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小麦抵抗叶锈菌侵染早期的防卫反应机制初探

标题: 小麦抵抗叶锈菌侵染早期的防卫反应机制初探
英文标题: Research on the Mechanism of Wheat Defense Responses at Early Infection Stage of Puccinia Triticina
作者: 王亦飞
出版时间: 2010-01-01
所在大学: 河北农业大学
关键词: 小麦,叶锈菌,细胞间隙液,防卫反应,β-1,3-葡聚糖酶
英文关键词: Wheat (Triticum aestvum L),Puccinia triticina,Intercellular washing fluids (IWF),Defense response,β-1,3-glucanase
论文级别: 硕士
学位: 学位论文
导师: 王冬梅
专业: 生物化学与分子生物学
提交时间: 2010
摘要: 本试验以小麦(Triticum aestivum L.)品种洛夫林10和叶锈菌(Puccinia triticina)生理小种165与260为研究对象,洛夫林10与小种165构成亲和组合,与260构成不亲和组合,对不同亲和性组合中叶锈菌早期侵染结构的形成过程进行了研究.在接菌后2h、4h、6h统计不同亲和性组合叶锈菌的芽管长度;在接种后4h、6h、8h统计各组合中接菌叶段上附着胞形成数;在接种后8h、10h、12h统计各组合中接菌叶段上成功侵染点的个数,并对这些数据进行了方差分析.结果表明:在接菌后4h,孢子萌发芽管的长度在两个组合间达到了极显著差异水平;6h和8h各叶段上附着胞个数也存在差异,不亲和组合叶段上附着胞个数比亲和组合少的多,达到极显著差异;在接种后8h-12h不亲和组合中各叶段上侵染点个数明显少于亲和组合,差异达到极显著水平.初步证明了在不同亲和性组合间锈菌侵染结构的发育程度存在很大差异.为了进一步论证这种差异存在的原因,我们设计试验初步探讨了叶锈菌生理小种260对小麦叶片抗病性的诱导.给洛夫林10小麦先接种260,之后4h用水冲掉该菌,然后再接种165,发现165的发育和侵染受到了抑制,表明接种260有可能诱发了洛夫林10对亲和小种的抗性反应,而且这种抗性反应直接对叶锈菌的生长发育产生了抑制作用.结合本实验室的前期工作,我们认为在这种早期抗病反应诱导过程中,β-1,3-葡聚糖酶起到了很重要的作用,随后我们对接种叶锈菌后的小麦叶片IWF进行提取,并通过western-blotting技术对其中β-1,3-葡聚糖酶进行检测,发现在接种后4h,不亲和组合中该酶的表达量已经很高,而在亲和组合与对照中相对较低.并且对接菌叶片中该酶的活性进行了测定,发现其在不亲和组合中4h活性很高.而且对该酶作用于叶锈菌芽管的产物用DNS法检测,发现该产物是一种还原糖.以上试验初步证明质外体中的β-1,3-葡聚糖酶在抑制锈菌孢子的发育及诱导小麦抗叶锈菌侵染的早期防卫反应过程中可能发挥重要作用.综上所述,在小麦与叶锈菌互作过程中,不同亲和性组合间叶锈菌早期侵染结构的形成存在显著差异,即不亲和性组合中叶锈菌夏孢子萌发芽管的生长以及气孔下泡囊的产生均受到不同程度的抑制,且这种抑制作用有可能与寄主表达的早期抗病性反应-β-1,3-葡聚糖酶的高表达有关.本研究为进一步探讨小麦与叶锈菌互作的早期识别机制及小麦抗叶锈菌侵染的早期防卫机制奠定了基础.
英文摘要: In this paper, it set wheat (Triticum aestvum L) Lovrin 10 and Puccinia triticina physiological race 165 as compatible combinations and Lovrin 10 and physiological race 260 as incompatible combinations, and studied the formation of the early-stage infection structure of Puccinia triticina in different compatible combinations. The germination rates of Puccinia triticina spores of different compatible combinations and their germ tube lengths 2h, 4h, and 6h after inoculation and the number of infection points 8h, 10h and 12h later were recorded and calculated by statistic analysis. The results showed that significant differences were observed 4-6h after inoculation; 8-12h later, the number of infection points in incompatible combinations was significantly less than that in compatible combinations and the differences were extremely significant. These preliminarily proved that the developments of infection structures of different compatible combinations differed greatly.To further confirm the reason of these differences, the induction of disease resistance of Puccinia triticina physiological races to wheat leaves was studied. It first inoculated physiological race 260 in Lovrin 10, and then washed it down 4 hours later and inoculated 165. The development and infection of 165 was inhibited. It showed that inoculating 260 might induce resistance of Lovrin 10 to compatible race and this resistance could directly inhibit the development of Puccinia triticina.Based on previous studies, it is believed thatβ-1,3-glucanase played an important role in this early-stage resistance induction. The intercellular washing fluids (IWF) of leaves inoculated Puccinia triticina were extracted and theβ-1,3-glucanase in them was tested by western-blotting technology. It was found that the content of this enzyme was really high 4 hours after inoculation in incompatible combinations, which was relatively lower in the compatible combinations and in the control combinations. The activity of this enzyme was tested and proved quite high 4 hours after inoculation in incompatible combinations. The product of this enzyme to puccinia triticinia germ tube was also tested by DNS method and it proved that this product is a reducing sugar. According to the tests above,β-1,3-glucanase might played an important role in induction of the early-stage defense response of wheat to Puccinia triticina. In summary, in the interaction of wheat and Puccinia triticina, the formation of the early-stage infection structure of Puccinia triticina in different compatible combinations differed greatly; in incompatible combinations the growths of the germination and the germ tube of the Puccinia triticina and the formation of the stoma vesicle was inhibited by different levels, which might be related to the early-stage resistance of the host-the high content ofβ-1,3-glucanase. This research can provide a basis to further study of the early identification mechanism of the interaction of wheat and Puccinia triticina and the early defense mechanism of wheat to Puccinia triticina.