Analysis Of Silique Shatter Resistance And Its Qtls Mapping In Brassica Napus L.

During harvesting of rapeseed, the silique shatter loss proportion usually reaches up to10%to30%. In order to decrease the yield loss caused by silique shatter, it is necessary to breed varieties with silique-shatter resistance, which are suitable for combine harvesters. Since Brassica napus is the main rapeseed type used in production in China, breeding practice requires that theoretical researches of silique-shattrer resistance be carried out in advance.This study firstly improved random impact test (RIT) and set up silique-shatter resistance index (SSRI) as a measurement for this trait. Then a slilque-shatter resistant line H155and a slilque-shatter susceptible line Qva were selected from the Brassica germplasm. Silique-shatter resistant trait and their influencing factors were also studied.During construction of DH mapping population, microspore cultural techniques were improved for higher efficiency and lower costs. A linkage map with19linkage groups was constructed and13QTLs which were related to silique-shatter resistance were mapped. These results can be helpful in molecular assistant selection in rapeseed silique-shatter resistant breeding.The main results in this study are listed as followings.1Setting up a test system for silique-shatter resistance and screening silique shatter resistant lines1.1Random impact test (RIT) was improved and silique-shatter resistance index (SSRI) was also firstly set up for evaluating silique-shatter resistant ability in rapeseed. Results indicated that there were extensive variance of SSRI in Brassica napus germplasm, which SSRI ranged from0.000to0.7675and the coefficient of variance (CV) was as high as114.4%. In the screened germplasm, about59.38%were silique-shatter sensitive accessions,32.75%were silique-shatter susceptible accessions. Only4.01%accessions with more than0.5of SSRI. Two silique-shatter resistant accessions (H155and98009) with more than0.7of SSRI were obtained.1.2Results showed that SSRI detected by RIT were stable and credible.The differences of SSRI among accessions reached1%significant level. However the differences among years and replications did not reached significant level.1.3Results of simple correlation analysis revealed that SSRI had no correlation with the angle between stalks and inflorescences, angles between stalks and silique and stalk length.All these parameters decide silique distribution in space and are directly related to scraping and silique shatter loss during harvesting. Silique number per cm along inflorescences and branches had significant (1%) negative correlation with SSRI. Silique length, silique width, beak length, valve thickness and seed number per silique had significantly (1%) positive correlation with SSRI.1.4There were differences in the valve structures between silique-shatter resistant accession H155and silique-shatter susceptible accession Qva.Compared with Qva, the cells in endopericarp of H155arranged more tightly and the cell wall lignified more seriously, and even the thin-walled cells in mesocarp were lignified and formed thick lignified layer.There were more vascular boundles in the valves of H155, which enhanced the mechanical strength.2Influencing factors analysis of silique-shatter resistance in Brassica napus2.1Results revealed that genetic differences were the crucial influencing factors of silique-shatter resistance.The SSRI of the same accessions among different replications did not reached to signifficant level. The SSRI of different accessions got to signifficant difference. The SSRI CV of silique from different branches ranged from18.67%to93.57%.Except for Ningyou No.10, siliques from the first branch had the biggest SSRI for other4accessions.The SSRI of siliques from the second branch and other above branches had not significant difference.2.2SSRI differed because of siliques from different portions in plants in different accessions, which CV of SSRI ranged from18.67%to93.57%. For most accessions, the siliques from the first branch had the largest SSRI. Siliques from the second branche and upper branches hand similr SSRI.Siliques from middle portion and lower portion had similar SSRI. SSRI of siliques from top portion were less than those from middle and lower portion in inflorescence by55.04%and60.43%, respectively. Siliques from the lowest portion had the largest SSRI.2.3There were difference of SSRI changes during dehydration bentwen silique-shatter resistant accessions and silique-shatter susceptible accessions.Compared with silique-shatter resistant accessions, SSRI of silique-shatter susceptible accessions decreased more sharpy. Results implied that water content in silique had linear regression relationship with SSRI (y=0.0149x-0.4779).The coefficient of determination (R2) was0.4284. Consequently SSRI enhanced while water content in silique increased.3Technique improvements in isolated microspore culture3.1Sampling time was put forward from traditional time (3days after anthesis) to9days before anthesis. Results showed the embryo yields were not significantly different from improved sampling time to that by traditional sampling time.The improved sampling time could increase total sampling time by12days and the total suitable sampling time was doubled.3.2In improved sampling method, suitable buds were harvested from donor plants in field and small buds were left to grow up and one donor plant could be used for several times. In addition, buds from the same donor plants had the same genetic background.3.313%sugar solution substituted B5medium as extraction solution. Results confirmed that embryo yield with improved extraction solution was not significantly different from that of B5medium. Sugar solution could save reagents and labour for preparation B5medium.3.4In the improved microspore incubation procedure, the microspores were directly incubated at32℃in dark till the embryos were visible. Compared with the traditional incubation procedure, the embryos were visible on the tenth day, which decreased the time for embryo visibility by four days.4Construction of linkage map and QTLs mapping of silique shatter resistance4.1The frequency distributions of SSRI in P1(H155), P2(Qva), F1,F2, BC1F1and BC2F1were continuous.The proportion of silique-shatter susceptible plants in populations of F1, F2, BC1F1and BC2F1were high and no plants had higher nor lower SSRI than those of both parents. The SSRI frequency distributions of ZZ DH lines in Zhengzhou and Wuhan were similiar and they distorted to silique-shatter susceptible parent Qva. The SSRI distribution of ZZ DH lines belong to muliti-normal distrbution and it was continuous. Therefore silique-shatter resistance trait was controlled by quantitative minor-effect additive genes and environmental factors affected it.4.2A linkage map with175polymorphic loci and19linkage groups was constructed, which covered1382.8cM and its mean marker interval was7.9cM. The genetic distance of linkage groups ranged from33.5cM to107.7cM and the number of markers in linkage groups varied from4to20. 4.3QTL analysis of silique-shatter resistance was conducted with composite interval mapping method of Windows QTLCart V2.5software and the mean SSRI in Zhengzhou and Wuhan. Nine QTLs were obtained in Zhengzhou and four QTLs were obtained in Wuhan(LOD>2.0). In Zhengzhou,9QTLs were located in N1, N7, N8, N15and N18linkage groups, respectively. These QTLs could explain49.0%phynotype. There were epistatic interactions among these QTLs and their contribution to phynotype was about45.9%. These QTLs and their epistatic interactions in Zhengzhou could totally explain94.9%variance. In Wuhan,4QTLs were located in N1, N4, N7and N18linkage groups, respectively.These QTLs could explain38.6%phynotype. There were also epistatic interactions among these QTLs and their contribution to phynotype was about12.8%. These QTLs and their epistatic interactions in Wuhan could totally explain51.4%variance.4.4There were three QTLs which were detected both in Wuhan and in Zhengzhou. qSSRI2detected in Zhengzhou and qSSRI10detected in Wuhan were all located in N1linkage group and their positions were adjacent. qSSRI4from Zhengzhou and qSSRI12from Wuhan were all located in N7linkage group. Their positions were also adjacent. qSSRI9from Zhengzhou and qSSRI13from Wuhan were located in N18linkage group. Their positions were also adjacent.4.5The results of QTL mapping in this study were consistent with the results of association analysis of Raman et al (2011).And the results of Raman et al (2011) aggreed with the findings of a comprehensive transcriptome analysis of silique development and dehiscence in Arabidopsis and Brassica(Jaradat et al.,2010).