【作者】 黄春荣；

【导师】 鲁国东； 王宗华；

【作者基本信息】 福建农林大学 ， 植物病理学， 2010， 硕士

【摘要】 稻瘟病菌（Magnaporthe oryzae）是目前研究丝状真菌的重要模式生物。为了探索水稻与稻瘟病菌互作的分子机理,本文从稻瘟病菌全基因组基因芯片中挑选了三个该菌侵染水稻后表达上调倍数比较高的基因,对它们进行基因敲除,同时对其中的两个基因进行过量表达,分析它们的表型变化,以初步确定它们在稻瘟病菌侵染致病过程中的作用。首先构建了三个基因的敲除载体和MGG₀6834.6与MGG₀3891.6基因的过量表达载体,然后通过原生质体转化,获得基因的缺失突变体和过量表达突变体。突变体表型分析发现,MGG₀3356.6缺失突变体在菌落形态,孢子萌发,致病性方面与野生型没有显著差别,而在疏水性表面上附着胞形成率比野生型菌株KU70低;MGG₀6834.6缺失突变体的表型与对照菌株也没有明显变化,但过量表达突变体的产孢量降低,且附着胞形成提前,而致病性减弱;MGG₀3891.6的缺失突变体的表型与野生型菌株没有显著差别,但过量表达突变体的附着胞形成滞后,且致病性也有所减弱。RT-PCR分析表明,MGG₀6834.6和MGG₀3891.6在野生型菌株中表达量很低。这些结果表明它们可能在稻瘟病菌侵染致病过程中充当一定的作用。同时,我们利用SDS的胁迫来观察细胞壁的完整性,实验结果发现,敲除MGG₀3356.6和MGG₀3891.6基因对细胞的完整性影响不明显,而敲除MGG₀6834.6基因后病菌对SDS抗性较弱,说明细胞完整性减弱,因此MGG₀6834.6可能是细胞完整性的调控因子。更多还原

【Abstract】 Magnaporthe oryzae is one of model organisms in the research of Filamentous fungi. To explore the interaction between Magnaporthe oryzae and its host in molecular level, three genes which high expression during fungal infection were chosen for functional analysis. The knock-out mutans of all three genes and overexpression mutants of two genes were obtained to test their phenotypes in this paper.The gene deletion mutants of MGG₀3356.6, MGG₀6834.6 and MGG₀3891.6, and gene overexpression mutants of MGG₀6834.6 and MGG₀3891.6 were obtained. The results indicated that there is no obvious difference between MGG₀3356.6 deletion mutant and wild type strain in fungal colony morphology, conidial germination and pathogenicity. However, the appressoria formation on the hydrophobic surface was slightly low in the mutant. The deletion of MGG₀6834.6 and MGG₀3891.6 genes did not effect the fungus obviously, while the overexpression mutant of MGG₀6834.6 showed low conidiation, fast appressoria formation but less virulence, and the overexpression mutant of MGG₀3891.6 showed slow appressoria formation, as well as less virulence. The expressions of MGG₀6834.6 and MGG₀3891.6 in wild type were low by RT-PCR analysis. The results suggestted that these two genes may play certain roles during infection of Magnaporthe oryzae.Meanwhile, the cell wall integrity of three deletion mutants was tested by SDS stress. The results showed that the mutant of MGG₀6834.6 but no MGG₀3356.6 and MGG₀3891.6 demonstrated weak resistance to the SDS stress, suggesting that the MGG₀6834.6 gene may be a regulator in the cell wall integrity.更多还原