QTL Mapping Of Silique And Seed Related Traits Including Oil Content In Brassica Napus L.using A High Density Sequence-based Linkage Map

Rapeseed(Brassica napus L.)is a major oil crop around the world.It is not only a leading source for vegetable oil,but also an important source for industrial materials such as biofuel and lubricants.In China,oilseed rape is the most important oil crop and provides about 40% of the vegetable oil supply.Seed-related traits,including seed oil content(OC),silique length(SL),seeds per silique(SS),and seed weight(SW),are primary targets for oil yield improvement.However,these four traits are controlled by multiple genes and environmentally sensitive,and the genetic basis of these traits are largely unknown at present.To dissect the genetic basis of these traits,we developed a population consisting of 192 recombinant inbred lines(RILs)derived from a cross between two parents with distinct OC and silique length.The RILs were grown in three environments including two winter environments and one summer environment,and OC,SL,SS and SW were surveyed in these three environments.Quantitative trait locus(QTL)analysis was conducted for these traits with a high-density genetic linkage map containing 1526 sequence-based molecular markers.A total of 37 consensus QTLs were detected for these traits.The main results are as follows:Genomic DNA from the parents and RILs was double digested with SacI and MseI.Restriction fragments were ligated to SacI and MseI adaptors containing barcode sequences.Fragments in the size range of 400–450 bp were recovered and amplified with PF and PR primers and submitted to sequence on the HiSeq2000 platform.A total of 68,450,311 high-quality PE90 reads were obtained for the 192 RILs after quality filtering.A total of 1,812 polymorphic RAD tags were identified.A subset of 1,700 polymorphic RAD markers containing 3,765 SNPs was identified.A genetic linkage map containing 1,329 RAD markers,107 INDELs and 90 SSRs were constructed.The linkage map was consisted of 19 linkage groups(LGs)and covered a total length of 1610.4 cM,with an average of 1.06 cM between adjacent loci.Alignment of “352” short reads to the B.napus reference genome indicated that homeologous nonreciprocal transposition(HNRT)or homeologous exchange(HE)from A2 to C2 occurred in the genome of “352”.HNRTs resulted in the presence of a duplication of a chromosomal region and loss of the corresponding homeologous region.These HNRTs increased the level of heterozygosity in the RILs,and thus affected our discovery and genotyping of SNPs.The sequences of all loci on the genetic linkage map were aligned to the B.napus reference genome sequence,Alignments indicated that all LGs had a good colinearity with the B.napus reference genome ?Darmour-bzh? with several minor inconsistencies.The RILs derived from the cross between “M201” and “352” were also grown in each of the three environments,and the four traits including OC,SL,SS and SW were investigated.Two-way ANOVA of individual traits across environments indicated that the genotypes of RILs(G),growing environment(E)and genotype-environment interactions(G × E)had significant effects on the four traits.In addition,the environments also have significant effects on the performance of these four traits.The broad-sense heritability of OC,SL,SS and SW was 78.8%,89.4%,78.5% and 81.9%.Genome-wide QTL analysis was conducted using the phenotypic data of all four traits.A total of 54 QTLs for OC,SL,SS and SW were detected in the three environments.Meta-analyses integrated these 54 QTL into 37 consensus QTL.A total of 18 QTLs were identified that controlling seed oil content.These QTL individually explained 4.7–12.8% of the phenotypic variation.Eleven QTL were detected for silique length.These QTL individually explained 4.2–9.5% of the phenotypic variation.A total of 13 QTL were detected for seed number per silique.These QTL individually explained 5.4–8.5% of the phenotypic variation.Twelve QTL were detected for seed weight.Individual QTL explained 3.1–11.7% of the phenotypic variation of SW.We examined the confidence intervals of consensus QTL for different traits by meta–analysis.overlapped consensus QTLs were integrated into seven unique QTL.A significant finding of this study was that one consensus QTLs(cqOCA10b)for OC were identified in three environments.The OC QTLs cqOCA10 b located on A10 were strongly expressed across the three environments.The interval between markers M174 and M782(representing an approximately 6 cM region of chromosome A10)represented approximately 613.8 kb of the B.napus genome.