Cloning And Functional Analysis Of Abcdm1 Gene Of Alternaria Brassicicola

Black spot,caused by Alternaria brassicicola,is one of the important diseases in cruciferous vegetables,which seriously affects the yield,quality and safety of cruciferous vegetables.However,there are no effective measures for prevention and control up to now.Conidia play an important role of the fungal pathogenicity.Therefore,finding the gene related to sporulation of A.brassicicola will be helpful for understanding the pathogenic mechanism and provides a new strategy for the prevention and treatment of the disease.In this study,nineteen conidiation-deficiency mutants were obtained from the mutant library.The flanking sequence of insertion of mutant M85 was obtained and proved to be a transcription factor Ndt-80 of the yeast homologous gene.We named it as Abcdm1?A.brassicicola conidiation-deficiency mutant?.The Abcdm1 gene was cloned and analysed.And we adopt the method of protoplast of PEG mediated transformation,to obtain a knockout mutant?Abcdm1and one complementation C and analyze the biological function,the results are as follows:Conidiation-deficiency mutants from the mutant library were screened and nineteen conidiation-deficiency mutants were obtained.We get thirteen mutants without conidiation,namely M17,M37,M51,M56,M73,M85,M131,M96,M101,M133,M159,M204 and M324.Six mutants'sporulation capacity decreased,namely M15,M108,M122,M183,M258and M177.The flanking sequence of insertion of mutant M85 was amplified and named as Abcdm1.Amplification and analysis of Abcdm1 gene show that the full-length gene were 1905 bp,cDNA were 1632 bp,encoding 543 amino acid.Abcdm1 regulates colony morphology and conidiation of A.brassicicola.The mutant's hypha is fluffy.The hypha is white and can't produce melanin.However,The growth rate of colony is consistent with that of wild type strain.We couldn't find any conidium by microexamination.However;the mutant can produce chlamydospore in the 1.5 M sucrose.Abcdm1 regulates stress sensitivity of A.brassicicola.?Abcdm1 showed increased sensitivity to KCl and NaCl.But?Abcdm1 can grow normally in 30mm H₂O₂.Abcdm1 regulate external penetration of A.brassicicola.The mutant lost external penetration.The pathogenicity of mutant is consistent with that of WT.Abcdm1 regulates nutrition metabolism.Mutant?Abcdm1 respond differently on carbon source and nitrogen source.Adding glucose to the basic medium,we found that the mutant can't barely grow.However,adding BSA to the basic medium,the mutant can grow.To sum up,Abcdm1 of A.brassicicola was involved in regulating stress sensitivity of the response,the generation of secondary metabolites,penetration in vitro and nutrition metabolic response.