In planta Agrobacterium-Mediated Transformation of Rice

The floral-dip transformation, the simplest technique, is no requirement of tissue culture procedure, and can directly transfer the interest gene into plant reproductive cells. It has been successfully applied to various plant species. In this study, the optimal conditions of a floral-dip method for production of transgenic rice variety RD41 were explored. The simple and effective inoculation medium was composed of Murashige and Skoog (MS) medium, 5% sucrose, 44 nmol/L benzylaminopurine, and 0.075% surfactant Tween-20 with pH 5.7. The transformation efficiencies of Agrobacterium tumefaciens strains AGL1 and EHA105 were compared with the Agrobacterium density at OD600 = 0.8-1.0 and the co-cultivation at 25 °C for 48 h. A. tumefaciens strain EHA105 gave slightly higher transformation efficiency than AGL1, with statistically non-significant difference. The floral-drop transformation using the optimal floral-dip conditions showed higher transformation efficiency than the floral-dip method, but the dropped flowers turned brown and died within 2 d. Production of transgenic rice variety RD41 by the floral-dip method was achieved using A. tumefaciens strain EHA105 with the optimal conditions. Screening for the gusA gene by PCR using the gusA specific primers in the T0 lines, there were 4 transgenic lines from 286 T0 lines (1.4% transformation efficiency). However, histochemical glucuronidase (GUS) assay demonstrated that only three of four transgenic lines exhibited gusA expression. These results indicated that floral-dip transformation is a potential tool for production of the transgenic rice, which can be used for molecular breeding via genetic engineering in the future.