Primary Infection Suppression Of Blumeria Graminis F. Sp. Tritici Regulated By Pm40Gene And ESTs Analysised In Wheat

Wheat Powdery mildew caused by Blumeria graminis f.sp.tritic(Bgt)i is considered a major disease in the main wheat regions of China. The main strategy for management of Bgt is use of resistant cultivars. But, because of pathogen virulence variance, the resistance of new cultivars was lost soon. It has to be a serious threat to the yield and quality of wheat, and Bgt is biotrophic plant fungal pathogen, cannot be achieved in culture vitro. Therefore, the study of pathogenesis and genetic variability of Bgt is a great significance for preventing wheat powdery mildew. In recent years, with the rapid development of cell biotechnology and molecular biotechnology, which expanded the ideas and methods of studies of vivo parasites, which is difficult to in-depth in the past. New technology can not only study more intuitively the process of interaction between fungi and host, also identified and analysed the functional genes during the process.This study conducted a series of experiments on the cellular level and molecular level, which infect wheat carrying Pm40by pathogen. Observed the process and variety of invasion of Bgt in the primary infection, and constructed SSH-cDNA library, obtained the high quality ESTs from it, and annotated the gene and classified depend on functions, The main results were obtained as followed:1.To reveal the mechanism of Pm40-mediated Bgt resistance in wheat at early infection stage, we carried out the cytological observations using wheat line L699and L658, carrying Pm40gene and presenting immunity to Bgt.CN26was used as the susceptible control,and Nipponbare was used as the affinity control. In wheat-Bgt interaction sites on leaves, we observed the length changes of germ tube of Bgt、papillae formed、hypersensitive reaction, and haustoria formed. Appressorium and germ tube malformation occurred in L699and L658, which was probably caused by the failure of multiple penetration attempts of Bgt, and the length of germ tube was longer in L699and L658than in CN26. Papillae formed at a similar time in challenged epidermal cells in L699and L658and CS. However, the papillae were persisted in longer time in L699and L658than in CN26.For the small number of Bgt that successfully penetrated the epidermal cells of L699and L658, haustorium-dependent hypersensitive reactions were induced to suppress further development of haustoria, arresting onset of powdery mildew. In the affinity interaction sites on leaves, there is no papillae formed, but the length of germ tube was longer in Nipponbare and blank control than in L699and L658and CN26. Therefore, the major mechanism of Pm40-mediated Bgt resistance involves the malformation of appressorium and germ tube、papillae formed and hypersensitive reaction2. Wheat cultivar L699and Bgt was used as the initial materials to construct a wheat SSH-cDNA library using mixed RNA samples isolated from wheat, leaves24h,48h and72h after inoculation. Classified ESTs according to function, after sequencing and bioinformatics analysis, finally, analyzed and discussed the expression of ESTs. In order to lay the foundation of further clarification both the molecular mechanism of wheat-Bgt interaction and of the separation of resistance related genes3.We got483positive clones in SSH-cDNA library, and randomly selected120positive clones sequencing, which produced103high quality ESTs, and got58non-redundant differently expressed ESTs. Functional annotation and category showed that there is12%disease and defense gene,13.7%energy gene,8.6%protein synthesis gene,25.8%primary metabolism gene,10.3%cell structure,8.6%protein destination and storage,6.8%transporters gene,12%signal transduction gene, and1.7%intracellular traffic gene in known functional genes. Meanwhile, there is16.5%differently expressed ESTs of unknown function.4.To discuss and analyse the known functional genes, we infer some kinase and protein involved the process of signal transduction of resistance, which includes MAP kinase, ZmMEK kinase, GTP binding protein, Ran binding protein, etc. Meanwhile, we also found ethylene-responsive protein and MAPK4which is adownstream material of signal pathway of Salicylic acid, and infer the salicylic acid signal pathway and ethylene signaling pathway may exist in the process of resistance, and would have effected in plant resistance.