The Functional Analysis Of The Transcription Factor Ss-FoxE2in Sclerotinia Sclerotiorum (Lib.) De Bary

Sclerotinia sclerotiorum （Lib.） de Bary is one kind of filamentous fungi whichcan cause a variety of plant diseases and huge losses to agricultural production.Apothecia is the very important part of S. sclerotiorum and formed from sclerotia,under suitable environmental conditions, it can release large quantities of ascosporesthat initiate new disease cycles.In S.sclerotiorum, there were4Forkhead-box transcription factor homologousgenes, Ss-FoxE1, Ss-FoxE2, Ss-FoxE3and Ss-FoxE4respectively, and we infer thatthey closely related to sexual reproduction of S. sclerotiorum. So, in this research, westudied the role of Ss-FoxE2in S. sclerotiorum. According to S. sclerotiorum genomesequence,890bp upstream fragment and1024bp downstream fragment of Ss-FoxE2were cloned and ligated into pXEH respectively to construct the knock-out vector forSs-FoxE2. In one hand, protoplasts of S. sclerotiorum have been prepared by Lysingdecomposition of hyphae. The knock-out vector for Ss-FoxE2was transformed intoprotoplasts of S. sclerotiorum and transformants were screened on PDA medium withhygromycin resistance, first transformants were verified with fast PCR process andsouthern blot for further validation. In the other hand, we cloned the whole Ss-FoxE23.4kb fragment including upstream and downstream sequence, then ligated intocomplement vector and transformed to S. sclerotiorum Ss-FoxE2knock-out mutantprotoplast. Transformants were screened on PDA medium with G418resistance andverified by PCR process. After phenotype and pathogenic analysis among S.sclerotiorum wild type, S. sclerotiorum Ss-FoxE2knock-out mutant and complementmutant, we found that there was no difference in hyphae growth, number and weightof sclerotia and pathogenicity, but no apothecia formation in Ss-FoxE2knock-outmutant, this indicated that Ss-FoxE2has a very significant role in S. sclerotiorum sexual reproduction, it will provide new theoretical foundation and technical supportfor S. sclerotiorum disease control.