Genome-Wide Association Study of Xian Rice Grain Shape and Weight in Different Environments

Abstract

Drought is one of the key environmental factors affecting the growth and yield potential of rice. Grain shape, on the other hand, is an important factor determining the appearance, quality, and yield of rice grains. Here, we re-sequenced 275 Xian accessions and then conducted a genome-wide association study (GWAS) on six agronomic traits with the 404,411 single nucleotide polymorphisms (SNPs) derived by the best linear unbiased prediction (BLUP) for each trait. Under two years of drought stress (DS) and normal water (NW) treatments, a total of 16 QTLs associated with rice grain shape and grain weight were detected on chromosomes 1, 2, 3, 4, 5, 7, 8, 11, and 12. In addition, these QTLs were analyzed by haplotype analysis and functional annotation, and one clone (GSN1) and five new candidate genes were identified in the candidate interval. The findings provide important genetic information for the molecular improvement of grain shape and weight in rice.

Keywords: GWAS; QTLs; grain shape; rice.

Figure 1

Box plots of six traits of rice grain shape and weight under drought stress (DS) and normal water (NW) and phenotypic correlations of the six traits in different environments. 2017DS: 2017 drought stress; 2017NW: 2017 normal water; 2018DS: 2018 drought stress; 2018NW: 2018 normal water. (a) Grain circumference; (b) grain length; (c) grain length to width ratio; (d) grain size; (e) grain width; (f) thousand grain weight; (g) 2017 drought stress; (h) 2017 normal water; (i) 2018 drought stress; (j) 2018 normal water. ‘a’, ‘b’ and ‘c’ are based on whether the t-test is significant between each other. ‘*’, ‘**’, and ‘***’ refer to significant correlations (p < 0.05, p < 0.01, and p < 0.001).

Figure 2

Genetic structure analysis of 275 Xian rice accessions. (a) Phylogenetic tree; each branch corresponds to a rice accession. (b) Principal component analysis on about 0.4 million SNPs in 275 rice accessions. PC1 and PC2 refer to the first and second principal components, respectively. Red points represent the 275 rice accessions, with each point representing one rice accession. A shorter distance between the points indicates a closer relationship. (c) Heatmap of kinship from R Package “pheatmap”. (d) LD decay. Y–axis is the average r² value of each 250−kb region, and X–axis is the physical distance between markers.

Figure 3

Manhattan plots of the GWAS results for GC, GL, GLWR, GS, GW, and TGW obtained with GLM. (a) GC under drought stress; (b) GC under normal water; (c) GL under drought stress; (d) GL under normal water; (e) GLWR under drought stress; (f) GLWR under normal water; (g) GS under drought stress; (h) GS under normal water; (i) GW under drought stress; (j) GW under normal water; (k) TGW under drought stress; (l) TGW under normal water.

Figure 4

Identification of candidate genes for GC. (a) Based on six SNPs in all evaluated rice accessions, four haplotypes of LOC_Os03g29260 were identified. In the gene structure diagram of LOC_Os03g29260, the promoter is indicated by white frame; the exon is represented by blue frame; and the intron and intergenic region are marked by blue lines. A thin black line represents the genomic location of each SNP. Haplotypes with fewer than 10 accessions are not shown. (b,c) Local Manhattan map under drought stress and normal water. Red dotted lines represent candidate regions for associated SNPs. (d) Linkage disequilibrium heatmap. (e,f) Based on GL of LOC_Os06g15480 haplotype under drought stress and normal water, differences between haplotypes were statistically analyzed using Tukey’s test, ‘a’ and ‘b’ are based on whether the t-test is significant between each other.

Figure 5

Dentification of candidate genes for GL. (a) Based on the three SNPs in all evaluated rice accessions, three haplotypes of LOC_Os05g02500 were identified. In the gene structure diagram of LOC_Os05g02500, the promoter is indicated by white frame; the exon is represented by blue frame; and the intron and intergenic region are marked by blue lines. A thin black line represents the genomic location of each SNP. Haplotypes with fewer than 10 accessions are not shown. (b,c) Local Manhattan map under drought stress and normal water. Red dotted lines represent candidate regions for associated SNPs. (d) Linkage disequilibrium heatmap. (e,f) Based on GL of LOC_Os05g02500 haplotype under drought stress and normal water, differences between haplotypes were statistically analyzed using Tukey’s test, ‘a’ and ‘b’ are based on whether the t-test is significant between each other.

Figure 6

Identification of candidate genes for GLWR. (a) Based on the tree SNPs in all evaluated rice accessions, two haplotypes of LOC_Os03g37930 were identified. In the gene structure diagram of LOC_Os03g37930, the promoter is indicated by white frame; the exon is represented by blue frame; and the intron and intergenic region are marked by blue lines. A thin black line represents the genomic location of each SNP. Haplotypes with fewer than 10 accessions are not shown. (b,c) Local Manhattan map under drought stress and normal water. Red dotted lines represent candidate regions for associated SNPs. (d) Linkage disequilibrium heatmap. (e,f) Based on GL of LOC_Os03g37930 haplotype under drought stress and normal water, differences between haplotypes were statistically analyzed using Tukey’s test, ‘a’ and ‘b’ are based on whether the t-test is significant between each other.

Figure 7

Identification of candidate genes for GS. (a) Based on 24 SNPs in all evaluated rice accessions, seven haplotypes of LOC_Os04g08350 were identified. In the gene structure diagram of LOC_Os04g08350, the promoter is indicated by white frame; the exon is represented by blue frame; and the intron and intergenic region are marked by blue lines. A thin black line represents the genomic location of each SNP. Haplotypes with fewer than 10 accessions are not shown. (b,c) Local Manhattan map under drought stress and normal water. Red dotted lines represent candidate regions for associated SNPs. (d) Linkage disequilibrium heatmap. (e,f) Based on GL of LOC_Os04g08350 haplotype under drought stress and normal water, differences between haplotypes were statistically analyzed using Tukey’s test, ‘a’, ‘b’ and ‘c’ are based on whether the t-test is significant between each other.

Figure 8

Identification of candidate genes for GW. (a) Based on 13 SNPs in all evaluated rice accessions, three haplotypes of LOC_Os01g07500 were identified. In the gene structure diagram of LOC_Os01g07500, the promoter is indicated by white frame; the exon is represented by blue frame; and the intron and intergenic region are marked by blue lines. A thin black line represents the genomic location of each SNP. Haplotypes with fewer than 10 accessions are not shown. (b,c) Local Manhattan map under drought stress and normal water. Red dotted lines represent candidate regions for associated SNPs. (d) Linkage disequilibrium heatmap. (e,f) Based on GL of LOC_Os04g08350 haplotype under drought stress and normal water, differences between haplotypes were statistically analyzed using Tukey’s test, ‘a’ and ‘b’ are based on whether the t-test is significant between each other.

Figure 9

Identification of candidate genes for TGW. (a) Based on one SNP in all evaluated rice accessions, two haplotypes of LOC_Os02g15580 were identified. In the gene structure diagram of LOC_Os02g15580, the promoter is indicated by white frame; the exon is represented by blue frame; and the intron and intergenic region are marked by blue lines. A thin black line represents the genomic location of each SNP. Haplotypes with fewer than 10 accessions are not shown. (b,c) Local Manhattan map under drought stress and normal water. Red dotted lines represent candidate regions for associated SNPs. (d) Linkage disequilibrium heatmap. (e,f) Based on GL of LOC_Os02g15580 haplotype under drought stress and normal water, differences between haplotypes were statistically analyzed using Tukey’s test, ‘a’ and ‘b’ are based on whether the t-test is significant between each other.