Supplementary Table 1. Numbers of differentially expressed genes

Number of genes
Treatment / Tissue / 3 hours / 6 hours / Total
Desiccation / root / 4337
(5225) / 1697
(2048) / 4892
(5897)
shoot / 9590
(11623) / 10698
(13013) / 12909
(15655)
Salt / root / 122
(141) / 1579
(1880) / 1634
(1946)
shoot / 0
(0) / 0
(0) / 0
(0)
Cold / root / 5529
(6794) / 5388
(6623) / 7814
(9633)
shoot / 2515
(3068) / 5442
(6644) / 6124
(7467)
ABA / root / 3111
(3751) / 8931
(10688) / 9335
(11183)
shoot / 0
(0) / 749
(927) / 749
(927)

The number of probes is in parentheses.

Plant materials

Barley seeds of cultivar “Haruna Nijo” were grown on two layers of No. 2 filter paper (8 cm diameter, ADVANTEC, Tokyo, Japan) wetted with 4.5 ml distilled water containing 0.0125 % (w/v) iminoctadine-triacetate as fungicide in 9 cm Petri dishes sealed with Parafilm (Bemis, Neenah, WI, USA). The dishes were incubated at 24 C under short day conditions (8 h day length) for 4 days. After that, the germinated seedlings were transferred for stress and hormone treatments. For non-treated controls, the seedlings were incubated with distilled water containing the fungicide.

Stress and plant hormone treatments

Desiccation stress

The 4-day-old germinated seedlings were transferred onto dry paper towels on a bench and incubated at room temperature, approximately 23 C, for 3 and 6 hours.

Salt stress

Three 4-day-old germinated seedlings were transferred onto two layers of No. 2 filter paper with 4.5 ml of 0.2 M sodium chloride (NaCl) solution in a Petri dish sealed with Parafilm. The dishes were incubated at 24 C for 3 or 6 hours.

Cold stress

Three 4-day-old germinated seedlings were transferred onto two layers of No. 2 filter paper with 4.5 ml of distilled water in a 9 cm Petri dish sealed with Parafilm. The dishes were incubated at 4 C for 3 or 6 hours.

Abscisic acid (ABA) treatment

Three 4-day-old germinated seedlings were transferred onto two layers of No. 2 filter paper with 4.5 ml of 50 M ABA solution in a 9 cm Petri dish sealed with Parafilm. The dishes were incubated at 24 C for 3 or 6 hours.

Microarray Analysis

Shoots and roots were isolated from the seedlings treated with each stress or hormone condition for 3 or 6 hours. Total RNA was extracted from the shoots and roots using TRIzol reagent (Life technologies, Carlsbad, CA, USA) and RNA quality was checked using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). The RNAs (400 ng aliquots) were labeled with a Quick Amp Labeling Kit (Agilent Technologies, Santa Clara, CA, USA) according to the manufacturer’s instructions. Aliquots of Cy3-labeled (0 hour) and Cy5-labeled (3 and 6 hours) cRNAs (825 ng each) were used for hybridization to the Agilent Oligo Microarray (44K, custom-made; Agilent Technologies, Santa Clara, CA, USA) with 42,491 probes corresponding to the sequences of 36,632 barley full-length cDNA clones (Sato et al. 2009; Matsumoto et al. 2011), which are available from the bex-db (Barley full length cDNA database) site ( Four biological replicate sets of samples were analyzed. After hybridization, microarray slides were scanned (scanner model G2505B; Agilent Technologies, Santa Clara, CA, USA) and data were analyzed using Feature Extraction software (version 9.5; Agilent Technologies, Santa Clara, CA, USA) with the default settings. Subsequently, the data were analyzed using GeneSpring GX 7.3 software (Agilent Technologies, Santa Clara, CA, USA). False positives were controlled by measuring the false discovery rate (Benjamini and Hochberg 1995). This barley Oligo microarray is registered as GPL16130 in Gene Expression Omnibus (GEO) at National Center for Biotechnology Information (NCBI). A complete set of microarray data from this study was deposited to the GEO repository under the accession number GSE41518.