Progeny of this plant , which contained two T-DNA inserts, demonstrated segregation for the inserts and for expression of the NOS gene in the selfed R1 progeny. N PTII activity was observed in the R2 generation, indicating inheritanc e and expression of the foreign DNA over at least two generations. Att empts to repeat this procedure were unsuccessful.