Integrating GWAS And WGCNA To Screen Candidate Genes For Biological Yield Of Brassica Napus L.

Biomass yield is especially important for Brassica napus L.planted in winter,as it is an important basis for high yields of crops.In this study,two-year phenotypic data of natural populations composed of resequencing materials were used for genome-wide association analysis.We integrated the results of transcriptome sequencing(RNA-seq)of biomass yield-extreme materials.A WGCNA network constructed by integrating transcriptome data of six parts of six Brassica napus materials(stalks,leaves,main inflorescence and lateral branch seeds 30 days after anthesis,and main inflorescence and lateral branch keratin 30 days after anthesis).We screened candidate genes related to biological yield,and finally verified the expression of these genes.The main results are as follows:1.Biological yield has positive effect on other economic yieldAccording to the relevant analysis,it was found that biological yield in natural populations of Brassica napus had positive effects on most yield-related traits in the past two years,indicating that the biological yield of Brassica napus is the basis and guarantee for other yield-related traits.2.Detection of SNP loci closely related to yield traitsK + PCA model was the best model for biomass analysis of natural population in two years,and 14 significant loci were detected in the best model(P < 1 / 385692 or P < 0.05 / 385692).They were distributed on A03,A07,C01,C03,C04,C06 and C09 chromosomes respectively,and these loci accounted for 5.64%-7.98% of phenotypic variation.3.Transcriptome differential gene analysis of extreme phenotypesTranscriptome differentially expressed genes were analyzed using six tissues and organs with high biomass yield material "CQ45" and low biological yield material "CQ46".6820 significantly differently expressed genes were detected in leaves,17,309 significantly differently expressed genes were detected in the main inflorescence horns 30 days after flowering,and 5431 significantly differently expressed genes were detected in the lateral branch horns 30 days after flowering.Leaf and keratin are the main photosynthetic organs in rapeseed.The GO and KEGG analysis showed that the differential genes are related to biological functions such as glucosyltransferase activity(GO: 0046527),and the enriched metabolic pathway is mainly concentrated in photosynthesis(GO: 0015979).6948 significantly differently expressed genes were detected in the main inflorescence seeds(30 days after flowering),and 17309 significantly differently expressed genes were detected in the lateral branch seeds(30 days after flowering).The seed is the storage organ of rapeseed,and its GO and KEGG analysis shows that the function of the differential genes is related to microtubule motor activity(GO: 0003777),thioglucosidase activity(GO: 0019137),Starch and sucrose metabolism,and more genes are involved in nutrient reservoir activity(GO: 0045735),cell cycle(GO: 0007049)and other metabolic pathways.12,867 significantly differently expressed genes were detected in the stem.GO and KEGG analysis showed that the differential genes were mainly concentrated in biological processes such as plant-type secondary cell wall biogenesis(GO: 0009834)and cell wall biogenesis(GO: 0042546).4.Through WGCNA analysis,some gene modules with high correlation with tissue location were obtainedAccording to 36 groups of transcriptome data,the mad of each gene was calculated.The first 5% genes with mad value were selected to construct WGCNA(a total of 5052 genes).These genes were analyzed by WGCNA and screened into 15 gene modules.The correlation between the tissues and the modules was calculated.It was found that the co expression modules of five genes were significantly correlated with leaves,stems and seeds 30 days after anthesis.The expression level of me and is also highly related to the overall expression level of the module,indicating the accuracy of the screening and analysis of the target module.Go and KEGG analysis of key modules showed that most of the functions and biological processes involved in these modules were concentrated in photosynthesis,energy metabolism,carbohydrate metabolism and lipid metabolism Metabolism)and other biological processes,which are highly related to the tissue sites that are significantly related to them.5.Integrating the hub gene of key modules in WGCNA,significant SNP locus of association analysis and extreme phenotype difference gene to determine candidate genes.According to GWAS analysis,there were 755 genes of 500 kb before and after SNP locus,27610 genes of transcriptome difference of leaf,stem,main sequence and lateral seed at 30 days after anthesis,and main sequence and lateral pericarp at 30 days after anthesis of extreme phenotype material.178 genes were obtained by combining GWAS related genes and transcriptome differential genes.According to the function of Arabidopsis homologous genes of these genes,the candidate genes for biological yield,BnA07g19320 D and BnC03g33610 D,were screened.The homologous genes of Arabidopsis are cp12-2 and gapc1,which play an important role in energy,carbohydrate metabolism and photosynthesis.At the same time,the top 10 connected genes in the key modules obtained by WGCNA were selected as hub genes,and the homologous genes of these genes in Arabidopsis were found through sequence alignment,and then the function of these homologous genes was sorted out.Finally,the candidate genes of BnC03g73810 D,BnA09g35380D,BnA04g06420 D,BnA04g04350D and BnC08g48810 D were identified by WGCNA.Their Arabidopsis homologous genes are hcef1,Hog1,SBPase and ACT2,which play an important role in the arwen cycle,carbon assimilation and material accumulation of photosynthesis.