公开(公告)号：US20150344897A1

公开(公告)日：2015-12-03

申请号：US14422661

申请日：2012-09-14

申请人： Haibo Wang ,  Fushuang Dong ,  Mengyu Lv ,  Yanmin Zhang ,  Zhiheng Ren ,  Fan Yang ,  Xinchao Liang ,  Wenbo Zuo ,  Xueping Shi ,  Huanhuan Zhang ,  Yiping Gao ,  He Zhao ,  Xian Xu ,  Guozhong Sun ,  Jianfang Chai ,  Yongwei Liu ,  Jinyong Zhu ,  Qiufen Han ,  Qiang Zhang ,  Huijie Ma ,  Zhanwu Wang ,  Junfeng Guan

发明人： Haibo Wang ,  Fushuang Dong ,  Mengyu Lv ,  Yanmin Zhang ,  Zhiheng Ren ,  Fan Yang ,  Xinchao Liang ,  Wenbo Zuo ,  Xueping Shi ,  Huanhuan Zhang ,  Yiping Gao ,  He Zhao ,  Xian Xu ,  Guozhong Sun ,  Jianfang Chai ,  Yongwei Liu ,  Jinyong Zhu ,  Qiufen Han ,  Qiang Zhang ,  Huijie Ma ,  Zhanwu Wang ,  Junfeng Guan

IPC分类号： C12N15/82

CPC分类号： C12N15/8205 ,  C12N15/8207

摘要： The present invention is a method of shoot apical meristem transformation for monocot plant via sufficient and micro wounding (SMW). The technical process includes: expose the apical meristem by removing the coleoptile away when the shoot grows to 0.2-2 cm after 1-2 days of seed germination; make sufficient and micro wounding transformation to the apical meristem by stabbing and brushing for 2-3 times using the SMW brush having 100-5000 bristles which is 4-20 μm in diameter for each one and 0.5-3 mm in exposed length, and dipped with the Agrobacterium tumefaciens containing binary vector harboring exogenous genes; develop the treated meristems directly to normal plants after co-cultivation; promote the plants to develop big spikes and set more seeds; harvest the seeds of T0 plants separately; detect and identify the transformation results in T1 generation which is bred from each individual T0 plant. The advantages of the invention are independent of tissue culture, unlimited in genotype, unnecessary to carry resistant marker, simple and large scale to perform, and applicable to all monocot plants which can set seeds. The transformation efficiencies for wheat, rice and maize using this method are 49%, 66.3%, and 100%, respectively.

摘要翻译： 本发明是通过足够和微伤（SMW）的单子叶植物的茎顶分生组织转化的方法。 技术过程包括：通过在种子发芽1-2天后枝条生长至0.2-2厘米时，通过去除胚芽鞘来揭开顶端分生组织; 使用具有直径为4-20μm，暴露长度为0.5-3mm的直径为4-20μm的100-5000个刷毛的SMW刷，通过刺入和刷刷2-3次来对顶端分生组织进行充分和微小的伤口转化，并浸渍 含有包含外源基因的二元载体的根癌农杆菌; 共培养后将经处理的分生组织直接发育至正常植物; 促进植物发展大穗，种植更多种子; 分别收获T0植物的种子; 检测和识别从每个T0植物繁殖的T1代转化结果。 本发明的优点独立于组织培养，基因型无限，不需携带抗性标记，简单大规模进行，适用于可以设置种子的所有单子叶植物。 使用该方法对小麦，水稻和玉米的转化效率分别为49％，66.3％和100％。