Efficient Agrobacterium - mediated transformation and the usefulness of a synthetic GFP reporter gene in leading varieties of Japonica rice

Abstract

We have established an efficient Agrobacterium - mediated transformation method in some leading varieties of Japonica rice (Oryza sativa, L.), including Koshihikari. Scutellum calli were induced from mature seeds on our revised medium, KA1, with high frequencies (50 to 70%) and were used for coculture with Agrobacterium tumefaciens EHA101 which carries binary vector harboring either β glucuronidase (GUS) gene or synthetic green fluorescent protein (sGFP(S65T)) gene driven by CaMV 35S promoter. Scutellum calli at 3 weeks old were highly efficient for the regeneration of transformants. The transformation efficiencies ranged from 15 to 34% in seven leading varieties of nonglutinous rice. The presence of the foreign genes in the genome was confirmed by southern blot analysis, and the expression of sGFP(S65T) gene was detected in several tissues of transformants with bright fluorescent signals under a fluorescent microscopy. The present study demonstrates the usefullness of sGFP(S65T) gene as a reporter in transformed rice plants.