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作 者:魏松红[1] 曹远银[1] 张艳贞[2] 张领兵[2] 王罡[2] 杨家书[1]
机构地区:[1]沈阳农业大学植物免疫研究所,沈阳110161 [2]解放军军需大学植物基因工程研究中心,长春130062
出 处:《中国农业科学》2003年第8期905-909,共5页Scientia Agricultura Sinica
基 金:国家"十五"重点科技攻关专题资助项目 (2 0 0 1BA5 0 9B0 3 3 );"国家植物转基因中试及产业化基地建设"专项 (J99 B 0 0 1);辽宁省博士启动基金资助项目 (2 0 0 2 10 3 9)
摘 要:将携带pti5 vp16基因的pJZ170质粒附着微弹用基因枪轰入辽春 10号、铁春 1号和丰强 3号小麦幼胚 ,2d后检测瞬时表达 ,GUS染色 ,幼胚表面有明显的蓝点 ,单个幼胚蓝点数多达 85个 ;对经ppt2次筛选后的转基因小麦植株进行PCR检测 ,检出 6株阳性植株 ;对部分PCR检测呈阳性的植株进行斑点杂交和Southern印迹杂交检测 ,转基因植株均产生杂交信号。对PCR呈阳性的转基因小麦植株接种小麦白粉病菌 ,接种 7d后 ,病害表型多数为抗病 ;接种 14d后 ,病害表型为抗病、中抗及中感 ,病害严重度明显轻于对照。The immature embryos of wheat cultivars Liaochun10, Tiechun1 and Fengqiang3 were bombarded with gold particles coated with pti5-vp16 by gene gun and disease resistant regenerated plants were obtained. In order to confirm that the plants are genuine transformed ones, a series of molecular tests were conducted. Firstly, transient expression test on embryos two days after bombardment was done. There were many obvious blue spots produced on the surface of bombarded embryos after GUS staining, with the highest blue spot number being 85 on one embryo. Secondly, PCR test was performed with DNA from the regenerated plants obtained after double selection with ppt. 6 plants were found PCR test positive. At last, further verification analysis using dot hybridization and southern blotting techniques was carried out on those PCR positive plants and the strong hybridization signals appeared as expected. All the above tests were uniformly indicated that the disease resistant regenerated plants were true transgenic plants. When inoculated with Blumeria graminis , transgenic wheat plants of PCR positive results were mostly resistant(R) after 7 days, and resistant, moderate resistant(MR), moderate susceptible(MS) at 14 days respectively. The disease severity of them was distinctively lighter than that of control.
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