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首页> 美国卫生研究院文献>Molecules and Cells >Development of an Efficient Agrobacterium-Mediated Transformation System and Production of Herbicide-Resistant Transgenic Plants in Garlic (Allium sativum L.)
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Development of an Efficient Agrobacterium-Mediated Transformation System and Production of Herbicide-Resistant Transgenic Plants in Garlic (Allium sativum L.)

机译：农杆菌介导的高效转化系统的开发和大蒜中抗除草剂的转基因植物的生产

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The genetic improvement of garlic plants (Allium sativum L.) with agronomical beneficial traits is rarely achieved due to the lack of an applicable transformation system. Here, we developed an efficient Agrobacterium-mediated transformation procedure with Danyang, an elite Korean garlic cultivar. Examination of sGFP (synthetic green fluorescence protein) expression revealed that treatment with 2-(N-morpholino) ethanesulfonic acid (MES), L-cysteine and/or dithiothreitol (DTT) gives the highest efficiency in transient gene transfer during Agrobacterium co-cultivation with calli derived from the roots of in vitro plantlets. To increase stable transformation efficiency, a two-step selection was employed on the basis of hygromycin resistance and sGFP expression. Of the hygromycin-resistant calli initially produced, only sGFP-expressing calli were subcultured for selection of transgenic calli. Transgenic plantlets produced from these calli were grown to maturity. The transformation efficiency increased up to 10.6% via our optimized procedure. DNA and RNA gel-blot analysis indicated that transgenic garlic plants stably integrated and expressed the phosphinothricin acetyltransferase (PAT) gene. A herbicide spraying assay demonstrated that transgenic plants of garlic conferred herbicide resistance, whilst non-transgenic plants and weeds died. These results indicate that our transformation system can be efficiently utilized to produce transgenic garlic plants with agronomic benefits.
机译：由于缺乏适用的转化系统，鲜有农艺性状的大蒜植物的遗传改良很少。在这里，我们开发了一种高效的农杆菌介导的转化方法，它与韩国著名的大蒜栽培种Danyang一起使用。对sGFP（合成绿色荧光蛋白）表达的检查表明，在农杆菌共培养过程中，用2-（N-吗啉代）乙磺酸（MES），L-半胱氨酸和/或二硫苏糖醇（DTT）处理可实现最高的瞬时基因转移效率愈伤组织来源于离体苗的根。为了增加稳定的转化效率，基于潮霉素抗性和sGFP表达采用两步选择。在最初产生的抗潮霉素的愈伤组织中，仅将表达sGFP的愈伤组织继代培养以选择转基因愈伤组织。从这些愈伤组织产生的转基因小植株生长到成熟。通过我们的优化程序，转换效率提高了10.6％。 DNA和RNA的凝胶印迹分析表明，转基因大蒜植株稳定整合并表达了膦丝菌素乙酰转移酶（PAT）基因。除草剂喷雾试验表明，大蒜的转基因植物具有抗除草剂的能力，而非转基因植物和杂草则死亡。这些结果表明，我们的转化系统可以有效地用于生产具有农艺优势的转基因大蒜植物。

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