Functional Analysis Of Cat1 And CcP Gene In Fasurium Oxysporum F.sp.cubense

The banana fusarium wilt caused by the pathogenic fungus Fusarium oxysporum f.sp.cubense (Foe) is a devastating soil-borne vascular disease in banana plants. We can't effectively control the banana fusarium wilt bacause of the lack of resistant cultivars and effective chemical fungicide. But also the chemical fungicide may lost in the field and damage to the environment and effect human health. Therefore, disease-resistant varieties cultivation becomes a very urgent alternative strategy. Better understand the pathogenic mechanism by explore the pathogenic genes of Foe is beneficial to the research of disease-resistance breeding.It was found that the cat1 and CcP gene in Fusarium oxysporum f. sp. cubense race 4(Foc4) was up-regulated induced by Musa sp. AAA. cv. Baxi in our previous proteomics analysis. In order to investigate the function of cat1 and CcP gene during the host infection process of Foc4, the cat1 and CcP gene in Foc4 was respectively deleted by the homologous recombination method, and was respectively complemented. The phenotype and pathogenicity of the knockout mutants and complement mutants were then analyzed.The results are listed as follows:1. Homologous arms of left and right side of cat1 gene and CcP gene in Foc4 were amplified, and then the knockout vectors with GFP (green fluorescent protein) were constructed successfully.2. PEG-mediated protoplast transformation method was used to do transformation,we obtained the cat1 knockout mutant and CcP knockout mutant with GFP (green fluorescent protein) and HPH (hygromycin B phosphotransferase) double labeling. And the catl complement mutant and CcP complement mutant with G418 resistance were obtained by Fusion PCR method. Furthermore, the phenotype and pathogenicity of the complementation mutation is similar to the wild-type strain.3. There were no apparent difference between the catl knockout mutant and the wild type strain on the morphology mycelium and spores, colonial growth, osmotic stress resistance and cell wall selective pressure. However, the mutants showed significant decrease in the abilities of exogenous oxygen stress resistance, penetrability of cell wall and the pathogenicity to Musa spp. AAA. cv. Baxi. These results suggested that the cat1 gene in Foc4 should be involved in pathogenesis by scavenging host-derived ROS.4. Compared to the wild type of Foc4, the ability of exogenous oxygen stress resistance decreasing in the CcP knock out mutant while the mutants was similar to the wild type on the morphology of mycelium and spores, colonial growth, osmotic stress resistance, ability of cell wall selective pressurepenetr and colonization ability in living leaf of banana.5. Both cat1 knockout mutant and CcP knockout mutant showed significant decrease in the ability of cellulose utilization. It is suggested that the two gene maybe involved in the process of host cellulose degradation.