MicroRNA调控下的RNAi途径关键基因及NBS-LRR类抗病基因在SMV侵染大豆中的作用研究

发布时间：2018-02-23 04:24

  本文关键词： 大豆花叶病毒 大豆 microRNA 抗病基因 RNAi途径 微阵列分析　出处：《内蒙古大学》2016年博士论文　论文类型：学位论文

【摘要】：植物通过不同层次的防卫反应来抵御外源病毒的侵染。RNA干扰(RNA interference, RNAi)是植物针对病毒侵染的一个重要的应答反应,AGO (Argonaute)和Dicer/DCL (Dicer-like)蛋白是RNAi途径中的关键成分。效应蛋白诱导的免疫反应(Effector triggered immunity, ETI)一般指的是抗病基因(Resistance gene, R)介导的抗病毒机制。迄今为止发现的R蛋白多数属于NBS-LRR家族蛋白。MicroRNA (miRNA)是一类由生物体内源产生的、具有重要调节作用的非编码小RNA分子。近年来发现miRNA在植物的抗病反应中也发挥着很重要的作用。大豆花叶病毒(Soybean mosaic virus, SMV)是一种世界性大豆病害,严重影响着大豆的经济和商品价值。有研究表明miRNA在大豆种粒斑驳形成过程发挥着主要作用,同时发现SMV的侵染影响大豆内源小干扰RNA (small interference RNA, siRNA)的积累水平。但是,目前没有一个系统的研究SMV侵染与大豆miRNA表达调控之间关系的报道。本论文中我们利用microarray技术在敏感大豆品系中筛选到了45个SMV响应的miRNAs。在上调的miRNAs中miR168a、miR403a、miR162b和miR1515a的预测靶标分别为AGO1、AGO2、DCL1和DCL2; miR1507a、miR1507c和miR482a的预测靶标为NBS-LRR家族的10个抗病基因。我们通过双荧光素酶报告系统与RLM-RACE实验证实了上述7个miRNAs对其10个靶标基因表达的调控作用。AGO1、DCL1、DCL2和5个NBS-LRR类抗病基因的表达受SMV侵染的抑制,同时相应miRNAs的丰度上调,并且它们之间的表达在SMV侵染后不同时间点均表现出相反的表达模式。我们的结果表明SMV会通过上调一些miRNAs来抑制RNAi途径相关基因及NBS-LRR家族抗病基因的表达,从而导致敏感大豆品系的感病。
[Abstract]:Plant defense response to foreign virus infection by different levels. RNA interference RNA interference (RNAi) is an important response response to virus infection in plants ago and Dicer/DCL Dicer-like proteins are the key components of the RNAi pathway. The immune response induced by Effector triggered (ETI) is generally referred to as the antiviral mechanism mediated by resistance gene resistance (RIA). Most of the R proteins found so far belong to NBS-LRR family proteins. MicroRNAs are produced from biological sources. Noncoding small RNA molecules with important regulatory functions. In recent years, miRNA has been found to play an important role in plant disease response. Soybean mosaic virus (Soy bean mosaic virus) is a worldwide soybean disease. Some studies have shown that miRNA plays a major role in the formation of soybean seed mottles, and it is also found that the infection of SMV affects the accumulation level of small interference RNAs (siRNAs) in soybean. There is no systematic study on the relationship between SMV infection and the regulation of miRNA expression in soybean. In this paper, we used microarray technique to screen 45 SMV responsive miRNAs in sensitive soybean strains. The predicted targets were AGO1, AGO2, DCL1 and DCL2, respectively, and the predicted targets of miR1507a- miR1507c and miR482a were 10 resistant genes of the NBS-LRR family. We confirmed the regulatory effect of these 7 miRNAs on the expression of their 10 target genes by using double luciferase reporting system and RLM-RACE experiments. The expression of DCL2 and 5 NBS-LRR class resistance genes were inhibited by SMV infection. At the same time, the abundance of miRNAs was upregulated. Our results showed that SMV could inhibit the expression of RNAi pathway related genes and NBS-LRR family resistance genes by up-regulating some miRNAs. Thus, sensitive soybean strains are susceptible to the disease.
【学位授予单位】：内蒙古大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S435.651

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