| Black rot is one of the most important diseases affecting crucifers and is widespread worldwide.In production,chemical control is the main focus,and the massive use of chemical pesticides has caused great damage to the environment and ecology.Production practices show that growing resistant varieties is an economical,safe,and effective way to prevent black rot in crucifers.Investigation of locally predominant flora,analysis of mutualistic relationships between pathogenic bacteria and hosts,and screening of host target proteins can streamline the cultivation of cabbage and the selection and breeding of multi-resistant black rot cultivars.In this study,genome sequencing and genetic diversity analysis were performed on black rot pathogens isolated from major Cabbage growing areas in China,using Xanthomonas campestris pv.campestris and black rot susceptible Cabbage as test material.The type III effector Xop R was selected by genome-wide comparative analysis,and the target proteins interacting with the effector protein Xop R were studied using the constructed Cabbage c DNA library to provide a basis for elucidating the mechanism of interactions between pathogenic bacteria and plants and to obtain new ideas for breeding black rot-resistant Cabbage.The main results are as follows;(1)Isolation and identification of the black rot fungus was carried out in the main Cabbage growing areas in nine different provinces and cities in China.The strains were typed using molecular biology,and the indigenous black rot bacteria were classified into three subtypes represented by the BJSJQ20200612 strain,the GSXT20191014 strain,and the race1 strain,with the BJSJQ20200612 strain having the largest distribution and the GSXT20191014 strain being the most virulent.(2)Based on whole-genome sequencing,molecular markers from genus to species were integrated from previous studies,and molecular markers specific to BJSJQ20200612 strain and GSXT20191014 strain were developed,which can rapidly identify black rot strains in the field in different regions.(3)Genome-wide analysis showed that there were 26 and 47 insertion loci in the BJSJQ20200612 strain and the GSXT20191014 strain,respectively,and these loci contained 6 and 10 virulence-associated virulence factors,respectively.Analysis of the virulence factors(T3SS,LPS,and EPS)of the BJSJQ20200612 strain and the GSXT20191014 strain showed that there were genetic nucleotide differences between the strains.Evolutionary tree analysis showed that the BJSJQ20200612 strain was more closely related to MAFF 302021,and the GSXT20191014 strain was more closely related to the race2 strain(HRIW 3849A).(4)Sequence analysis revealed that gene Xop R is a highly conserved III type effector.To investigate the function of Xop R,a deletion mutant ΔXop R or an overexpression vector p YBA1144/Xop R were constructed.Comparison of the pathogenicity of kale revealed that ΔXop R was less pathogenic than the wild strain HRIW 3811,and overexpression of p YBA1144/Xop R Agrobacterium-injected tobacco revealed that Xop R could inhibit BAX-induced cell death.(5)Using a disease-prone Cabbage c DNA library to search for the Xop Rinteracting target protein Bo SGR1,Xop R was confirmed to interact with Bo SGR1 using yeast two-hybrid,subcellular localization,and bimolecular fluorescence complementation.The q RT-PCR analysis showed that Bo SGR1 can be induced by Xcc,SA,Me JA and ETH.Silencing of Bo SGR1 gene increases black rot resistance in Cabbage,and overexpressed Arabidopsis thaliana was more susceptible to black rot.Bo SGR1 is a susceptibility gene in Cabbage and can be used as a new target for breeding disease resistance. |
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