X射线辐照对发根农杆菌介导芸芥遗传转化的影响

Effects of X-ray Irradiation to Genetic Transformation of Eruca sativa Mill Mediated by Agrobacterium rhizogenes

探讨了X射线辐照对芸芥发状根诱导的影响,为研究辐射对转基因技术的协同作用提供一定的实验依据。以芸芥无菌苗的子叶为材料,通过5—20Gy的X射线辐照和不等浸染时间的联合处理,研究苗龄、预培养时间、辐照剂量及其浸染时间等因素对芸芥发状根诱导的影响作用,并通过聚合链酶式反应(PCR)对所得发状根进行了分子水平的鉴定。15GyX射线辐射能提高芸芥发状根的诱导频率,且有量效关系,其中先浸染后辐照比先辐照后浸染效果更显著;PCR结果也表明,发根农杆菌R1000的rolB基因已成功地被转入并整合到该发状根的基因组中。X射线辐照对芸芥发状根的诱导具有一定的促进作用,且最佳推荐诱导剂量为15Gy或稍多。

To investigate the effects of X-ray irradiation to the hairy root induction of Eruca sativa Mill, and to provide basic experimental data for concerted reaction of irradiation to the Genetic transformation technology. The aseptic cotyledons of Eruca sativa Mill were jointly treated with the X-ray at dose of 5, 10, 15, 20 Gy respectively and the different infected time, then the influence of explant age, pre-culturing time, dose of X-ray and infected time were studied. Furthermore, the hairy roots were testified in molecular level by the polymerase chain reaction（PCR）. The irradiation of X-ray at dose of 15 Gy can improve the frequency of Eruca sativa Mill hairy root induction in a dose-dependent manner. Moreover, the transfor- mation frequency in pre-infection plus irradiation group is higher than that in pre-irradiation plus infection group at the same irradiation dose. In addition, the PCR analysis also demonstrated that rolB gene of T-DNA from Ri plasmid had been integrated into the genome of the transformed roots. The irradiation of X-ray has some positive effects on the hairy root induction of Eruca sativa Mill, and the optimal revulsive dose is 15 Gy or a little bit more.