Cell culture freezing medium

The fetal thyroid organoid culture medium consisted of Advanced DMEM/F‐12 supplemented with penicillin–streptomycin (Invitrogen), HEPES buffer (10 mm, Gibco), B27 supplement (1×, Invitrogen), GlutaMax (1×, Invitrogen), N‐acetyl‐L‐cysteine (1 mm, Sigma‐Aldrich), retinoic acid (50 ng mL⁻¹, MCE), and GFs mentioned in the figures, R‐spondin1 (200 ng mL⁻¹, OrganRegen), Noggin (100 ng mL⁻¹, OrganRegen), EGF (50 ng mL⁻¹, SinoBiological), FGF 10 (100 ng mL⁻¹, OrganRegen), and A83‐01 (500 nm, Tocris). 10 µm Y‐27632 (Sigma‐Aldrich) was added for primary culture. Forskolin (10 µm, Selleck, S2449) or TSH (0.3 mIU mL⁻¹, Sigma, T9265) were added for stimulating maturation. Fetal thyroid organoids were passaged every 10–14 days. To avoid the influence of passage, the authors used the organoids from passage 2 to passage 3 for RNA‐seq and ATAC sequencing. To prepare frozen stocks, organoids were mixed with Cell Culture Freezing Medium (Gibco) and frozen following standard procedures. Light microscopy images were captured with an Olympus FV3000 camera.