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首页> 中文期刊> 《植物生理与分子生物学学报》 >农杆菌介导的高羊茅高效遗传转化和转基因植株再生

农杆菌介导的高羊茅高效遗传转化和转基因植株再生

         
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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 has been developed for the production of transgenic plants in tall fescue (Festuca arundinacea Schreb.) using A.tumefaciens-mediated transformation. Two-months-old suspension cultures served as excellent explants for transformation. The explants were inoculated with an A. tumefaciens strain EHA105 carrying a plasmid pDBA121 containing genes for hygromycin phosphotransferase (hpt) and phosphinotricin acetyltransferase (bar). The commercial herbicide Basta was used as a selective agent. Inclusion of acetosyringone (ACS) 20 mg/L in the co-culture medium led to an increase in transformation efficiency.The efficiency of transformation was highly dependent on the genotype, the explant, the culture medium and selective agent used. Tall fescue transformation efficiency is 2.85-10.9 plants per gram fresh weight (FW) of suspension cultures. This is much higher than the corresponding values reported before (2-5 plants).So far more than 300 transgenic plants have been obtained, the fertility of some transgenic plants had decreased. Stable integration and high expression of the transgenes were confirmed by PCR analysis and Southern blot hybridization or herbicide Basta spraying test.

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