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首页> 美国卫生研究院文献>Iranian Journal of Biotechnology >Agrobacterium-Mediated Transformation of the Oryza sativa Thaumatin-Like Protein to Canola (R Line Hyola308) for Enhancing Resistance to Sclerotinia sclerotiorum
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Agrobacterium-Mediated Transformation of the Oryza sativa Thaumatin-Like Protein to Canola (R Line Hyola308) for Enhancing Resistance to Sclerotinia sclerotiorum

机译：农杆菌介导的水稻稻草甜蛋白样蛋白向油菜的转化（R系Hyola308）以增强对核盘菌的抵抗力

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>Background: Canola is an agro-economically oilseed crop. Yield loss due to fungal disease of stem rot caused by Sclerotinia sclerotiorum is a serious problem in canola cultivation. Thaumatin-like proteins are large groups of the pathogenesis-related proteins which provide resistance to the fungal infection in response to invading pathogens and play a key role in plant defense system. >Objectives: Transformation of the rice tlp into canola via Agrobacterium-mediated transformation and evaluation of the antifungal activity of the expressed TLP in the transgenic events on the S. sclerotiorum growth was subject to investigation. >Materials and methods: The canola (R line Hyola308) was used for transformation experiment. The vector, pBITLPRA1, was used for the stable transformation. The PCR and southern blotting techniques were used to confirm transgene’s presence in the transgenic canola events. Antifungal activity of transgenic plants was evaluated by the radial diffusion and spore germination assays. T2 transgenic plants were evaluated by the intact leaf inoculation method in greenhouse assay. >Results: In this study, pBITLPRA1 construct containing tlp gene was introduced into canola and the transformed plants were verified by PCR. The glucanase activity of tlp gene in T0 generation was measured and transgenic plants with high activity were assessed by Southern blot analysis to confirm the copy number of the gene. Also, antifungal activity of the single copy T0 transgenic plants against Sclerotinia sclerotiorum was evaluated by radial diffusion and spore germination assays. In greenhouse assay, evaluation of T2 transgenic plants by the intact leaf inoculation method demonstrated that following the infection with S. sclerotiorum, there was a significant reduction in the lesion’s diameter in transgenic lines compared to the non-transgenic ones. >Conclusions: These results revealed that expression of TLP has an inhibitory effect against fungus compared to non-transgenic plants both in vitro and in vivo (i.e., greenhouse condition). These transgenic lines could be used as the additional sources of disease resistance for canola breeding program.
机译：>背景：双低油菜籽是一种经济上经济的油料作物。油菜菌核盘菌引起的茎腐真菌病引起的产量损失是油菜栽培中的严重问题。索马甜蛋白样蛋白是与发病机理相关的蛋白的一大类，它们对入侵的病原体提供对真菌感染的抗性，并在植物防御系统中发挥关键作用。 >目的：尚需调查通过农杆菌介导的转化将水稻tlp转化为双低油菜籽，并评估转基因菌核菌中STP的抗真菌活性。 >材料和方法：将双低油菜籽（R系Hyola308）用于转化实验。使用载体pBITLPRA1进行稳定转化。 PCR和Southern印迹技术用于确认转基因油菜籽事件中转基因的存在。通过径向扩散和孢子萌发试验评估了转基因植物的抗真菌活性。通过完整的叶接种方法在温室试验中评估T2转基因植物。 >结果：在这项研究中，将含有tlp基因的pBITLPRA1构建体导入油菜中，并通过PCR验证了转化的植物。测量t0代中tlp基因的葡聚糖酶活性，并通过Southern印迹分析评估具有高活性的转基因植物，以确认该基因的拷贝数。同样，通过径向扩散和孢子萌发试验评估了单拷贝T0转基因植物对菌核菌的抗真菌活性。在温室试验中，通过完整的叶接种方法对T2转基因植物的评估表明，在感染了核盘菌之后，与非转基因品系相比，病原体直径明显减小。 >结论：这些结果表明，与非转基因植物相比，TLP的表达在体外和体内均具有抑制真菌的作用（即温室条件）。这些转基因品系可用作油菜育种计划的其他抗病源。

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