High-efficiency improvement of transgenic torenia flowers by ion beam irradiation

To shorten the time required for breeding and optimize the risk-cost/benefit ratio of genetically modified ornamental plants, we applied heavy-ion beam irradiation to wild-type and genetically modified torenia (Torenia fournieri Lind. CV. ‘Crown Violet’ plants in which petal color and pattern had been modified by controlling two anthocyanin biosynthesis-related genes encoding chalcone synthase (CHS) and dihydroflavonol-4-reductase (DFR). Ion beams of 12C6+ and 20Ne10+ were applied to 11,500 leaf disks from wild type and five transgenic lines, and over 3,200 regenerated flowering plants were then investigated for visible phenotypes. The mutation rate after whole irradiation averaged 10.4%, and the maximum rate in the initial screening was 44.2% (20Ne, 30 Gy). Mutant phenotypes were observed mainly in flowers and showed wide variation in color and shape. Mutation efficiencies for petal color and coloration pattern were higher in transgenic plants than in wild-type plants, while those for petal shape and corolla divergence were almost equivalent in the two plant groups. Mutation spectrums in petal color in transformant-based mutants were obviously wider than those in wild-type plants. Among these mutants, a class B gene-deficient mutant was investigated as a model case for further study to facilitate the control of flower phenotype. Expression of the TfGLO gene was found to be repressed in this line, probably due to dysfunctioning of the upstream signaling. We propose that the combination of genetic engineering and ion beam irradiation greatly facilitates improvement of agrobiological and commercial traits within a short period. We also discuss characteristic changes observed at high frequency in torenia flowers and the mutant-based approach to the identification of useful genes.