title: hESC_Chir3d
organism: Homo sapiens
description: Protocols: Cells were detached after 4 hours, 24 hours, and 72 hours of CHIR treatment (n = 6 wells per time point). The cell suspensions were filtered through a 40 mm cell strainer (KKE3.1, Carl Roth), and cell concentrations were evaluated using a hemocytometer in a Leica DM 204 IL LED microscope and adjusted to 1'000 cells/µl. hESCs were cultured in Essential 8TM medium (A1517001, ThermoFisher Scientific) in plates and flasks coated with 10 mg/ml Vitronectin (A14700, ThermoFisher Scientific). Cells were passaged by incubation in 0.5 mM EDTA-PBS (stock: 0.5 M EDTA, AM9260G, Thermo Fisher Scientific). Cells were treated with 10 mM GSK-3 inhibitor CHIR99021 (SML1046-25MG, Sigma-Aldrich) for 4 hours, 24 hours, and 72 hours (n = 6 wells per time point). Single cells were run in a 10X Chromium Controller and samples were further processed according to the manufacturer's indications (10X Genomics; v3 protocol). Library preparation was performed according to the manufacturer's indications (10X Genomics; v3 protocol)