Sucrose

3, 3′-dithiobis sulfosuccinimidylpropionate (DTSSP) and beta-D-Lactose were purchased from Thermo Scientific (Pittsburgh, PA). Sucrose was purchased from MP Biomedicals (Solon, OH). Ouabain octahydrate, cis-diammineplatinum dichloride (CDDP), doxorubicin hydrochloride (DXR) and thiazolyl blue tetrazolium bromide (MTT) were purchased from Sigma-Aldrich. Pyrazolo pyrimidine (PP2), a Src kinase inhibitor, was purchased from Tocris Bioscience (Bristol, UK).
Customized polyclonal rabbit anti-Gal-3 antibody was created by Pierce Biotechnology; mouse anti-V5 and purified mouse IgG were purchased from Invitrogen; polyclonal goat anti-Na⁺/K⁺-ATPase alpha1 and monoclonal mouse anti-Mdr-1 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA); polyclonal rabbit anti-Mdr was purchased from Oncogene Research Products (Cambridge, UK); mouse anti-beta-actin was purchased from Sigma-Aldrich; rabbit anti-Src and anti-p-Src were purchased from Cell Signaling Technology (Beverly, MA); purified rabbit IgG was purchased from ZYMED (San Francisco, CA); monoclonal mouse anti-phosphoserine was purchased from abcam (Cambridge, MA).

Seeds of sunflower (H. annuus L.) RHA280 were harvested from plants grown under greenhouse conditions as previously described (Zhang and Finer 2015 (link)), and stored in sealed plastic bags at 4°C in the dark for up to 1 yr. After pericarps were removed manually, high-quality kernels (those without developmental defects or necrotic regions) were used for transformation. Kernels were surfaced sterilized with 5% (v/v) commercial bleach (8.25% [w/v] sodium hypochlorite; Clorox, Oakland, CA) for 20 min, and rinsed with sterilized distilled water 8–10 times. After the embryo axis was removed by making a cut 1–2 mm from the cotyledonary node perpendicular to the longitudinal axis, the remaining cotyledons were immersed in liquid shoot induction medium (SIM). After overnight immersion, the seed coat was easily removed from the cotyledons with minimal damage to cotyledon tissues. SIM was composed of Murashige and Skoog salts (Murashige and Skoog 1962 (link)), Gamborg’s B5 vitamins (Gamborg et al.1968 (link)), 30 g L⁻¹ sucrose (MP Biomedicals, Solon, OH), 1.5 mg L⁻¹ 6-benzylaminopurine, and 0.2 mg L⁻¹ 1-naphthaleneacetic acid. The medium pH was adjusted to 5.7, and sterilized by autoclaving at 121°C for 20 min. All medium components were obtained from Sigma-Aldrich® (St Louis, MO) if not otherwise specified.

RNA for Start-Seq experiments were prepared from control or NF-YA KD cells, as previously described^{62 (link)}. Briefly, mouse ESCs were grown as described for RNA-Seq, and capped RNAs were isolated essentially as previously described^{68 (link)}. In brief, ~2 × 10⁷ ESCs were trypsinized and collected by centrifugation. After washing with ice-cold 1× PBS, cells were swelled in 10 ml of swelling buffer [10 mM Tris (Sigma, T2663) pH 7.5; 10 mM NaCl (Sigma, S5150); 2 mM MgCl₂ (Sigma, M2670); 3 mM CaCl₂ (Sigma, C1016); 0.3 M sucrose (MP Biomedicals, 152584); 0.5% IGEPAL CA-630 (Sigma, I3021); 5 mM dithiothreitol (Sigma, D0632); 1 mM PMSF (Sigma, P7626); protease inhibitors (Roche, 4693159001), SUPERase-IN RNAse inhibitor (Ambion, AM2694)] by incubating for 15 min on ice followed by 14 strokes with a loose pestle. The dounced cells were spun for 5 min at 500x g, the supernatant (cytoplasm) was discarded, the pellet resuspended in 30 ml of swelling buffer, and spun as above. The supernatant was discarded and the nuclei pellet was resuspended in 1 ml of swelling buffer, aliquoted and stored at -80 °C. Libraries were prepared according to the TruSeq Small RNA Kit (Illumina, RS-200–0012). To normalize samples, 15 synthetic capped RNAs were spiked into the Trizol preparation at a specific quantity per 10⁶ cells, as previously described^{69 (link)}.