禾谷镰刀菌Metacaspases基因的功能研究

发布时间：2018-04-12 11:28

本文选题：禾谷镰刀菌 + metacaspase　；参考：《山东农业大学》2017年硕士论文

【摘要】：禾谷镰刀菌(Fusarium graminearum)是导致小麦赤霉病(Fusarium head blight,FHB)的主要致病真菌,也可侵染大麦(Hordeum vulgare)和其他禾谷类作物。小麦赤霉病不仅导致产量的损失,而且会产生一些真菌毒素,如脱氧雪腐镰刀菌烯醇(Deoxynivallenol,DON)。DON毒素是该病原物主要的真菌毒素,DON对人畜健康和安全造成巨大危害,并且是病原菌侵染植物过程中的重要毒力因子。病害控制主要是选择抗病品种和杀真菌剂,但效果不佳。凋亡作为重要的生理学过程之一,是细胞程序性死亡的常见形式。半胱天冬氨酸酶(metacaspase)是一种半胱氨酸依赖的蛋白酶,是原核生物和真菌中参与凋亡过程的重要因子。目前,Metacaspase在植物病原真菌中尚没有相关研究和报道。通过NCBI(National Center for Biotechnology Information)搜索和BLAST比对,在禾谷镰刀菌数据库中发现2个metacaspase基因,分别命名为FgMCA1和FgMCA2。系统发育树和序列比对分析发现:2个FgMCA同时包含metacaspase特征性结构域:P20和P10,2个FgMCA蛋白都属于第一类metacaspase。通过Split-marker PCR方法获得FgMCA基因敲除突变体(?mca1和?mca2)。基因敲除后发现:相对于野生型,2个敲除突变体?mca1和?mca2生长速度和分生孢子产量均有所下降。小麦穗和玉米须接种实验发现,2个突变体的致病性显著下降;与野生型相比,DON毒素产量和TRI基因表达水平也显著降低。在含有过氧化氢的CM培养基上,FgMCA的2个敲除突变体提高了对H2O2的抗性。在H2O2诱导的条件下,2个突变体的凋亡率均下降。对FgMCAs蛋白的亚细胞定位研究发现,2个蛋白都定位于细胞质中。综上所述,F.graminearum基因组中含有2个FgMCA基因,遗传分析发现它们参与调控菌丝生长、分生孢子形成、致病性和DON产量,敲除突变体提高了对H2O2的抗性水平,降低了H2O2胁迫下的细胞凋亡率。
[Abstract]:Fusarium graminearum (Fusarium graminearum) is the leading cause of wheat scab (Fusarium head, blight, FHB) the main pathogenic fungi, can also infect barley (Hordeum vulgare) and other cereal crops. Wheat scab not only lead to loss of output, but also can produce some fungal toxins, such as deoxynivalenol (Deoxynivallenol DON).DON toxin is one of the main pathogens of DON mycotoxin, causing great harm to human and animal health and safety, and is an important virulence factor of pathogen infection in plants. Disease control is mainly the selection of resistant varieties and fungicides, but the effect is poor. Apoptosis as an important physiological process is one of the common forms of. Programmed cell death. Caspase dependent (metacaspase) is a cysteine protease, is an important factor in the apoptosis process in prokaryotes and fungi. At present, Me Tacaspase in plant pathogenic fungi. There is no relevant reports by NCBI (National Center for Biotechnology Information BLAST) search and comparison, 2 metacaspase genes found in Fusarium graminearum database, named FgMCA1 and FgMCA2. phylogenetic tree system and sequence alignment analysis showed that: 2 FgMCA also contains the metacaspase characteristic domains P20 and P10,2 FgMCA proteins belong to the first class of metacaspase. through the Split-marker PCR method to obtain the FgMCA gene knockout mutant (? Mca1? Mca2). Gene knockout found: compared to wild-type, 2 knockout mutants? Mca1 and? Growth rate and spore yield decreased mca2 and wheat. Corn inoculation experiment showed that the pathogenicity of 2 mutants decreased significantly; compared with the wild type, the expression of DON and yield of TRI toxin gene were also reduced significantly. In the presence of peroxide Hydrogen on CM medium, FgMCA 2 knockout mutants enhanced resistance to H2O2. After H2O2 induction, the apoptosis of 2 mutants decreased. To study the subcellular localization of FgMCAs protein found that 2 proteins were localized in the cytoplasm. In summary, F.graminearum genome contains 2 FgMCA gene, genetic analysis showed that they are involved in the regulation of mycelial growth, conidia formation and pathogenicity and DON yield, knockout mutant improves the level of resistance to H2O2, decreasing the apoptosis rate under H2O2 stress.

【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S435.121.45

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