农杆菌介导的高羊茅高效遗传转化和转基因植株再生(英文)  

Highly Efficient Transformation and Plant Regeneration of Tall Fescue Mediated by Agrobacterium tumefaciens

作　　者：胡张华[1] 陈锦清[1] 吴关庭[1] 金卫[1] 郎春秀[1] 黄锐之[1] 王伏林[1] 刘智宏[1] 陈笑芸[1] 

机构地区：[1]浙江省农业科学院病毒学与生物技术研究所,杭州310021

出　　处：《植物生理与分子生物学学报》2005年第2期149-159,共11页Journal Of Plant Physiology and Molecular Biology

基　　金：浙江绿洲农业股份有限公司资助~~

摘　　要：用带有质粒pDBA121(含hpt基因和bar基因)的农杆菌EHA 105转化高羊茅(Festucaarundinacea Schreb.)胚性悬浮细胞,建立了可重复的、高效的农杆菌介导的高羊茅遗传转化系统.商业用的除草剂Basta直接用于转化细胞的筛选.基因型、受体材料的类型、培养基成分和筛选剂影响农杆菌介导的转化频率.悬浮细胞的农杆菌转化效率为每克悬浮细胞再生2.85～10.9株转基因植株,大大高于基因枪法的高羊茅转化效率(2～5株).经PCR分析和Southern杂交检测表明,bar基因已整合进入高羊茅基因组,转基因植株Basta喷洒试验表明bar基因已成功地实现高水平的表达.此转化系统的建立为高效地将外源有用基因导入高羊茅并高效稳定地表达奠定了基础.An efficient and reproducible system hasbeen developed for the production of transgenic plantsin tall fescue (Festuca arundinacea Schreb.) using A.tumefaciens-mediated transformation. Two-months-oldsuspension cultures served as excellent explants fortransformation. The explants were inoculated with anA. tumefaciens strain EHA105 carrying a plasmidpDBA121 containing genes for hygromycinphosphotransferase (hpt) and phosphinotricinacetyltransferase (bar). The commercial herbicideBasta was used as a selective agent. Inclusion ofacetosyringone (ACS) 20 mg/L in the co-culture me-dium led to an increase in transformation efficiency.The efficiency of transformation was highly depen-dent on the genotype, the explant, the culture mediumand selective agent used. Tall fescue transformationefficiency is 2.85–10.9 plants per gram fresh weight(FW) of suspension cultures. This is much higher thanthe corresponding values reported before (2–5 plants).So far more than 300 transgenic plants have beenobtained, the fertility of some transgenic plants haddecreased. Stable integration and high expression ofthe transgenes were confirmed by PCR analysis andSouthern blot hybridization or herbicide Basta spray-ing test.

关 键 词：高羊茅 根癌农杆菌 遗传转化 胚性悬浮培 养细胞 转基因植株 

分 类 号：S688.4[农业科学—观赏园艺]