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Kenjirou Ozawa, Yuhya Wakasa, Yuko Ogo, Kouki Matsuo, Hiroyuki Kawahigashi, Fumio Takaiwa, Development of an Efficient Agrobacterium-Mediated Gene Targeting System for Rice and Analysis of Rice Knockouts Lacking Granule-Bound Starch Synthase (Waxy) and β1,2-Xylosyltransferase, Plant and Cell Physiology, Volume 53, Issue 4, April 2012, Pages 755–761, https://doi.org/10.1093/pcp/pcs016
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Abstract
We have developed a high-frequency method for Agrobacterium-mediated gene targeting by combining an efficient transformation system using rice suspension-cultured calli and a positive/negative selection system. Compared with the conventional transformation system using calli on solid medium, transformation using suspension-cultured calli resulted in a 5- to 10-fold increase in the number of resistant calli per weight of starting material after positive/negative selection. Homologous recombination occurred in about 1.5% of the positive/negative selected calli. To evaluate the efficacy of our method, we show in this report that knockout rice plants containing either a disrupted Waxy (granule-bound starch synthase) or a disrupted Xyl (β1,2-xylosyltransferase) gene can be easily obtained by homologous recombination. Study of gene function using homologous recombination in higher plants can now be considered routine work as a direct result of this technical advance.