D1000 hs dna screentape analysis

SMARCA4 (BRG1) levels of the ammonium sulfate nuclear extracts were normalized via BCA protein quantification and silver stain analyses for HA-SS18 and HA-SS18-SSX conditions. Protein was diluted to a final reaction concentration of 150 μg ml⁻¹ in REAA buffer (20 mM HEPES, pH 8.0, 50 mM KCl, 5 mM MgCl₂) containing 0.1 mg ml⁻¹ BSA, 1 mM DTT, 20 nM nucleosomes (EpiDyne nucleosome remodeling assay substrate ST601-GATC1, EpiCypher). The REAA mixture was incubated at 37 °C for 10 min, and the reaction was initiated using 1–2 mM ATP (Ultrapure ATP, Promega) and 0.005 U ml⁻¹ DpnII restriction enzyme (New England Biolabs). The REAA reaction mixture was quenched with 20–24 mM EDTA and placed on ice. Proteinase K (Ambion) was added at 100 mg ml⁻¹ for 30–60 min, followed by either AMPure bead DNA purification and D1000 HS DNA ScreenTape analysis (Agilent) or mixing with GelPilot Loading Dye (QIAGEN) and loading onto 8% TBE gel (Novex 8%TBE Gels, Thermo Fisher). TBE gels were stained with either SYBR-Safe (Invitrogen) or Syto-60 red fluorescent nucleic acid stain (Invitrogen), followed by imaging with ultraviolet light on an Alpha Innotech AlphaImager 2200 and/or with 652-nm light excitation on a Li-Cor Odyssey CLx imaging system (LI-COR).