水稻纹枯病菌菌核形成过程中特异基因的表达与分析

Analysis of differentially expressed genes of Rhizoctonia solani during the sclertium formation

以水稻纹枯病GD-118菌株菌丝形成期的mRNA为驱赶子(driver),以菌核形成期的mRNA为检测子(tester),采用抑制消减杂交(SSH)结合虚拟Northern杂交筛选,构建了水稻纹枯病菌菌核形成过程中特异表达产生的差减cDNA文库;进一步用PCR筛选显示克隆中插入的片段主要分布在200～500bp之间;随机挑取克隆应用虚拟Northern技术以菌丝、菌核cDNA为探针来验证消减有效性,经测序和同源性检测剔除重复序列之后,获得了从菌丝到菌核形成相关的28条特异表达的基因片段;所筛选的特异表达序列主要涉及胁迫响应、转录调控、信号传导、合成代谢等.这为进一步分析水稻纹枯病菌菌核形成过程的相关基因提供了科学线索.

Subtractive cDNA library of differentially expressed genes during the sclertium formation of Rhizoctonia solani,GD-118 strain,was constructed by using suppression subtractive hybridization（SSH） technique combined with virtual Northern blot.The hypha mRNA was used as the driver and the sclertium mRNA was applied as the tester.Further PCR screening showed that the size of the inserts was mainly between 200 to 500 bp.The effectiveness of the subtractive hybridization was verified by virtual Northern blot of colonies randomly selected using hypha,sclertium cDNA sequence as probes.Finally,28 specific sclertium formation-related gene fragments were obtained after sequencing and homology-blasting,which were mainly related to stress response,transcription regulation,signal transduction and metabolism in the pathogen.Results above provide scientific clues to understand the mechanism of the sclertium formation of R.solani.