Method Evaluation Of Sclerotinia Resistance Identification And Allelic Variation Analysis Of Resistance-related Genes And The Other Traits Genes In An Oilseed Rape Pedigree

China is one of the largest country producing rapeseed in the world, but edible plant oil produced domestically is serious shortage. Huge import of edible plant rapeseed or oil has led to serious and negative effect on domestic production of oilseed rape. It is urgent for China’s rapeseed production to reach the targets of "double low","three highs" and "industrialization and mechanization", including increase of plant resistance to Sclerotinia disease. To help achieve these targets, we used a pedigree to study the allelic variations of the genes related to disease resistance, grain weight, and oil content, which will benefit molecular breeding. To do so, we first evaluated resistance evaluation method to be used for pedigree resistance identification. The main results are as follows:1. Evaluation Method Identification for Resistance of Sclerotinia sclerotiorumIn this study, S. sclerotiorum resistance of Zhongyou821and its pedigree was evaluated by the detached leaf inoculation, the in situ stem inoculation and natural disease nursery identification. The results showed that identification methods to be adopted are dependent on research purpose:natural disease nursery is a good method to comprehensively evaluate varietal resistance and avoidance and thus necessary for resistance identification of varieties which intend to be commercialized; the rest two methods are just to distinguish varietal ability to resist disease expansion and thus are complementary to the first method and suitable to be used for resistance genetic analysis.2. Cloning and allelic variation of S. sclerotiorum resistance-related gene BnERF104Two copies of BnERF104Ⅰ and Ⅱ were predicted from the rapeseed genome database. The two copies of BnERF104were cloned from Zhongyou821and its pedigree with the universal primer. The analysis indicates that:in Zhongshuang6,7,10, Huashuang3,4, and Huyou19, there are5SNPs in the copy II and a DNA fragment insertion. We can infer that the allelic variations were introduced by the other parents, and the allelic variations may make a contribution to the stronger ability to resist disease expansion of Huyou19than Zhongyou821.3. Cloning and allelic variation of seed weight gene BnTTG2Two copies of BnTTG2Ⅰ and Ⅱ were predicted from the rapeseed genome database. The two copies of BnTTG2were cloned from Zhongyou821and its pedigree with the universal primer. The analysis indicated that:the BnTTG2copy I has a SNP which leads to early termination in transcription. So the functional domain of the product was lost. It suggests that the SNP loci have a major impact on seed weight and it is related to smaller seed weight of Zhongyou821. The subsequent experiment proved that this SNP in Zhongyou821was acquired from its parent and passed on to some members of its pedigree.4. Cloning and allelic variation of oil synthesis gene BnBCCP2Five copies of BnBCCP2were predicted from the rapeseed genome database. One of the copies, which is with the highest expression level in ovule and the lower expression in other tissues, was selected for next analysis. This copy of Zhongyou821and its pedigree was cloned with the specific primer. The analysis shows that the allelic variations in the third and fifth intron exist in some members of the pedigree. The primer was designed to amplify the section between the two allelic variation sites. The results showed that the varieties with allelic variations have higher oil content than the ones that have no allelic variations.