Ms salts

Seeds of Arabidopsis accessions and T-DNA mutants were purchased from ABRC (Arabidopsis Biological Resource Center). Seeds were surface-sterilized with 5% (v/v) sodium hypochlorite and 0.05% (v/v) Tween 20 for 5 min and were placed on half strength Murashige and Skoog medium (0.23% (w/v) MS salts (Fujifilm Wako Pure Chemical, Osaka, Japan), 0.5% (w/v) sucrose, and 0.03% (w/v) gellan gum (Sigma-Aldrich, MO, USA), pH 5.8). After seeds for sowing were stored at 4 °C for 2 days in the dark, plates were placed in a vertical position under continuous light conditions at 22 °C to allow the seedlings to grow onto the surface of the medium.

Arabidopsis thaliana ecotype Columbia-0 (Col-0) was used throughout the experiments. Seeds of vip1 (SALK_001014C) [13] (link) were obtained from ABRC (Arabidopsis Biological Resource Center, https://abrc.osu.edu/). The T-DNA insertion was confirmed by genomic PCR analysis as previously described [13] (link). Seeds were surface sterilized and sown on 0.8% agar containing 0.5× MS salts (Wako), 1% (w/v) sucrose and 0.5 g/l MES, pH 5.8, with 0, 200 or 250 mM mannitol or 0.25 µM ABA, chilled at 4°C in the dark for 48 h (stratified), and germinated at 22°C. Plants were grown at 22°C under 16-hour light/8-hour dark condition (light intensity 120 µmol·m⁻²·s⁻¹). Arabidopsis transformation was performed by the Agrobacterium-mediated floral-dip method [19] (link), and T2 generation plants were used for analyses. GFP fluorescence in individual plants was checked immediately before measuring cotyledon lengths to confirm the expression of the transgenes.