超声波介导铁蛋白基因转化苹果的研究

Ultrasonic Wave-mediated Transformation of Apple with Ferritin Gene

以嘎拉苹果的叶片为转化受体,应用超声波法将菜豆铁蛋白基因导入受体中.实验表明,以0.5 W/cm2的声强、25℃、20 min超声波处理的转化效果最佳,抗性愈伤组织的比例高达24%.抗性愈伤组织经诱导获得了16个苹果抗性植株,有11株表现出GUS阳性,阳性比率为2.75%.经PCR检测,11株GUS阳性植株中有6株能扩增出目的条带;而Southern blotting实验表明,菜豆铁蛋白基因仅在2个苹果转基因株系的基因组中实现了完整插入,并且杂交的信号较强;进一步RT-PCR检测实验显示,外源菜豆铁蛋白基因在上述2株转基因植株中并未转录,发生了基因沉默.

Bean ferrritin gene was transformed into the leaves of the apple variety ‘Gala’ apple as the recipients through ultrasonic wave meditation. It was shown that while ultrasonic wave was applied at 0.5 W/ cm^2 and 25℃ for twenty minutes the most effective transformation occurred and the percentage of the resistant calluses reached as high as 24%. The resistant calluses were induced to form sixteen resistant plants,eleven of the resistant plants presented GUS positive,and the percentage of GUS positive resistant plants was 2.75%. of the eleven GUS positive plants. There were six plants from which target bands could be amplified by PCR and bean ferritin gene was tested by Southern blotting to be completely integrated into only two transgenic apple plants and presented strong hybridization signal in the two; RT-PCR analysis showed that in the abovementioned two plants exogenous bean ferritin genes was not transcribed and gene silencing occurred to it.