Abstract
Phytoalexin are low-molecular-weight compounds produced and accumulated after exposure to microorganism attack, elicitor and ultraviolet, and have been suggested to serve as plant antibiotics. Rice (Oryza sativa) produces 14 diterpenoid phytoalexins, including oryzalexins A-F, phytocassanes A-E, oryzalexin S, and momilactones A and B. We have identified 6 diterpene cyclase genes responsible for biosynthesis of these phytoalexins: OsCPS2, 4, OsKSL4, 7, 8, and 10. In this study, we attempted to produce diterpenoid-phytoalexin-deficient rice plants by knock down of OsCPS2/4, OsKSL4, or OsKSL7 to verify the roles of these phytoalexins on disease resistance in rice. We first found one line of rice plant in which Tos17 was inserted in the 3rd intron of OsCPS4 by PCR screening of Tos17 mutant panel. However, the level of the mature OsCPS4 mRNA in the line slightly lower than that in wild type rice, suggesting that Tos17 in the 3rd intron of OsCPS4 was excluded by splicing. This result showed that this line is not a null mutant for OsCPS4. Next, we prepared transgenic rice, in which each RNAi trigger (approximately 100bp) for OsCPS2/4, OsKSL4, and OsKSL7 was expressed under the control of rice ubiquitin promoter. However, specific knock down of the targeted genes were not observed in the transgenic rice plants. We have made again transgenic rice plants in which approximately 200bp trigger sequences were introduced, and will present the transcript levels in the transgenic plants.
