Fine Mapping Of The Resistant Gene Ve To Potato Verticillium Wilt And Candidate Genes Analysis

Verticillium Wilt caused by Verticillium dahliae is a common fungal disease,also known as potato early dying disease.Once infected,the pathogen will gradually infect all parts of the plant,inhibit its normal growth and then make a devastating impact on the yield.So far,no fungicides available have been found to control this disease and relevant researches are relatively scarce.Thus,exploring and taking advantage of the gene against Verticillium dahliae in potato to accelerate the process of resistance breeding are the most efficacious prevention measures and of vital meaning.In this study,diploid potato interspecific hybrids C545 and V67 are used as parents to construct BC₁ isolated population for genetic mapping.And we have obtained these results on the basis of the primary mapping research:1.Progeny population construction and linkage marker development:With the hybridization of the parent plants,287 new materials of BC1 population were obtained for selecting recombinants.Using the previous BSA+DNA-seq data as a reference,69pairs of In Del primers were designed at the initial mapping region of chromosome 5and 13 pairs of In Del markers associated with verticillium wilt resistance were selected by t-test among segregation population.2.Recombinants selection and fine mapping of target gene:20 recombinants were selected among 375 individuals in total.And the phenotype of 5 recombinants whose exchange loci were within the mapping region have been identified by several rounds of the root soak inoculation.Combining genotype and phenotype information of recombinants,the candidate region was narrowed down from 1.7 Mb to 238 kb and then perfecting the genetic linkage map.3.Candidate gene analysis:Parent materials and partial resistant or susceptible progeny individuals were selected to be inoculated with Verticillium dahliae.The plants’root were sampled at the time point of 0 h,6 h,12 h after the inoculation for RNA-seq.According to the sequencing result and subsequent q PCR experiment,we preliminary determined a candidate gene,which Gene ID is Soltu.DM.05G005120,related to verticillium wilt in the candidate region.4.Over-expression vectors construction:In accordance with the reference genome sequence of DM1-3,specific primers for the candidate gene were designed.And some sequence differences were discovered between resistant and susceptible materials,which allowing us to construct over-expression vectors with different CDS sequences for the following transgenic function verification and relating mechanism researches.