| Apple valsa canker,caused by the Valsa ceratosperma filamentous fungus Valsa mali（V.mali）,lead to severe tissue maceration and necrosis in apple tree bark（Wang et al.2014）.This disease is a highly destructive disease resulting in yield and fruit quality losses on apple（Malus sp.）production.Therefore,apple valsa canker is a serious threat to the apple industry in China.At present,the prevention and friendly to the environment control system is inefficient due to the pathogenesis is not clear in V.mali.Therefore,in-depth analytical pathogenesis will be beneficial to reveal law of development of V.mali.More importantly,it is very big significance in safeguarding apple production safety.Zn（Ⅱ）2Cys6 class transcription factors exist in the fungus,and only participate in the process of fungal life activities.In our previous studies,two Zn（Ⅱ）2Cys6 cys6 named Vmtf266,Vmtf1026 were up regulation during infection by bioinformatics and V.mali genomes analysis.Base on this predicted result,we further analyzed the gene function of the two transcription factors via the knockout,phenotypic analysis such as mycelium growth,conidiation and pathogenicity.The main results are as follows:1.After the using of Double-joint PCR,PEG mediated protoplast genetic transformation and PCR detection,we got 6 and 10 positive transformants of Vmtf266 and Vmtf 1026,respectively.Finally,we got single copy knockout mutant namedΔVmtf 266-19,ΔVmtf 1026-31 in Vmtf 266 and Vmtf 1026 via Southern blotting,respectively.2.The phenotype observation of knockout mutant,resistance to salt ion stress test and determination of irulence were found that Vmtf 266 and Vmtf 1026 mycelium growth were decreased significantly.But the amount of spore production was not significant difference compared with 03-8.The pathogenicity of Vmtf 266 and Vmtf1026 deletion mutants was significantly reduced than the wild type strains 03-8.Vmtf 266 is sensitive to K+stress and Vmtf 266,Vmtf1026 is sensitive to Na+stress.This study shows that Vmtf266 and Vmtf1026 are related to the growth and development,pathogenicity,and stress response in V.mali. |
