GWAS identifies novel loci linked to seedling growth traits in highly diverse barley population under drought stress

Abstract

Climate changes refer to long-term shifts in temperature and weather patterns that may cause drought, one of the major stresses hindering seed germination, plant growth, and crop productivity. Barley (Hordeum vulgare L) is considered one of the most drought-stress-tolerant cereals and may be used for elucidating genes for drought tolerance at seed germination and seedling stages that would pave the way toward improving the performance of all cereals. The current study was performed at IPK-Gatersleben (Germany) in 2023. Our aim was to explore the genetic basis of germination and seedling traits under drought stress (20% PEG₆₀₀₀ treatment) in a 198 global spring barley collection genotyped with 38,632 SNPs via Genotyping by Sequencing (GBS). The drought treatment significantly reduced the seed germination parameters and seedling traits in the genotypes of a global barley collection. Drought tolerance indices (DTI) for the measured germination and seedling traits indicate delayed and lower germination speed under drought stress than the control. The shoot fresh weight was the most affected trait, with a DTI of 37.4, followed by the seedling fresh weight SDLFW (DTI = 46.3) and root fresh weight (DTI = 47). In contrast, the root length DTI was the least affected trait by drought (78.2), followed by RDW DTI (72.8). GWAS was conducted using single-locus (CMLM) and multi-locus models (MLMM, Farm-CPU, BLINK), with significant marker-trait associations determined at -log10 (1.29E^-06) ≥ 5.88. In our present study, we identified 79 highly significant SNPs distributed across the seven barley chromosomes related to the germination and seedling growth parameters under both control and drought conditions. Gene annotation of these highly significant SNPs revealed that 35 SNPs were in the exonic regions of genes that play roles in important plant biological and physiological processes. Further analysis exhibited 35 high-confidence candidate genes influencing barley germination and seedling growth under control and drought conditions. These genes represent promising targets for breeding and genetic enhancement efforts to improve drought tolerance in barley, potentially extending these benefits to other cereal crops.

Keywords: Barley; Climate change; Drought stress; Drought tolerance index; GWAS; PEG.

Fig. 1

(a–f) Boxplots for germination traits in 198 barley genotypes under control and drought stress conditions (PEG 20%). (a) Germination percentage after 2 days; (b) Germination percentage after 4 days; (c) Germination percentage after 6 days; (d) Germination percentage after 8 days; (e) Germination percentage after 10 days; (f) Germination speed. Significance level: *p < 0.05; **p < 0.01, ***p < 0.001; ****p < 0.0001.

Fig. 2

(a–h). Boxplots of seedling growth parameters in 198 barley genotypes under control and drought stress conditions (PEG 20%). (a) Seed weight (b) Seedling fresh weight (c) Shoot length (d) Shoot fresh weight; (e) Shoot dry weight; (f) Root weight; (g) Root fresh weight; (h) Root dry weight. Significance level: *p < 0.05; **p < 0.01, ***p < 0.001; ****p < 0.0001.

Fig. 3

Histogram illustrating the impact of simulated drought imposed by 20% PEG on germination rate, germination speed and the measured seedling traits under drought expressed as DTI calculated as a percentage of their values under the control conditions for all barley genotypes.

Fig. 4

Pearson correlation coefficients of BLUEs values of the studied traits between (A) control and (B) drought (PEG20%) in 198 barley genotypes. The degree of significance for all correlations was P < 0.001. The color reflects the strength of the correlation. Black crosses indicate non-significant correlations.

Fig. 5

Manhattan plots generated with MLMM, Farm-CPU, CMLM, and BLINK models showing the distribution of SNPs associated with G% after 2 days (A), 4 days (B). 6 days (C). 8 days (D), germination speed (E) seed weight (F), shoot length (G) and root length (H) under the control conditions and drought stress conditions imposed by 20% PEG solution for the 198 barley accessions used in the current study.