Highly efficient visual selection of transgenic rice plants using green fluorescent protein or anthocyanin synthetic genes

Visual selection enables transformation in plants that are sensitive to the stress of antibiotic selection. Furthermore, we proposed a method of isolating transformed cells in which the gene of interest is expressed at high level using visual selection. However, visual selection was inefficient and laborious because of the technical difficulties involved in clonal propagation of transformed calli selected using GFP (9.4-20.5% in the model japonica rice variety Nipponbare in our previous report). Very recently, we have discovered that an indica model cultivar, Kasalath, is very highly competent for Agrobacterium-mediated transformation compared to Nipponbare, suggesting that visual selection would be achieved more efficiently in Kasalath than in Nipponbare. We confirmed that the number of transformed cells emitting green fluorescence in Agrobacterium co-cultivated callus of Kasalath under no antibiotic pressure was >50-fold higher than the number obtained in Nipponbare. We succeeded in clonal propagation of GFP emitting cells at a frequency of 62.2-85% in Kasalath. Moreover, we showed that anthocyanin, that is an intrinsic pigmentation and thus might be more acceptable to consumers, can be used as an intrinsic marker in visual selection of rice. Thus, we propose that combination of the use of cultivar which has high transformation competency and visual selection system could enhance efficient production of transgenic rice expressing foreign gene product at higher level.