Genome-Wide Association Study Of Microelement In Grain Of Wheat Variety Population And Screening Of Excellent Germplasm

As the largest grain ration crop of the word,wheat is an important source of food and nutrition for human beings.With the development of social economy and the further improvement of living standards,people are increasingly paying attention to the role of microelements in grain crops on human nutrition and health,which puts forward requirements and challenges for improving the microelements in wheat grains.In this study,272 wheat varieties(lines)recently selected from the Huang-Huai wheat region were used as experimental materials to determine the content of iron(Fe),manganese(Mn),copper(Cu),zinc(Zn),selenium(Se),cadmium(Cd),chromium(Cr)and arsenic(As)in wheat grains under three environments in two years by inductively coupled plasma mass spectrometry,and to screen out high-quality wheat varieties(lines)with high microelements contents.Based on the transcriptome data obtained by the group,GWAS and WGCNA analyses are performed to obtain significantly associated markers and hub genes,with a view to discovering molecular markers or candidate genes significantly associated with microelements in wheat grains,and laying a certain foundation for further cloning of related genes and biofortification breeding of wheat microelements.The results are as follows:(1)Determination of microelements in wheat grains and screening of excellent germplasms.The 272 wheat varieties(lines)were planted in Tai’an and Rizhao in 2020-2022,respectively,and the contents of Fe,Mn,Cu,Zn,Se,Cd,Cr and As in wheat grains are determined by inductively coupled plasma mass spectrometry.Phenotypic description statistical analysis showed that there were significant differences in the content of 8 microelements among 272 wheat varieties(lines),with a coefficient of variation of 9.63-51.45%.Among them,the grain Cr content(GCr C)variant is the largest,and the grain Zn content(GZn C)variant is the smallest.Correlation analysis shows that there is a significant positive correlation between the contents of five beneficial microelements(Fe,Mn,Cu,Zn and Se),while the contents of three heavy metal elements(Cd,Cr and As)mostly show a negative correlation.According to the microelement content of different varieties(lines),a total of 50 materials with high beneficial microelement content were selected,11 materials with high beneficial microelement content and 13 materials with high beneficial microelement and low heavy metal element content were selected.(2)Genome-wide association study of grain microelement content and prediction of candidate genes.Using TASSEL 5.0 software,a mixed linear model(MLM+PCA+K)was used.Genome-wide association analysis was performed for the microelement content of grains by combining the genotype data of 176,357 SNP/In Del markers obtained in the previous study.The results showed that a total of 4483 loci were detected,which were significantly correlated with the content of 8 microelements in grains at the p<0.001 level,involving 3551 markers.Among them,there are 439,443,418,466,586,401,631,and 253 markers significantly associated with grain Fe content(GFe C),grain Mn content(Mn),grain Cu content(GCu C),grain Zn content(GZn C),grain Se content(Se),grain Cd content(GCd C),grain Cr content(GCr C),and grain As content(GAs C),respectively.Further screening identified stable pleiotropic markers detected in the 2+AV environment,include 152 stable markers significantly associated with only a single element and 31 pleiotropic markers significantly associated with more than 2 elements at the same time.Based on the annotation results of the Chinese spring genome,the five genes with the highest number of significantly associated markers co-located within the same candidate gene interval were selected,including Traes CS7B02G056600,Traes CS2B02G088600,Traes CS5B02G530500,Traes CS6B02G452200 and Traes CS7A02G564300.Based on the genotypic data of 272 wheat varieties(lines)for haplotype analysis,all the above five candidate genes classified the test materials into two haplotypes,which differed significantly in GSe C,GMn C,GCr C,GZn C and GCd C,respectively.(3)Weighted gene co-expression network analysis.FPKM normalization was performed on the transcriptome data of 272 wheat varieties(lines),and the genes with low confidence,average standard deviation less than 10% and outliers were filtered out,resulting in 12,579 genes for WGCNA analysis.Through analysis and screening,19 modules were identified,and GO enrichment and KEGG pathway analysis were performed on the red and blue modules,resulting in a total of 10 hub genes.Among them,only one gene was detected repeatedly in GWAS.Traes CS7B02G105600 research found that it is related to Cd stress.At the same time,using the National Rice Data Center,it is found that the rice homologous genes of these genes play a role in calcium content and low temperature stress resistance.