稻曲病菌优势真菌病毒对寄主菌致病力分化机制研究

Differentiation of pathogenicity on ustilaginoidea virens strains as one of key factors, which manipulates screening and identification resistant rice materials to rice false smut in the experiments, mycoviruses as one of important biological factors hosting in ustilaginoidea virens, it was a pity how to regulate the development and pathogenicity differentiation of its host have still been poorly understood yet. Small RNAs originated from different ustilaginoidea virens strains will be isolated and sequenced by deep sequencing, thus different mycoviruses as well as the relative genomes will be obtained and verified through assembling all sequences from RT-PCR, flanking sequences from 5'-RACE and 3'-RACE, respectively. Therefore, genetic diversity and structure of mycoviruses in ustilaginoidea virens in China will be further characterized; simultaneously the effect on biological characterization in ustilaginoidea virens by dominant mycoviruses will be elucidated. Under agrobacterium-mediated transformation, the relative fragments within 10kb preliminary mapped are responsible for pathogenicity of ustilaginoidea virens through its mutants with GFP gene insertion, when as the targets dominant mycoviruses how to regulate pathogenicity differentiation in host fungi strains will be displayed, as well as the molecular mechanism in the study. Take overall, there are expanding basic knowledge for contributing to resistance breeding and also developing novel fungicides origin from mycoviruses to rice false smut on rice production.

稻曲病菌致病力分化是抗性育种中抗稻曲病材料筛选和鉴定的关键，真菌病毒是稻曲病菌中常见的生物因子，是否参与其生长发育和致病力分化的研究未见报道。本研究利用小分子RNAs深度测序技术，挖掘稻曲病菌的新真菌病毒，通过RT-PCR和旁侧序列RACE技术验证组装基因组，研究稻曲病菌真菌病毒的遗传多样性和优势种群，阐明优势种群对稻曲病菌生物学特性的影响；同时结合农杆菌介导遗传转化获得稻曲病菌突变体库进行致病相关基因的初步定位，研究真菌病毒对稻曲病菌致病力分化的影响和分子调控机制，为稻曲病抗病育种和真菌病毒源杀菌剂的挖掘提供理论依据。

小分子RNAs测序证实了稻曲病菌体内真菌病毒存在遗传多样性，明确了稻曲病菌真菌病毒优势种群为Ustilaginoidea virens partitivirus和Ustilaginoidea virens RNA virus，并建立相应病毒RT-PCR检测方法。生物测定分析表明携带优势病毒寄主菌株系DQ1112、DQ1121以及不携带优势病毒寄主菌株系△DQ1112、△DQ1121在生长速率、产孢量、稻曲球大小和发病时间存在差异。克隆了Ustilaginoidea virens partitivirus的 CP，RDRP和3个未知功能的hypothetical protein基因分别进行克隆，并对其原核表达；发现纯化的CP蛋白对分生孢子和菌丝具有明显的影响，导致致病性明显降低；以CP蛋白为真菌病毒和寄主真菌之间的互作研究对象，发现△DQ1112和成功表达的CP菌株DQ1112-CP在菌丝生长速率、产孢量方面存在差异，致病力分析表明， virus和病毒的CP基因对供试菌株致病力存在相似的影响，推测CP基因可能与供试菌株相关致病基因一定存在互作；酵母双杂交证实CP基因与前期定位的致病相关基因片段没有发生互作，但是对其编码基因进行突变和回补，其菌株的致病力存在比较明显的影响，表明CP基因可能参与其他致病相关基因致病机理，有待进一步研究。