Identification Of Effector In Racel Of Fusarium Oxysporumf.sp.conglutinans

Cabbage Fusarium wilt, caused by Fusarium oxysporum f. sp.conglutinans (Foc), has been one of widespreadfungal vascular diseasesin cabbage planting area of Northern China. The disease spreads quickly, resulting insevere damageto cabbage planting industry. Up to now, due to the lack of safe and effective chemical control measures against Foc, using of resistant varieties has become the major way to control the diseases. However, the resistance to Fusarium wilt would lost as a result of the rapid evolution of virulence genes in Foc, especially effectors, which play important roles in the interactions between Foc and their hosts. Therefore, understanding the pathogenic mechanisms of Foc is vital to the defence and control of Foc. In this study, we focused on the identification and verification of Foc effectors and obtained following results:1. Comparative genome and transcriptome analyses of racel and race2 of Foc revealed that 17 secreted protein genes were expressed only in race 1. For these genes, three genes (FOCA1-01935, FOCA1-11385 and FOCA1-13811) were not expressed in vitro but expressed in planta, which could be specific secreted protein genes in race 1 of Foc.2. We collected xylem sap of cabbage inoculated with Foc race 1 for seven days and analysed the proteome by mass spectrometry. Compared with the transcriptome, we obtained four secreted protein genes. Then, q-PCR experiments confirmed that a gene FOCA1-16510was upregulated in planta.3. Functional study of four genes (FOCA1-01935, FOCA1-11385, FOCA1-13811 and FOCA1-16510) was conducted using gene knockout procedure. The gene encoding hygromycin B phosphotransferase was chosen as a selectable marker. Combining the split marker strategy with protoplast transformation approaches mediated by PEG-CaCl2, we obtained four gene deletion mutants respectively, which were verified by Southern hybridization.4. In these four mutants, the virulence of deletion mutant FOCA1-01935 was reduced while the mutant FOCA1-11385 increased virulence on cabbage. However, the growth rate, germination rate and production of conidia of four deletion strains showed no significant difference compared with wild type isolates.