| A method for single-spore isolation of Exserohilum turcicum was established. The main research contents included the population of physiological races and genetic polymorphism of E. turcicum in northeastern China, and isolates of physiological races of E. turcicum identifying resistance of 11 main varieties of Liaoning. The resualts were as follows:1. A method for single-spore isolation of E. turcicum with general light microscope was appllied in the research. Spores displacing on the corn leaf were transfered to Water Ager firstly, then single-spore of Exserohilum turcicum was pickked with simple homemade needle at low magnification in order to achieve purification. Although the method was slightly sophisticated, it was simple, practical, less contaminating and highly effective. The application of the method is contributive to rapid identification of physiological races and DNA genetic polymorphism of E. turcicum. It is more suitable for other single-spore isolation of the pathogenic fungi with large conidiospores.2. The physiological race of E. turcicum was identified among 113 isolates collected from 28 areas of Liaoning, Jilin and Heilongjiang provinces in 2009 on corn inbred lines with Ht1, Ht2, Ht3 and HtN resistance genes, and its population dynamics was investigated. The results showed that 12 races were identified among these fungal isolates, including physiological races of 0,1,2,3, N,12,13, 1N,2N,123,12N and 123N respectively. Those races not only can infect the maize with single Ht gene, but also that with all kinds of resistance gene, and it means that the race component is in pluralistic. Among these races population, races 0 and 1 were predominant with occurrence frequency of 47.8% and 19.5%. The virulence frequencies against genes Ht1, Ht2, Ht3 and HtN were 37.2%,21.2%,4.4% and 15.9% respectively. The physiological differentiation of E. turcicum was obvious, and their distribution and interspecific varieties became complex. The increasing epidemic of northern corn leaf blight in northeastern China was ascribed to the increasing virulence frequencies of such major races as 0 and 1, and the resistance-losing of the cultivars.3. A suitable Universally Primed PCR (UP-PCR) amplification system was established and optimized in the dNTP, primers, MgCL2, Taq DNA polymerase and DNA template concentration. The number of amplification cycles and melting temperature (Tm) were optimized respectively.7 polymorphic primers were screened out for studying the genetic diversity among 24 isolates of E. turcicum coming from the major corn planting areas in the Northeast of China. A total of 75 bands ranged between 250bp and 2000bp were amplified among the tested isolates, and 72.0% were found to be polymorphic, indicating the high genetic diversity in the population of E. turcicum. The dendrogram based on UP-PCR results revealed that the 24 isolates were clustered into 8 groups at the threshold of genetic similar coefficient of 0.796. All above results indicated that there was abundant genetic diversity among E. turcicum colony in the Northeast of China; there was no distinct relationship between the fungal genetic diversity and their geographic distribution; there was some correlation between fungal genetic diversity and physiological races but not simple genetic pedigree-races relationship between them. UP-PCR analysis revealed the phylogenetic relationships and difference of the tested isolates of E. turcicum, which provided a new method for analyzing the genetic evolution and the physiological differentiation of E. turcicum.4. The resistance of 11 major maize cultivars in Liaoning province to 14 different physiological races of E. turcicum was identiffied by inoculating niblet seed with inocula into leaf whorl. There was obvious discrepancy in different physiological races to the cultivars. Identification of disease resistance and pedigree analysis showed that the cultivars resistance to different physiological races had correlation with the consanguinity of the cultivars. |
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