Mapping And Analysis Of QTLs Associated With Disease Resistance To Sclerotinia Sclerotiorum In Brassica Napus

Sclerotinia (stem rot) disease in rapeseed, caused by fungal pathogen Sclerotiniasclerotiorum, is the most important disease in the worldwide and especially seriouslytaking place in the southeast coastal area and Yangtze River Area of China and hasrestricted the development of rapeseed industry for a long time. Till now, rapeseedcultivars with complete resistance to this disease haven't been identified, nor have relatedspecies. Resistance to Sclerotinia sclerotiorum in rapeseed belongs to quantitative traitand easily affected by environment condition and other factors. There is no widelyaccepted method to identify resistance. These questions have made it difficult for breedersto select really resistant materials. The objectives of our study were to 1) screen moreprecise identification resistance methods, 2) QTL mapping the resistance to Sclerotinia,which will be expected to help understand the mechanisms of disease resistance, and canbe used for selecting high-quality cultivars with strong resistance to Sclerotinia.We compared different identification resistance methods at seedling and maturingstage in 8 resistant and susceptible materials. We obtained one DH (doubled haploid)population derived from a cross between zhongyou821 and Bao604. Then, We did QTLmapping of resistance to Sclerotinia by utilizing the resistance performance of DHpopulation at different growth stage and molecular markers such as SSR, RAPD andRFLP. The main results are followed below:1 Comparison of different identification resistance methodsThe resistance at seedling stage was evaluated by petiole inoculation technologyby placing mycelial plugs on detached leaves and immersing petioles into oxalic acidsolution. The resistance at maturing stage was evaluated by placing mycelial plugs andtoothpicks on axilla to induce stems disease. All these methods showed effective foridentifying resistant and susceptible plants. There was significant correlation betweenmycelial plugs inoculation method and toothpick inoculation method. There weresignificant correlation between only the petiole inoculation technology at seeding stageand mycelial plugs and toothpicks inoculation methods at maturing stage.2 QTL Mapping resistant genes conferring Sclerotinia sclerotiorum in B.napusTwenty-one QTLs controlling resistance were detected by Composite IntervalMapping Method (CIM) with a LOD score more than 2.5. The single QTL accounted forphenotype variation from 10.21% to 36.06%. Ten QTLs were totally detected by mycelialtoothpicks inoculation in 2004 and 2005, accounting for phenotypic variation from10.21% to 36.06%. Only one QTL was detected by mycelial petals, accounted for 13.62%of the phenotypic variability. Ten QTLs were detected by mycelial plug inoculations in2006 and 2007, accounting for phenotypic variation from 11.63% to 15.10%. No commonQTLs has been detected by three methods, during four years. But there was one QTLdetected in linkage group N12 by mycelial toothpick inoculation method in 2004-2005and by petiole inoculation method in 2005, one QTL was detected in linkage group N14 and N4 by mycelial plug post-inoculation 3d to 7d in 2006-2007, and there were also onecommon QTL detected by mycelial toothpick post-inoculation 3d to 7d in 2005 and bymycelial plug inoculation-post 3d in 2006.3 Utilization of expressed sequence tags (EST)By analyzing the resistance genes involved in the procedure post-inoculated 0.5 to8.5 hours by Suppression Subtractive Hybridization (SSH) and reverse northern blot, 50unrepeated ESTs were identified. According to the putative function analysis, these ESTsfrom Brassica napus were related to the primary metabolism, energy metabolism,secondary metabolism, disease defence, cell structure, transcription, protein synthesis,and signal transduction, which were involved in the oxgen burst and jasnonade pathway.Mapping genes with known function onto the whole genome of Brasscia napus withresistance-relevant EST markers. We found out a resistance-related EST marker in LG13linked with one common QTL which was also detected by mycelial toothpickpost-inoculation 3d in 2004 and 2005.