果糖-1，6-二磷酸酯酶在稻瘟病菌致病过程中的功能研究.docxVIP

3 果糖-1，6-二磷酸酯酶在稻瘟病菌致病过程中的功能研究 摘要：稻瘟病菌 (Magnaporthe oryzae)引起的稻瘟病是水稻生产上的主要病害之一，严重威胁着全球粮食产 量。研究已经表明，环腺苷酸(cAMP)信号通路在稻瘟病菌营养生长，附着胞形成，有性生殖，无性繁殖及致病过程 中均发挥重要作用。本研究运用cAMP水解酶高亲和性磷酸二酯酶MoPdeH进行免疫共沉淀(Co-IP)试验，并经酵母双杂 交(Y2H)验证， 发现MoPdeH与MGG_08895编码的蛋白果糖-1，6-二磷酸酯酶(MoFbp1)相互作用。MoFbp1是糖异生途径 的关键酶，广泛存在于有机体内。为了进一步阐明MoPdeH在稻瘟病菌致病过程中的作用机制，本研究通过双酶切法 构建pCX62_MoFBP1敲除载体，将其转入野生型菌株Guy11原生质体中， 运用同源重组的方法获得2个敲除突变体，并 对敲除突变体进行营养生长、产孢量、分生孢子萌发、附着胞形成、致病力等生物学表型进行测定。 关键词：稻瘟病菌，高亲和性磷酸二酯酶MoPdeH，果糖-1,6-二磷酸酯酶，基因敲除 Knockout Analysis of Pathogenicity Related Genes of Magnaporthe oryzae Abstract：Rice blast caused by Magnaporthe oryzae is one of the main diseases in rice production, which seriously threatens the global grain yield. Studies have shown that cAMP signaling pathway plays an important role in vegetative growth, appressorium formation, sexual reproduction, asexual reproduction and pathogenicity of Magnaporthe oryzae. In this study, cAMP hydrolase high affinity phosphodiesterase MoPdeH was used for immunoprecipitation (Co-IP) analysis and further yeast two-hybridization (Y2H) verification. Found that MoPdeH interacted with MGG_08895-encoded protein fructose 1, 6-bisphosphatase (MoFbp1). MoFbp1 is a key enzyme in the gluconeogenic pathway and widely found in organisms. In order to further clarify the mechanism of MoPdeH in the pathogenesis of Magnaporthe oryzae, the knockout vector pCX62_MoFBP1 was constructed by double enzyme digestion and transferred into protoplast of the wild-type strain Guy11. Two knockout mutants were obtained by homologous recombination method. Nutritional growth, sporulation, conidial germination, appressorium formation, pathogenicity and other biological phenotypes of the mutants were determined. Key words: Magnaporthe oryzae, high affinity phosphodiesterase MoPdeH, Fructose-1, 6- bisphosphatase, gene knockout 目录 1前言1 1.1稻瘟病菌概述1 1.2 稻瘟病菌中cAMP信号通路2 1.3 果糖-1, 6-二磷酸酯酶研究进展3 1.4 本研究目的及意义4 2材料与方法5 2.1 材料5 2.1.1供试菌株及植物5 2.1.2培养基5 2.1.2.1 CM培养基5