| Sheath blight (SB) disease is one of the three most serious diseases in rice worldwide, and causes severe loss in yield and quality under favorable environmental condition. The resistance of rice to SB is a typical quantitative trait, and resources with strong resistance against this disease in the existing rice germplasm are not found.Rice SB inoculation identification system involves the inoculation method, inoculation time, investigation time, investigation standard and other factors, and the appropriateness of the system can affect genetic and breeding research progress of the resistance to SB in rice. Appropriate inoculation identification system can not only reduce the strength of the research work, but also improve the reliability, repeatability, and accuracy of the test results. In view of this point, five rice cultivars with different levels of resistance to SB were adopted to identify their resistance to SB at seedling stage in artificial climate incubator or temperature-controlled greenhouse, and the test results were verified through a field experiment during rice adult stage, then a SB inoculation identification system in rice seedling stage was established preliminarily. Based on the above study, five rice cultivars were inoculated with toothpick-inserting inoculation method during different times in rice adult stage (the late tillering stage, the early booting stage) in field and temperature-controlled greenhouse. The lesion length was measured during3-11days after inoculation, and the SB disease rating at the30th day after heading was investigated with the reference of three different investigation standards. Then, some new inoculation identification methods in rice adult stage were developed and some new investigation standards of SB disease rating at the30th day after heading stage in field were also explored in this study.YSBR1developed by our own research group is a new germplasm with stable and high resistance to sheath blight, which has been identified by using many methods in different experimental sites for many years. However, information of its genetic background relevant to SB resistance has been in scarce yet. A F2clonal populations (F2CP) derived from the cross between YSBR1and Lemont (a susceptible cultivar) was constructed, and the identification of resistance to SB and the investigation of agronomic traits for F2CP in a multi-year experiment were carried out in this study. At the same time, the QTLs mapping of resistance to SB in YSBR1and agronomic traits were conducted, and the interrelation between resistance to SB and agronomic traits were also analyzed. On the basis of this study, the two cultivars (YSBRl, Lemont) were used as materials to inoculate SB pathogens, and their leaf sheaths were sampled at10and20hours after inoculation respectively for gene chip expression profiling analysis and screening of differentially expressed genes, especially YSBR1specific differentially expressed gene. Furthermore, the pathway and GO (gene ontology) of YSBR1specific differentially expressed gene and its integration with QTL detected in YSBRl were analyzed.The main results were as follows:1. The optimal relative humidity for SB mycelium to develop infection structures and to infect rice seedling was approximately85%. The SB resistance levels in5cultivar seedlings differed significantly (P<0.01) and could be divided into two groups, relatively susceptible (Lemont, WuYujing3) and relatively resistant (YSBR1, Jasmine85, Teqing). The symptom severeness degrees of the seedlings at different leaf-age (4-leaf stage and5-leaf stage) were significantly different and the average disease rating at4-leaf stage was significantly higher than that at5-leaf stage. The differences of resistance among different cultivars in greenhouse were smaller than that in the field, but the resistant-to-susceptible order among the cultivars were consistent with that in the field. Rapid identification technique of rice SB resistance in seedling stage proposed in this study could be used for screening resistance of rice germplasm on a large-scale scope or for primary resistance identification.2. The SB resistance levels evaluated with the lesion length among5cultivars inoculated at the late tillering stage in temperature-controlled greenhouse differed significantly(.P<0.01) and could be divided into three groups, relatively susceptible (Lemont), moderate susceptible to moderate resistant(Wuyujing3, Jasmine85, Teqing) and relatively resistant (YSBRl), and the resistant-to-susceptible order among five cultivars were consistent with the results tested in field verifying experiment at30th day after heading. Although the SB resistance levels among5cultivars inoculated at the same time in field differed significantly at1%levels, the resistant-to-susceptible order among five cultivars was inconsistent with that of the30th day after heading. Furthermore, the SB resistance levels among5cultivars inoculated at the early booting stage in field differed significantly at1%levels, and the resistant-to-susceptible order among five cultivars were consistent with that of the30th day after heading. These results indicated that the lesion length used as investigation index to evaluate rice SB resistance at the late tillering stage in greenhouse or at the early booting stage in field was feasible.3. There were no differences between the SB disease rating data measured with the common Rush’s investigation standard and with the "order of leaf sheath" investigation standard in five rice cultivars inoculated at the late tillering stage in different years and locations, and the correlation index of SB disease rating data between two investigation standards in different years and locations were0.9455,0.9846(P<0.01), respectively, which indicated that the "order of leaf sheath" investigation standard as a new standard could replace Rush’s investigation standard completely in evaluating rice SB disease rating. The difference of the SB disease rating data measured with the relative disease rating investigation standard among five rice cultivars inoculated at the early booting stage was not only significant (P<0.01), but also greater than that among the cultivars inoculated at the late tillering stage by means of the "order of leaf sheath" investigation standard. The resistant-to-susceptible order among the five cultivars between two investigation standards were consistent completely, which showed that the relative disease rating investigation standard as a new standard could also be used for evaluating rice SB disease rating in some feasible conditions. The application scopes of relative disease rating investigation standard in genetic research of resistance to rice SB was limited.4. In two years experiment(2009-2010), twelve resistant QTLs were located, of which qSB-1Le, qSB-5Le and qSB-8Le were from susceptible parent Lemont, and remaining QTLs were all from YSBR1. Among QTLs detected, qSB-2Y and qSB-12Y could be considered as the major effective QTLs, with the average contribution rate of28%and17%in2009and2010, respectively. Furthermore, the QTLs of13major agronomic traits were located and their relationship with the SB QTLs were analyzed. There existed significant correlation (mostly negative, P<0.01or P<0.05) between four agronomic traits (plant height, growth duration, panicle length, effective panicles) and SB disease rating. This result showed that the incidence of rice SB was affected by the traits of plant height, growth duration, panicle length, effective panicles, of which plant height had the most powerful effect on the SB disease rating.5. The difference of SB resistance levels within a short-term (5days after inoculation) between Lemont and YSBR1was significant (P<0.05), and the infection cushion and lesion appaered in10and20h after inoculation, respectively. The quality of gene chips of different biological replicates were reliable,2826differentially expressed genes in Lemont whole genome and748differentially expressed genes in YSBR1whole genome were screened out by using our screening standard respectively. Furthermore,247specific expressed differentially genes in YSBR1were also screened out. The specific expressed differentially genes in YSBR1were distributed among12rice chromosomes, and31genes fell into the detected QTL intervals, with the proportion to the total of12.55%. There were9and3YSBR1specific expressed differentially genes fell into the intervals of two major QTLs (qSB-2Y, qSB-12Y). Four and2genes were upregulated, and5and1genes were downregulated in qSB-2Y and qS.B-12Y intervals, respectively. |
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