根癌农杆菌介导的山定子遗传转化的研究

RESEARCH OF Agrobacterium tumefaciens MEDIATED GENETIC TRANSFORMATION OF Malus baccata BORKH

以山定子叶片作为根癌农杆菌介导转化的受体,通过比较在卡那霉素筛选培养基上的抗性芽百分率,研究激素配比、乙酰丁香酮、脯氨酸、共培养时间、预培养时间、侵染时间对转化的影响。结果表明:预培养不利于山定子转化;菌液中添加乙酰丁香酮和脯氨酸对转化无明显影响;侵染10min,共培养3d抗性芽百分率最高;转化后叶片再生培养基的最佳激素配比为NAA1.0mg/L+BA2mg/L+TDZ4mg/L。抗性植株经PCR检测,初步证明外源目的基因已整合到山定子基因组中。

Several factors, such as hormone combination, Acetosyringone （AS）, Proline （Pro）, co-culture time, pre-culture time and infection time, which affect Agrobacterium tumefaciens mediated gene transformation of Malus baccata Borkh, were studied by comparing rate of the kanamycin resistant shoot. The results showed that pre-culture of the leaves was not beneficial to transformation. There was no evident to improve transformation by adding AS and Pro into bacterial suspension. The highest rate of the resistant shoot was obtained under the condition of infection for 10 min and co-culture for 3d. The optimal hormone combination of leaf regeneration medium after transformation was NAA 1.0mg/L ＋ BA 2 mg/L ＋ TDZ 4 mg/L. The PCR analysis showed that the target gene was integrated into the genome of Malus baccata Borkh.