Hordeum vulgare
Protocols: We used as an object barley plants (Hordeum vulgare (L.)), cultivar 'Sebastian'. Treatments included: untreated control ('C'), ABA application alone ('A'), ABA application followed by drought stress ('AD'), and drought stress alone ('D'). Analyses were done at the end of the drought period (75 days, 'T1') and after recovery (85 days, 'T2'). Seeds germinated in Petri dishes were transferred to a soil mix in 13 ×13 ×13 cm pots. Growth was maintained under optimal water conditions (15% v/v) until 60DAS. Starting at 60 DAS, we divided plants into two groups for treatment: one received a 50 µM ABA spray ('A') and the other water ('C'). This led to four treatment designs mentioned above: Optimal water ('C'); ABA-treated under optimal water ('A'); ABA-treated, then drought from 65 DAS-75 DAS ('AD') and Drought from 65 DAS-75 DAS ('D'). Drought began at 65 DAS (heading stage: Z5.0; Zadok scale). The soil moisture was monitored daily using the TDR EasyTest (Institute of Agrophysics, Polish Academy of Sciences, Poland). In the Control treatment, the soil moisture remained at 15% vwc. Drought lasted for 10 d (soil moisture remained at 1.5-3% vwc), after which moisture was restored to 15% vwc until 85DAS in the recovery phase. Growth was maintained in a controlled greenhouse until harvest. The experimental details are shown in Figure 1. Leaf samples were collected at T1, T2, and T3 for transcriptomic analysis. Physiological analyses were performed at the the post-drought (T2) and post-recovery (T3) conditions. Clean reads were mapped to the barley reference transcriptome (BaRTv2.18) using Kallisto v. 0.43.0. Differential gene expression analysis was conducted using 3DRNAseq in R environment. The read counts and TPMs were generated using the tximport R package and Kallisto outputs, respectively. Based on the mean-variance trend, low-expression transcripts and genes were filtered out. Genes were considered to be expressed if specific criteria on counts per million reads (CPM) were met. Normalization of read counts was performed using the TMM method, and PCA indicated no distinct batch effects. We used as an object barley plants (Hordeum vulgare (L.)), cultivar 'Sebastian'. Treatments included: untreated control ('C'), ABA application alone ('A'), ABA application followed by drought stress ('AD'), and drought stress alone ('D'). Analyses were done at the end of the drought period (75 days, 'T1') and after recovery (85 days, 'T2'). Seeds germinated in Petri dishes were transferred to a soil mix in 13 ×13 ×13 cm pots. Growth was maintained under optimal water conditions (15% v/v) until 60DAS. Starting at 60 DAS, we divided plants into two groups for treatment: one received a 50 µM ABA spray ('A') and the other water ('C'). This led to four treatment designs mentioned above: Optimal water ('C'); ABA-treated under optimal water ('A'); ABA-treated, then drought from 65 DAS-75 DAS ('AD') and Drought from 65 DAS-75 DAS ('D'). Drought began at 65 DAS (heading stage: Z5.0; Zadok scale). The soil moisture was monitored daily using the TDR EasyTest (Institute of Agrophysics, Polish Academy of Sciences, Poland). In the Control treatment, the soil moisture remained at 15% vwc. Drought lasted for 10 d (soil moisture remained at 1.5-3% vwc), after which moisture was restored to 15% vwc until 85DAS in the recovery phase. Growth was maintained in a controlled greenhouse until harvest. The experimental details are shown in Figure 1. Leaf samples were collected at T1, T2, and T3 for transcriptomic analysis. Physiological analyses were performed at the the post-drought (T2) and post-recovery (T3) conditions. We used as an object barley plants (Hordeum vulgare (L.)), cultivar 'Sebastian'. Treatments included: untreated control ('C'), ABA application alone ('A'), ABA application followed by drought stress ('AD'), and drought stress alone ('D'). Analyses were done at the end of the drought period (75 days, 'T1') and after recovery (85 days, 'T2'). Seeds germinated in Petri dishes were transferred to a soil mix in 13 ×13 ×13 cm pots. Growth was maintained under optimal water conditions (15% v/v) until 60DAS. Starting at 60 DAS, we divided plants into two groups for treatment: one received a 50 µM ABA spray ('A') and the other water ('C'). This led to four treatment designs mentioned above: Optimal water ('C'); ABA-treated under optimal water ('A'); ABA-treated, then drought from 65 DAS-75 DAS ('AD') and Drought from 65 DAS-75 DAS ('D'). Drought began at 65 DAS (heading stage: Z5.0; Zadok scale). The soil moisture was monitored daily using the TDR EasyTest (Institute of Agrophysics, Polish Academy of Sciences, Poland). In the Control treatment, the soil moisture remained at 15% vwc. Drought lasted for 10 d (soil moisture remained at 1.5-3% vwc), after which moisture was restored to 15% vwc until 85DAS in the recovery phase. Growth was maintained in a controlled greenhouse until harvest. The experimental details are shown in Figure 1. Leaf samples were collected at T1, T2, and T3 for transcriptomic analysis. Physiological analyses were performed at the the post-drought (T2) and post-recovery (T3) conditions. Clean reads were mapped to the barley reference transcriptome (BaRTv2.18) using Kallisto v. 0.43.0. Differential gene expression analysis was conducted using 3DRNAseq in R environment. The read counts and TPMs were generated using the tximport R package and Kallisto outputs, respectively. Based on the mean-variance trend, low-expression transcripts and genes were filtered out. Genes were considered to be expressed if specific criteria on counts per million reads (CPM) were met. Normalization of read counts was performed using the TMM method, and PCA indicated no distinct batch effects. For RNA-seq analyses, plant tissue (2 cm long fragments of the third leaf located 3 cm below the leaf tip) was collected in four biological replicates, each containing fragments from 3 independent seedlings. Total RNA from cv. 'Sebastian' leaf tissue was extracted using the miRvana isolation kit (ThermoFisher Scientific, USA) per manufacturer guidelines. The concentration and quality were assessed using a NanoDrop (ND-1000) spectrophotometer, and RNA integrity was verified using an Agilent 2100 Bioanalyzer with an RNA 6000 Nanochip.  cDNA libraries prepared via Illumina TruSeq procedures were sequenced on an Illumina Novaseq 6000 sequencer at Macrogen: 2 × 150 bp PE reads, six samples per lane, and a full flow cell. The dataset had four replicates for both the control and drought conditions.
organism
Hordeum vulgare
organism
Hordeum vulgare
collection date
not collected
stimulus
drought
scientific_name
Hordeum vulgare
common name
barley
organism part
leaf
developmental stage
heading
genotype
wild type genotype
isolate
not applicable
cultivar
Sebastian
age
75
geographic location (country and/or sea)
not collected