水稻幼胚电激转化及转基因植株再生(英文)

Gene Transfer into Young Embryos via Electroporation and Regeneration of Plantlets in Rice

幼胚的遗传转化对研究植物胚胎发育相关基因的表达与调控具有重要意义 ,也为植物遗传改良提供新的技术。本研究借助一种自制的特殊装置 ,采用电激法将GFP基因转入 2 - 3天水稻幼胚 ,得到瞬时表达 ,4- 6天水稻幼胚经电激后再生了植株 ,并在愈伤组织阶段及R0植株中检测到GFP荧光的转基因植株 ,从而建立了水稻幼胚的遗传转化实验系统。在电容为 5 0 0 μF、电压为 30 0V/cm ,浓度为 10 0 μg/mL的条件下 ,幼胚GFP的电激转化频率可达 35 %。在pH 5 .8的电激缓冲液中 ,最高转化频率可达 40 %。在三种不同的启动子实验中 ,以Ubi启动子的转化频率最高。

Green fluorescent protein ( GFP ) gene was expressed transiently in 2-3 d old rice embryos by electroporation with the aid of a specially designed loading net. Under suitable conditions (500 μF capacitance, 300 V/cm Voltage, 100 μg/mL plasmid DNA), the percentage of embryos expressing GFP was up to 35%. The highest electroporation efficiency (40%) was obtained at pH 5.8 of the electroporation buffer. The GFP gene driven by the Ubi promoter produced the highest efficiency. Thus, on the basis of optimizing electroporation conditions, a transformation system has been developed for young embryos in rice. The electroporated 4-6 d old embryos regenerated plantlets under the controlled cultural conditions. Fluorescence microscopic observations indicated that GFP gene expressed in their calli and R0 plantlets.