The Function Of Cell Division Cycle Protein SsCdc28 In Sclerotinia Sclerotiorum

Sclerotinia sclerotiorum?Lib?de Bary is the main pathogen causing sclerotinia sclerosis in crops.This pathogen could infect most monocotyledonous and dicotyledonous plants,including most economic crops and oil crops.S.sclerotiorum is one of the pathogens that need to be closely controlled.The MADS-box gene family was an important transcriptional regulator widely distributed in eukaryotes.The SRF subfamily in the MADS-box family has a highly conserved domain,and SsMCM1 has a highly conserved domain of the SRF subfamily.The research team found that SsMCM1 was involved in the regulation of the growth and pathogenicity of S.sclerotiorum.Combined with S.cerevisiae MCM1 signaling pathway analysis,it was found that there was a protein SS1G₀2296 which was highly homologous to Cdc28 downstream of Saccharomyces cerevisiae MCM1.This gene was successfully cloned and named SsCdc28,and the function of SsCdc28 gene was analyzed by the following research contents.1.The SsCdc28 gene was successfully cloned from the wild-type S.sclerotiorum DNA as a template,and the full length was 1185 bp.Bioinformatics analysis of SsCdc28.2.In this study,NCBI was used to find the upstream and downstream sequences of SsCdc28 gene,and specific primers were designed.The wild type S.sclerotiorum DNA was used as a template to clone the 1246 bp fragment upstream nd the downstream 1225 bp fragment of SsCdc28 gene.The upstream and downstream fragments were ligated to the knockout vector pXEH to construct a S.sclerotiorum SsCdc28 gene knockout vector pXEH-Cdc28.3.Preparation of S.sclerotiorum protoplasts,and transformation of the constructed knockout vector pXEH-Cdc28 into protoplasts.The transformant-transformed transformants were verified by PCR amplification,and finally three positive SsCdc28 knockout mutants SsCdc28-7,SsCdc28-11,and SsCdc28-13 were obtained.4.Comparing the knockout mutant strains and wild-type S.sclerotiorum,the results showed that the mycelial growth rate of the knockout transformants became slower,the sclerotia could not be formed,the hyphae curved,the branching angle became larger,and the invasion the production of oxalic acid is significantly reduced,the ability to resist high osmotic stress is enhanced,and the pathogenicity is weakened.The above studies indicate that SsCdc28 is involved in the regulation of the growth and development of S.sclerotiorum and the production of oxalic acid and other substances,which in turn affects the growth and pathogenicity of S.sclerotiorum.The research further reveals the regulatory network of the transcription factor SsMCM1 and provides a foundation for exploring the growth and pathogenesis of S.sclerotiorum.