Lactic acid

Trypan blue staining and DAB staining were performed according to Xiao and Chye [68 (link)]. For trypan blue staining, rosettes of 4-, 5-, and 6-week-old wild-type, MACP2-Kos, and MACP2-OE plants were collected in 10 mL tubes and boiled for 5 min in trypan blue staining buffer composed of 12.5% phenol (Thermo, K2599312, Waltham, MA, USA), 12.5% glycerol (Guangzhou Chemical Reagent Factory, Guangzhou, China), 12.5% lactic acid, 48% ethanol (Guangzhou Chemical Reagent Factory, China) and 0.025% trypan blue. The rosettes were incubated for 10 min at room temperature and then decolorized five times in 70% chloral hydrate. For DAB staining, rosettes of 4-, 5-, and 6-week-old wild-type, MACP2-Kos, and MACP2-OE plants were collected and incubated in 1 mg/mL DAB (Sigma, D8001, USA) solution of 10 mM PBS (Vazyme, G101, pH 7.0, Nanjing, China) and 0.05% Tween 20 at 37 °C in darkness overnight and subsequently decolorized in 95% ethanol at 65 °C 3 times every 2 h.

Minimum essential medium (MEM), fetal bovine serum (FBS), penicillin/streptomycin, Hanks Balanced Salt Solution, sodium pyruvate and trypsin were purchased from Gibco (Carlsbad, CA, United States). 5-Alpha-dihydrotestosterone (5α-DHT), collagenase IA, pyridoxal 5-phosphate, O-(carboxymethyl) hydroxylamine hemihydrochloride chloride (AOAA), DL-propargylglycine (PAG), sodium hydrosulfide hydrate (NaHS × H₂O), sodium lactate, L-cysteine (L-Cys), bumetanide, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), glibenclamide (Glib), Iberiotoxin (IbTx) and tetraethylammonium chloride (TEA) were purchased from Sigma-Aldrich (St. Louis, MO, United States). HC067047 was purchased from Tocris (Bristol, United Kingdom). NaCl, KCl, MgSO₄, FeCl₃, HCl, BaCl₂, NaHCO₃, KH₂PO₄, CaCl₂, glucose, mannitol, lactic acid, normal saline, N,N-dimethyl-p-phenylenediamine sulfate, trichloroacetic acid, and zinc acetate were purchased from Guangzhou Chemical Pharmaceutical Factory (Guangzhou, China). Universal two-step detection kit (PV-9000) and DAB detection kit were purchased from ZSBIO (Beijing, China). Fluo-3 AM was purchased from Molecular Probes (Eugene, OR, United States). PBFI AM was purchased from Cayman Chemical (Ann Arbor, MI, United States).