摘要
应用农杆菌介导法首次向白桦 (Betula platy phyllaSuk .)基因组中导入抗虫基因 [蜘蛛杀虫肽与苏云金杆菌杀虫毒蛋白基因 (Bt基因 )C肽序列的嵌合基因 ]。转化结果表明 :共培养 2d的脱菌及防止腐烂的效果和转化率均高 ,液体共培养及液体除菌优于固体共培养及固体除菌 ;最佳外植体为大叶片 ;侵染 2个月之后产生抗性愈伤组织 ,转化率达到 2 2 %。卡那霉素抗性植株中 ,GUS阳性率为 4 3%。进行杀虫肽和nptⅡ基因的PCR(聚合酶链式反应 )均检测出目的带。转化植株目的基因的PCRSouthern(DNA印记杂交 )杂交呈阳性 ,证明获得了转基因植株。
The paper reported for the first time that transgenic plants of Betula platyphylla had been obtained through mediation with Agrobacterium , in which the chimeric gene of spider insecticidal peptide gene and the C peptide sequence of Bt gene was used as purpose foreign gene for this transformation experiment. The effects of duration of co culture, AS and the type of explants on the Agrobacterium mediated transformation were studied. It showed that 2 d co culture had been suitable for birch transformation to make the explants free of bacteria.The best plant material to be used as explant was the big leaf, on which the frequency of kanamycin resistance calli was 22% (Table 1). By GUS detection, 43 percent of the plants with kanamycin resistance showed positive result (leaves stained blue)(Table 2).Purpose bands were gotten by PCR amplification of npt Ⅱ or spide toxin gene (Figs.1,2). Kanamycin resistant plants also showed purpose band by PCR Southern blotting of the chimeric gene (Fig.3). It meant that insect resistant gene had been transferred and integrated into genome of the birch plants. The insect feeding test showed that the development of Lymantria dispar had been clearly retarded (Table 3).The foreign gene normally expressed in the transgenic plants as resistance to the insect ( Lymantria dispar )to remarkable degree.
出处
《植物生理与分子生物学学报》
CAS
CSCD
2003年第5期380-386,共7页
Journal Of Plant Physiology and Molecular Biology
基金
教育部重点项目 ( 0 2 0 93 )
东北林业大学校基金资助
关键词
白桦
遗传转化
分子生物学检测
抗虫性
Betula platyphylla
genetic transformation
molecular biology detection
insect resistance
