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以枳壳为对象 ,进行了农杆菌介导的上胚轴转化系统建立研究 ,得到了携带柑桔衰退病病毒外壳蛋白基因的枳壳转化植株。研究表明 :枳壳各类外植体中 ,上胚轴是较好的转化材料 ;上胚轴出芽部位主要在形态学上端。以卡那霉素为选择剂 ,外植体水平面放置时 ,选择剂的剂量是 50～ 70mg/ L。转化研究中发现感染液中乙酰丁香酮 10 0μmol/ L的加入促进转化外植体 Gus表达阳性率的提高。延迟 1a的选择培养 ,有助于 Gus阳性芽的增多。所得 Gus阳性无性系经 Southern blot杂交检测 ,证明外源基因已稳定整合到了植物基因组中 ,所得 Gus阳性植株确为携带柑桔衰退病病毒外壳蛋白基因的转化植株
Taking Citrus aurantium as the object, Agrobacterium tumefaciens-mediated hypocotyl transformation system was established and transgenic plants of Citrus aurantium which contained the gene of coat protein of Citrus recessive virus were obtained. The results showed that hypocotyls were the best transformation materials in all kinds of explant of Fructus Aurantii and the budding sites on epicotyls were mainly at the top of morphology. With kanamycin as a selective agent, the level of the explant when placed, the choice of dose is 50 ~ 70mg / L. In the transformation study, it was found that the addition of acetosyringone at a concentration of 100 μmol / L in the culture solution can promote the increase of the positive expression rate of Gus in transformed explants. Delayed selection and culture of 1a, contribute to the increase of Gus-positive buds. The obtained Gus-positive clones were confirmed by Southern blot hybridization, which proved that the exogenous genes were stably integrated into the plant genome. The resulting Gus-positive plants were indeed transformed plants carrying the coat protein gene of citrus recessive virus