Identification Of The Rapeseed's Resistance To Sclerotinia Sclerotiorum Using Aseptic Seedlings And Studies On Its Physiological Resistance

There are lots of methods to identify the rapeseed's resistance to S. sclerotiorum,but all of them more or less have limitations, and many factors influence them, such asdifficulties in controlling the conditions of disease development, the objectivity ofidentification standards and the classification of the resistance. Therefore, it is importantto find a new method which can be used in common.In this thesis, the technology system of using aseptic seedling to identify therapeseed's resistance to S. sclerotiorum is developed for the first time, and thebiochemical and physiological mechanism of the rapeseed's resistance to S.sclerotiorum are also studied. The main results as follows:1. The acquirement of the aseptic seedlings of rapeseed.The endophytes, which lives in the seed, has a strong impact on the cultivation ofthe aseptic seedlings. Among the 8 materials, the lowest one is No. 084 whose carrierrate is 70. 8%, and the highest one is Glycerin 5 whose carrier rate is up to 100%(Note:we disinfested the seeds surface, cultivated for 5 days and counted the number ofinfected seeds. From this experiment we got the carrier rate of the total number.)The part of the endophytes was first discovered by using the tissue isolationmethod. The endogenous fungus of the rapeseed can inhabit either the testas or theembryo, However, the endogenous bacteria of the rapeseed can only inhabit the embryo.By using different antiseptics to soak seeds, under constant temperatures, it is found thatalmost 50% of Thiran-quintozene WP have great germicidal action to the endogenousbacterium of rapeseed seed. The average carrier rates are 0 and 1.1%, M004 andGlycerin 5, which is obviously lower than that of sterile water and using conventionaldisinfectant, which is 0.1% mercury bichloride. The best condition for soaking seed of Thiran-quintozene wp: concentration100mg/L; prime temperature 37℃; time 7h.2. Identifying etticiem timing using aseptic seedlings to inoculate S. sclerotiorumTo ensure the identification of efficient timing inoculate S. sclerotiorum ondifferent types of rapeseed using aseptic seedlings with different timings.(for 24h, 42h,66h, 72h). The efficient timing is 42 to 66h after using the aseptic seedling to inoculate,because the occurring rate of the disease is high and have some remarkable difference inthe disease-resistance of the varieties in this period.3. The result of testing the resistance of the rapeseed to S. sclerotiorum by usingaseptic seedlings to inoculate.The materials are Brosica juncea var crispifolia and various types of rapeseed. Itshows that: M004 and R-888 belong to high resistant varieties, 085 is of low resistant,and 98C40 is in the middle susceptibility.4. The result of testing the resistance of the rapeseed to S. sclerotiorum obtainedfrom planting in the field.Do the experiments during initially flowering stage and the end flowering stagesrespectively. The result obtained from the initially flowering stage indicated that thediseases occurrence have great differences: the most susceptible variety is Brasicajuncea var crispitlia, which is similar to the result obtained from above. The index ofM004 and R-888 are the same which belong to high resistant varieties were the same,this was in agreement with the results obtained from other issues.The result of this experiment shows that the various types of rapeseeds havesimilar resistant abilities.5. The biochemical and physiological mechanism of resistance of the rapeseed tostem rotThe resistance was positively correlated with Ca²⁺ content in the tissue. The Ca²⁺content in M004, R-888, 085, were larger than that of Glycerin 5 and 98C40. In thetissue of R-888, The Ca²⁺ content was 16.4% larger than that of Glycerin 5. The higher content of Fe²⁺, the stronger susceptibility of rapeseed. Mn²⁺, Zn²⁺, Cu²⁺ had nocorrelation with resistance to stem rot.The experiment also indicated that the tannin content of the varieties treated byinoculation was higher than that of untreated varieties. The content was 25.9%, 25.4%43.3%, 40.0% and 16.7% (in treated M004, R-888, 085, Glycerin5, 98C40)lager thanthat of untreated species. But neither treated or untreated, the tannin content of thevarieties, which had higher resistance, was lager than 98C40.The resistance among the different varieties did not have an obvious correlationwith the malonic aldehyde content.The endogenous oxalic acid content in the rapeseed is related to resistance. Thehigher its resistance, The larger its content.. The endogenous oxalic acid content inM004, R-888, 085 were 22.2%, 16.0%, 10.4% larger than that of 98C40.After being inoculated, the PAL activity had increased, 25.9%, 11.5%, 41.4%,30.1% more than that of the untreated in leaves. But after inoculating in the leaves ofthe susceptible varieties, the PAL activity had decreased by 11.5%.After being inoculated with S. sclerotiorum, the SOD activity in various haddifferent changes. The increase of SOD activity in susceptible varieties is larger thanthat of the resistant varieties.After being inoculated, the varieties of decreased rapidly of respiration intensityhad higher resistance. Using aseptic seedlings method to identify the rapeseed'sresistance to Sclerotinia sclerotiorum can reflect the inborn differences of various typesof rapeseed's resistance. These differences are determined by their physiologicalresistances. For instance, the differences of the content of the mineral chemical element(Ca²⁺, Fe²⁺), and the content of secondary metabolite (eg: the endogenous oxalic acid,tannin), the differences of the activity of enzyme and the differences of the intensity ofrespiration. In conclusion, this new technology is accurate and reliable.