PMKs were isolated from newborn 2- to 3-day-old C57Bl/6 mice, as described previously (Lewis et al., 2014 (link), Mardaryev et al., 2016 (link)). PMKs were grown in EMEM calcium-free medium (Lonza, Wolverhampton, UK) with supplements (0.05 mM calcium, 4% fetal bovine serum, 0.4 μg/ml hydrocortisone, 5 μg/ml insulin, 10 mg/ml epidermal growth factor (EGF), 10−10 M cholera toxin, 2 × 10−9 T3, 2 mM l -glutamine, 100 U/ml penicillin, and 100 μg/ml streptomycin) at 33 °C, 8% CO2 until 60–70% confluent. PMKs were transfected with 100 nM p63siRNA or control siRNA using Lipofectamin 2000 (Life Technologies).
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