The Analysis Of Pathogeny For The Virulence Gene Of Verticillium Dahliae In Xingjiang

Cotton Verticillium Wilt is one of the most serious diseases of cotton in China On cotton yield and quality, even in the world, and it is a major problem to the sustainable development of the cotton industry in China. Due to the variation of V. dahliae, the resistance of cotton varieties to V. dahliae is easy to loss. Therefore, clarification the difference in virulence among the isolates of V. dahliae and causes of variation is valuable for the control of Verticillium wilt.Subject of the present study was typical 88 Verticillium fungus purified separated from Shihezi, Shawan, Wusu, Bole, Korla and other places. Partial isolates were characterized by morphology, molecular identification, ITS sequence analysis and virulence differentiation analysis, and transcriptome analysis of strain which had different virulence and genetic relationship. The main conclusions showed as fllowing:1. According to cultural characteristics on potato dextrose agar(PDA), 88 V. dahliae isolates were divided into four culture types(A-D). Type A(66,72.7%) produced massive microsclerotia, and it produced a white circle at the colony edge; Type B(8,9.0%) produced a few microsclerotia distributed at the edge of colony which formed a black circle; Type C(14,15.9%) produced massive microsclerotia, and it produced black mycelia at the colony edge; Type D(2,2.3%) did not produce microsclerotia under room temperature, and it had dense mycelium without pinch. The cultural characteristics of isolates from the southern had all kinds of culture, and thenorthern had three types(A、C、D).2. Based on the results of the pathogenicity tests, 24 V. dahliae isolates were clustered into three groups, which showed strong(average disease index varied 44.00~50.00), moderate(average DI varied 26.00~35.00) and weak(average DI varied 15.67~20.00) pathogenicity on cotton, respectively. Moderate virulent isolates distributed dominantly in China(50%). Moderate virulent isolates was dominant groups in Shihezi. The defoliating and nondefoliating pathotypes of V. dahliae isolates were identification by specific PCR amplification. Results indicated that 2 isolates were nondefoliating pathotypes for 7 strong virulent isolates, 5 isolates were defoliating pathotypes for 12 moderate virulent isolates, and 1 isolates were defoliating pathotypes for 5 wake virulent isolates.3. According to conducted a RNA-Seq analysis of V. dahliae that 3 different pathotypes, The SHZvd-4 and SHZvd-9 in pathogenicity difference was bigger, the correlation was higher. Analyses of these 1010 differentially expressed genes, including up-regulation gene 610 and down-regulation gene 400. The up-regulation gene mainly gathered in protein hydrolysis of biologicl process, molecular of cell component, hydrolytic enzyme and oxidase of Molecular Functional. The results showed that there was strong mutation bias: C:G to T:A, which is introduced by repeat induced point mutations(RIP). The V. dahliae genome contains many transposable elements, suggesting that they can escape damage by RIP. Finally, About 100 differentially expressed genes were selected.4. The relative expression of NPR1 gene and PR-1, PDF1.2 gene induced by NPR1 gene with different virulent isolates treatment at different times were detected by Real-time PCR. The results showed that the relative expression of NPR1 reached the maximum after treatment 24 h. The relative expression of Arabidopsis NPR1 trend is similarity after three kinds of different virulence of Verticillium treatment, but the relative expression of Arabidopsis NPR1 was high aftern the strong virulent isolates treatment, and the relative expression of Arabidopsis NPR1 was low aftern the weak virulent isolates treatment. PDF1.2 gene did not affect the invasion by pathogens, PDF1.2 gene expression more stable. The results of showed that the relative expression of PR1 reached the maximum after treatment 36 h. The relative expression of Arabidopsis PR1 trend is similarity after three kinds of different virulence of Verticillium treatment, but the relative expression of Arabidopsis PR1 was high aftern the strong virulent isolates treatment, and the relative expression of Arabidopsis PR1 was low aftern the weak virulent isolates treatment. Pathogen invasion signal is passed to NPR1 gene after treatment 12 h. The PR1 gene was regulated NPR1 gene after treatment 24 h, and the relative expression of PR1 began to increase. It activated the systemic acquired resistance(SAR) pathway.