Epibrassinolide ebl

Seedlings were transferred on solid MS medium with the indicated chemicals: propidium iodide (PI, 10 μM, Sigma-Aldrich or Thermofisher), hydroxyurea (HU, final concentration 5 mM, Sigma-Aldrich) for 24 hours, gibberellic acid (GA, final concentration 10 μM, Sigma-Aldrich) for 1 hour before ablation, paclobutrazol (PAC, final concentration 2 or 10 μM as indicated, Sigma-Aldrich) for 1 hour before ablation, epibrassinolide (EBL, Sigma Aldrich, final concentration 1 μM) for 1 hour before ablation, dexamethasone (DEX, Sigma Aldrich, final concentration 5 μM) for 1 hour before ablation, 1-Naphthylacetic acid f (NAA, Sigma Aldrich, final concentration 1 μM) or 1 hour before ablation, 6-Benzylaminopurine (BAP, Sigma Aldrich, final concentration 50 nM) for 1 hour before ablation.

Chinese kale cultivar “DSCH” germinated and grown in Zhejiang University (Hangzhou, China) was used as material in this study. Chinese kale seeds were sterilized for 30 s in 75% ethanol and washed with sterile water twice, and then immersed in 10% bleach for 2 min, followed by washing with sterile water five times. They were then soaked in sterile-distilled water for 48 h at 25°C in dark. Chinese kale sprouts were cultured in petri dishes with three pieces of wet filter paper in a plant growth chamber at 25°C with a 16-h-light/8-h-dark photoperiod. Five-day-old sprouts and six-month-old Chinese kale plants were used to collect different organs to analyze BoaMYB51 expression patterns. For epi-brassinolide (eBL) (Sigma), methyl-jasmonate (MeJA) (Sigma), SA (Sigma), and flagelin22 (flg22) (Chinese Peptide, Hangzhou, China) treatment, 5-day-old Chinese kale sprouts were grown in sterile half-strength Murashige and Skoog (MS) media at 25°C with a 16-h-light/8-h-dark photoperiod.
Arabidopsis mutant myb34myb51 (Frerigmann and Gigolashvili, 2014 (link)) was used for functional complementation studies. Arabidopsis seeds were sterilized for 12 min in 10% bleach and washed with sterile water 5 times, and then were stratified for 3 days at 4°C. Arabidopsis plants were grown in sterile half-strength Murashige and Skoog (MS) media at 22°C with a 16-h-light/8-h-dark photoperiod.

To prepare stock solutions, epibrassinolide (EBL; Sigma-Aldrich, http://www.sigmaaldrich.com), gibberellin A₃ (GA₃; Sigma-Aldrich), and paclobutrazol (PAC; Duchefa, http://www.duchefa.com) were dissolved in absolute ethanol at 5, 100, and 1mM, respectively. Picloram (PIC; Duchefa) was dissolved in DMSO at 50mM. Propiconazol (PCZ; BannerMax, Syngenta, http://www.syngenta.com) was dissolved in water at 5mM. Stock solutions were kept at –20ºC until use.