Identification Of The Pathogenicity-Defective Mutant AbS4b Of Alternaria Brassicicola And Functional Analysis Of The Related Gene

Alternaria brassicicola is an important pathogenic fungus of cruciferous vegetables.The black spot disease caused by it can cause severe decline in the yield and quality of cruciferous vegetables.In-depth research on the pathogenic mechanism of the pathogen will help prevent and cure black spot disease.Blue light can play an important role in the growth and development,sexual reproduction and asexual reproduction,metabolism,conidia production and pathogenicity of pathogenic bacteria.Therefore,this study started with the pathogenicity-defective mutant AbS4b and identified its mutant gene as the blue light regulator Abblr,and further studied the function of this gene.The results are as follows:?1?The pathogenic defective mutants in the transformant library of A.brassicicola was screened in vitro leaf inoculation method.We obtained 5 mutants with severely reduced pathogenicity,of which two were completely lost in pathogenicity,namely AbM1a and AbS4b.There are three strains with severely reduced pathogenicity,namely AbF7a,AbS2b,and AbS3b.The plasmid insert gene of mutant AbS4b was successfully amplified by hiTAIL-PCR technology,and was named as the blue light regulatory protein gene.The gene inserted by plasmid of mutant AbS4b was successfully amplified using hiTAIL-PCR technology,as the blue light regulator gene,which was named Abblr?Alternaria brassicicola blue light regulator?.?2?Abblr gene is 1432 bp in length containing a 52 bp intron.It encodes 459 amino acids,in which containing a PAS domain and a ZnF_GATA domain.?3?The Split-marker PCR recombinating technology was used to construct the targeting vector and using PEG-mediated transformation,Abblr gene was successfully knocked out and complemented.Functional studies found that the mutant?Abblr has a great change in colonies,showing a milky white color,the center was raised upward and aerial mycelia are slender and dense.The growth rate of the mutant is slow than that of WT in the minimal medium?MM?,but slightly higher than that of WT in rich medium?PDA?.The Abblr gene regulates the spore production of A.brassicicola,the mutant?Abblr loses its ability to produce spores,and the mycelial morphology appears partially enlarged in the minimal medium?MM?.The pathogenicity of the mutant?Abblr is severely lower than that of WT,the deletion of the Abblr gene reduced the colonization ability of A.brassicicola,but there is little effect on host penetration.The mutant?Abblr has a reduced ability to degrade H₂O₂ in vitro.The deletion of the Abblr gene leads to reduction of carotenoid-producing ability and repair ability after Blu-ray,conidia production cannot be induced under the condition of C-deprivation.In summary,the Abblr gene is involved in the regulation of colony morphology,vegetative growth,conidia production,pathogenicity,degradation ability of ROS,carotenoid-producing ability and blu-ray repair ability of A.brassicicola.