The Composition And Expression Of Carbohydrate-Active Enzyme Genes Of Rhizoctonia Cerealis Transcriptome

Wheat sharp eyespot caused by Rhizoctonia cerealis Van der Hoeven, was a worldwide disease of wheat. In China, wheat sharp eyespot has been an important threat of high and stable yield of wheat. R. cerealis causing wheat sharp eyespot in China belong to the AG-D anastomosis groups of binucleate Rhizoctonia. A lot of researchs on R. cerealis about rapid detections in field, chemical control and biological control, etc. have been done worldwide, but the studies in epidemiology and pathogenesis were lagging far behind other pathogens.Several recent studies have demonstrated that there is a strong relationship between the repertoire of carbohydrate-active enzymes in fungal genomes or transcriptomes and their lifestyle. Fungi with necrotrophic lifestyle have more CAZymes. In fungi or bacteria, the GH, PL, and CE families may involve in degrading plant cell wall, therefore play a role in pathogenicity.The transcriptomes of R. cerealis growing on induction medium were analysed in this study. We found that R. cerealis had a closer relationship with Laccaria bicolor and Coprinopsis cinerea, which were not the plant pathogenic fungi. There were a lot of CAZymes in R. cerealis, especially ones involving in degrading plant cell wall (cellulose, hemicellulose and etc.).From the digital gene expression profiling of R. cerealis on PSA, on induction medium and in planta, we found that there were a high number of different expression CAZyme genes involving in degrading plant cell wall. The gene expression of CAZymes degrading cellulose was lower in planta than that on induction medium, but the gene expression of CAZymes degrading hemicellulose was higher. We suggest that R. cerealis mainly hydrolyzed hemicellulose of wheat cell walls. The expression of CE3(unigene8051), CE12(unigene16042) and PL1(unigene629) was higher expressed in planta relative to that on induction medium. It shows that they may play an improtent role in infection. The expression of some CAZyme genes were validated by real time PCR, we found that the expressed patterns of them were consistent with the expression profiles, but slightly different in expression level at different infection stages.When degrading cereals hemicellulose, GH10xylanases have been shown to be unaffected by the presence of TAXI-like proteinaceous inhibitor, but GH11was inhibited. GH10may be the important pathogenic factor for cereals pathogens. We gained the full-length cDNA of two GH10genes by RACE and bioinformatics method, which provides the basis for studying the role of R. cerealis GH10in infection process.To further study the role of CAZymes in infection process, the kinds of enzymolysis enzyme, the enzymolysis time and the osmotic stabilizer types were optimized, and we gained an efficient and stable system for protoplast preparation. We compared morphology and pathogenicity, found that there was no difference between regenerated strains and original strain in. The system provided a basic for genetic transformation and protoplast fusion of R. cerealis. We also tried the PEG-protoplast and Agrobacterium tumefaciens mediated method to transform R. cerealis, and no transformant was obtained.