Pathogenicity Identification And Protoplast Preparation Of Rhizoctonia Solani And Resistance Identification Of Rice

Rice sheath blight caused by Rhizoctonia solani is one of the most important diseases on cultivated rice worldwide. Unfortunately, Pathogenesis mechanism and research at molecular level was not well studied till now, due to its complicated pathogenesis mechanism. Developing a genetic transfomation system for the plant pathogenic fungus R. solani and Constructing its mutagenesis library have great significance on pathogenesis mechanism researched, as well as studying the pathogenicity genes. A highly efficient method for preparation and regeneration of protoplasts is crucial for molecular genetic study in this fungus.In this study, the virulent strains of GD-118from Guangdong Province was choosed to explore the protoplast preparation and regeneration conditions. And a series of experiments were carried out on transformation according to the genetic transformation system of Magnaporthe oryzae. In addition, a total of240rice varieties and100of rice sheath blight fungus strains from Zhejiang province and Jiangxi province were studied on the resistance and pathogenicity. The results are as follows:1. The pathogenic difference with different strainsA total of100rice sheath blight fungus strains from Zhejiang province and Jiangxi province were studied on their pathogenicity differentiation, using five different resis-tance rice varieties. Statistical analysis showed the pathogenicity of different strains was significantly different. For example, Y-144, Y-12, Y-22, Y-116showed a strong pathogenicity, while Y-70, Y-17, Y-81, Y-18showed a weaker pathogenicity.2. The resistance difference with different rice varietiesA total of224rice varieties from Zhejiang province were studied by field test, resistance identification and statistical analysis, using five pathogenicity difference standard strains, C30, GD-118, E67, YN-7and YN-3. Our experiment found several strong resistance rice varieties, such as Z7334, G07-76, while some varieties were more susceptible to the disease than the highly susceptible Lemont. 3. The most efficient and easiest method for preparing protoplastsDifferent cell wall degrading enzymes and its combinations were tested. The results showed that R. solani cell wall was efficiently digested with of the enzyme mixture of15mg/mL Glucanex and10mg/mL Lywallzyme.3-3.5x107protoplasts were produced from1gram mycelium by the treatment. Furthermore, different virulence of Rice sheath blight Strains E67, C30, YN-7and YN-3could be digested effectly.4.Rice sheath blight fungus protoplast transformationIn this study, PEG-mediated transformation system of Rhizoctonia solani wasn’t really established according to transformation system of Magnaporthe grisea. We used20μg/mL hygromycinB as the selection concentrations and transformed plas-mid PCB1003, pTHPR1into rice sheath blight fungus protoplast. We could obtain transformants, but these transformants couldn’t be regenerated, and lost hygromycin B resistance.