摩西管柄囊霉（Funneliformis mosseae）抑制连作大豆根腐病发病机理的研究

The study aims to reveal the mechanism that F. mosseae inhibit the root rot in the continuous cropping soybean, and lays the foundation that AM inoculums applied to preventing the root rot of soybean. The varieties of the most representative soybean cultivars in Heilongjiang province that have different resistance to root rot are selected as materials. The inoculated F.mosseae inoculums were obtained after screening the soybean continuous cropping rhizosphere soil and propagated by the members of our research group. The variation of root rot pathogen variety and content in soybean root and rhizosphere are analyzed by Miseq sequencing, PCR-DGGE and qRT-PCR technology. The changes of defensive protein variety and content in the soybean root and rhizosphere are analyzed by LC-MS technology. GC-MS technology is used to analyze the change of poisonous and harmful secondary metabolite variety and content through in the soybean root and rhizosphere. Through pathogenic microorganism flora structure, defensive protein and poisonous metabolite molecular level reveals the mechanism that F. mosseae inhibit the root rot in the continuous cropping soybean. Determining the incidence of different varieties of soybean root rot, the index of biomass, yield and quality, it verify that F. mosseae have an inhibition effect for soybean root rot. Therefore, the study has an important significance to fill the mechanism of bio-control soybean root rot, realizes the goal of bio-control soybean root rot, and revitalization the industry of soybean.

本项研究的主要目的是揭示F. mosseae抑制连作大豆根腐病发病机理，为应用AM菌剂防治大豆根腐病发生奠定理论基础。试验选用黑龙江省具有代表性的不同抗根腐病大豆品种作为试验材料，接种自主筛选的F. mosseae菌剂，采用MiSeq测序、PCR-DGGE和qRT-PCR相结合方法分析大豆根系及根际土壤中根腐病致病菌种类及含量变化，采用LC-MS方法分析大豆根系及根际土壤中防御性蛋白种类及含量变化，采用GC-MS方法分析大豆根系及根际土壤中有毒有害次生代谢产物种类及含量变化，从病原微生物菌群结构、防御蛋白以及有毒代谢物变化等分子水平揭示F. mosseae抑制连作大豆根腐病发病机理，并且通过测定不同品种大豆根腐病发病率、生物量、产质量等指标，验证F. mosseae具有抑制大豆根腐病发生功效。所以本项研究为填补大豆根腐病生防机理，实现根腐病生防目标，振兴我国大豆产业具有重要的意义。

大豆作为重要的经济作物，在农业生产方面占据着重要的地位。但是近年来国内大豆种植面积日益减少，市场需求量不断提高，加速了大豆主产区连作现象的发生，造成大豆根腐病等病害愈来愈严重。丛枝菌根真菌（Arbuscular Mycorrhizal Fungi，AMF）被广泛应用于提高植物抗逆性研究中，但是，目前国内外研究人员鲜有从机理上解释AMF是如何抑制连作大豆根腐病发生以及对产品质量的提升。因此，本课题将F. mosseae接种于连作大豆土壤中，探究AMF减缓大豆根腐病发生情况，旨在揭示F. mosseae抑制连作大豆根腐病发病机理，为AM菌剂防治大豆根腐病奠定理论基础。. 本研究选用黑农48大豆品种作为试验材料，并种植于不同连作年限土壤中。一是采用iTRAQ-LC-MS/MS技术分析F. mosseae对大豆根系蛋白的影响；二是采用高通量测序技术测定F. mosseae对不同连作年限大豆根际土壤细菌的影响；三是利用代谢组学和转录组学联合分析，对大豆根系差异表达基因和差异代谢物进行鉴定；四是利用转录组学和蛋白质组学测序技术筛选F. mosseae和Fusarium. oxysporum互作过程中大豆所产生的差异基因及蛋白，从转录和翻译水平筛选出与抗病和促进大豆生长发育相关的基因及蛋白。. 本试验得出以下四个方面结果：一是采用iTRAQ-LC-MS/MS技术，测定出F. mosseae可以使大豆防御、能量代谢和信号转导相关的蛋白质表达量上调。二是采用高通量测序技术，测定出F. mosseae能够提高土壤有益菌属丰度，改善代谢功能，调节连作土壤理化性质。三是采用代谢组学和转录组学联合分析，测定出14个差异代谢途径和19个差异代谢物质，F. mosseae通过增加抗病基因的表达，进而提高抗逆性酶的活性，达到调控代谢产物。四是采用转录组学和蛋白质组学联合分析，测定出F. mosseae使抗病相关酶上调表达，也可以使与大豆生长发育相关的基因及蛋白上调表达，说明F. mosseae具有增强大豆根系抗病性、缓解连作带来的病害作用。.本课题旨在揭示F. mosseae抑制大豆根腐病发病的机理，填补大豆根腐病生防机理的研究空白，加快实现大豆根腐病生物防治目标，提升黑龙江大豆生产地位，保护非转基因大豆资源，增加农民收入，减少大豆进口量，重振我国大豆产业。