Genome-wide Association Study Of Seed Oil Content And Leaf Glucosinolate Content In Brassica Napus

Rapeseed(Brassica napus;AACC,2n = 38)is one of the most important oilseed crops worldwide and is the major source of edible oil in China.Seed oil content and glucosinolate content are important quality traits.Enhancement of the seed oil content has been a primary goal for B.napus breeding,and it is therefore crucial to discover novel elite allelic variations and develop high-throughput molecular markers for high oil content breeding.On the other hand,with the success of double-low breeding in rapeseed,leaf glucosinolate content decreased with the reduction of seed glucosinolate content,which made cultivars more susceptible to pest,birds and diseases.It is necessary to breed cultivars with high leaf/low seed glucosinolate content.However,the genetic basis for leaf glucosinolate content remains unclear.Genome-wide association study(GWAS)is an effective way to dissect the genetic basis of complex quantitative traits.In this study,we applied GWAS to dissect the genetic basis for the seed oil content and leaf glucosinolate content.The main results are as follows:1.Population structure,genetic relatedness and linkage disequilibrium of the association panelWe collected 521 accessions from the major breeding institutes across China and genotyped the panel of accessions with a Brassica 60K SNP array.Population structure analysis indicated that the 521 accessions could be divided into two major sub-populations and designated as P1 and P2.The P1 sub-population included 364 inbred lines,and the P2 sub-population included 54 inbred lines.The remaining 103 lines were classified into a mixed sub-population.The analysis of genetic relatedness revealed that most of the accessions in this study were distantly related.Approximately 56.4%of the kinship coefficients between inbred lines were equal to 0,and 22.7%of the kinship coefficients were between 0 and 0.05.We used 25,870 SNPs to measure the LD decay of the association panel.The LD decay rate was measured as the chromosomal distance at which the average pairwise correlation coefficient(r2)dropped to half of its maximum value.The LD decay of the association panel was 0.45-0.50 Mb.A considerably faster LD decay rate was observed for the A-subgenome(0.10-0.15 Mb)compared with the C-subgenome(1.15-1.20 Mb).2.Integration of QTLs for seed oil contentWe collected QTLs for seed oil content from six DH populations,DY,RNSL,Z5,SG,KN and TN,and mapped them to the reference genome of Darmor-bzh.There were 53 unique QTLs for seed oil content,32 QTLs in the A-subgenome and 21 QTLs in the C-subgenome,suggesting that seed oil content is a complex trait controlled by minor-polygene.3.Genome-wide association study for seed oil contentThe seed oil content of the association panel was investigated in three environments.The seed oil content has wide phenotypic variations(>20%),with broad-sense heritability as high as 87.4%.We identified 50 loci significantly associated with seed oil content using three statistical models,the general linear model,the mixed linear model and the Anderson-Darling test.Together,the identified loci could explain 79.7%of the total phenotypic variance,and 21 of these loci were located in previously reported QTL confidence intervals,and 29 of these loci have not been reported previously.4.Verification a novel locus OilA5 with linkage analysis and the cumulative effects of associated loci for the seed oil content in B.napusAll three models identified a locus at approximately 20.3 Mb on chromosome A5 that has not been reported in previous studies.Two allele-specific SNP markers Oil-A and Oil-G were developed according to the lead SNP marker Bn-A05-p22266340.The Oil-A and Oil-G were mapped to the peak of QTL on A5 linkage group of the ZD DH population,and could explain 20%of the total phenotypic variance with an additive effect 0.9,consistent with the GWAS results.We used 22 validated associated loci to understand the effects of allelic variations on seed oil content and found that the associated loci for the seed oil content exhibited cumulative effects,the seed oil content gradually increased with an increasing number of favorable haplotypes.5.Correlation between indolic glucosinolate content and leaf resistance to Sclerotinia sclerotiorumWe selected 34 accessions with low seed glucosinolate content and different leaf glucosinolate content to evaluate the leaf resistance to Sclerotinia sclerotiorum,and found that lesion size did not correlate with leaf-GLS and total aliphatic GLS content,but negative correlated with total indolic GLS content,especially indol-3-ylmethyl GLS content.This indicated that the indolic glucosinolate play important role in B.napus against Sclerotinia sclerotiorum.6.Genome-wide association study for leaf glucosinolate metabolismWe selected 366 accessions from the initial 521 accessions,and analysed leaf glucosinolate content using HPLC across two years.25 glucosinolate traits have wide phenotypic variations,with coefficient of variation from 17.1%to 207.9%,broad-sense heritability from 50.9%to 97.1%.We identified 177 associated loci for 25 glucosinolate traits,wihch could explain 5.70%-42.60%of the total phenotypic variance,respectively.In total,20 associated loci for secondary modifications of GLS were identified.We also identified 8 loci,GLS-A3,GLS-A8,GLS-A9,GLS-A10-2,GLS-C4-1,GLS-C7,GLS-C9-1 and GLS-C9-3,associated with Leaf-GLS,4 loci of them(GLS-A3?GLS-A9?GLS-C7 and GLS-C9-1)also associated with Seed-GLS.We detected GLS-A3 in low seed glucosinolate content panel including 168 accessions,suggesting that GLS-A3 was a major locus to control leaf glucosinolate content in low seed glucosinolate content panel.The locus could increase 0.73 ?mol/g FW of leaf glucosinolate content,accounting for 17.8%of total variation in leaves.