Fine Mapping, Map-based Cloning Of Cucumber Scab Resistance Gene Ccu And Expression Profiling Of Cucumber-Cladosporium Cucumerinum Interaction

Scab, is a very important disease of cucumber worldwide, and it is meaningful for usto further study of this disease.Cucumber types of"9930","9110gt","Gy14","Hardwickii"were employed in thepresent study. 9930 is a Northern china fresh market type that is susceptible to scab, 9110gtis a European green house type cucumber with scab resistance conferred by Ccu gene,Gy14 is a US processing type cucumber, and Hardwikii is a wild type originating fromIndia. Whole genomes of 9930,Gy14 and Hardwikii have been sequenced. Based on thesebackground, genome-wide analysis methods were used for studying cucumberNBS-encoding genes. The molecular markers and gene cloning technologies were used forfine mapping and map based cloning of Ccu candidates. The compatible and incompatibleinteractions between cucumber and the scab pathogen were studied using expressionprofiling supported by Illunima GA sequencing technology. The main conclusions are asfollows:1 Genome-wide analysis of cucumbers revealed that there were 58,60 and 60NBS-encoding genes in 9930, Gy14 and Hardwikii respectively, and the averagedistribution in genome was about 20 genes / 100 Mb genome, which was one-fifth thanthose in Arabidopsis, rice and poplar. In Kinase-2, Kinase-3, and GLPL domains,differences were found between different species. Seven frameshift mutations were foundamong the three cucumbers'NBS-encoding orthologouses.2 Fine mapping localizes Ccu into an R gene cluster. A F9 recombinant inbred lines(RILs 148 lines) population and a F2 population (2000 plants) were developed in this study.Ccu locus was delimit into a 140 kb genomic DNA region in the terminal of chromosome 2,it contained four nucleotide binding site encoding resistance gene analogs (RGAs), three ofthem were tandem repeats in this region. Comparative genomics analysis revealed that Ccugene region have syntenic regions in melon, Arabidopsis, poplar and grape (Figure 4-6 &4-7). It implied that Ccu homologous regions were originating from an old R gene cluster.3 Ccu candidates have been cloned. A 9110gt BAC library consists of 4100 individualclones was constructed, and BAC clones of H77-J7 and H74-D1 which contain Ccu regionwere selected, The H77-J7 clone was used for 11 kb library construction, and the targetclones which contain the Ccu candidate genes were screened.4 Expression profiling revealed the mechanism of cucumber scab diseases resistance.Gy14 (Scab Resistance) and 9930 (Scab Susceptible) were employed in this study.Twenty-four hours after inoculation, two HR pathways and a PR gene related diseaseresistance pathway (Figure 5-11 table 5-5) have been up-regulated in Gy14. On thecontrary, in 9930, the WRKY genes were up-regulated, but no PR genes were up-regulated, the plant did not detected the pathogen.In conclusion, Ccu has been fine mapped to a 140 kb DNA region. The Ccu candidateswere cloned by a map-based approach. It provided a platform for MAS of cucumber scabdisease resistance breeding and function analysis of Ccu gene. Expression profilingrevealed the mechanism of cucumber scab diseases resistance, which provides a model forcucumber diseases study.