All cell lines used in this study were derived from HeLa Flp‐in cells (a gift from S Taylor, University of Manchester, UK) (Tighe et al, 2008 (link)), which were authenticated by STR profiling (Eurofins). Cells were cultured in full‐growth media—DMEM supplemented with 9% FBS and 50 μg/ml penicillin/streptomycin. While doing fluorescence time‐lapse analysis, cells were cultured in Leibovitz's L‐15 media (900 mg/L D+ galactose, 5 mM Sodium pyruvate and no phenol red) supplemented with 9% FBS and 50 μg/ml penicillin/streptomycin, or DMEM (no phenol red) supplemented with 9% FBS and 50 μg/ml penicillin/streptomycin. Every 4–8 weeks, cells were screened to ensure a mycoplasma‐free culture.
Doxycycline (1 μg/ml), STLC (S‐trityl‐L‐cysteine: 10 μM), thymidine (2 mM), nocodazole (3.3 μM), MG132 (10 μM) and the MPS1 inhibitor AZ‐3146 (2.5 μM) were purchased from Sigma Aldrich; puromycin (1 mg/ml) and hygromycin B (200 μg/ml) from Santa Cruz Biotechnology; RO‐3306 (10 μM) from Tocris; the PLK1 inhibitor BI‐2536 (100 nM) from SelleckBio; the BUB1 inhibitor BAY‐1816032 (5 μM) from MedChemExpress; the SiR‐DNA far‐red DNA probe (1:10,000) from Spirochrome.
Doxycycline (1 μg/ml), STLC (S‐trityl‐L‐cysteine: 10 μM), thymidine (2 mM), nocodazole (3.3 μM), MG132 (10 μM) and the MPS1 inhibitor AZ‐3146 (2.5 μM) were purchased from Sigma Aldrich; puromycin (1 mg/ml) and hygromycin B (200 μg/ml) from Santa Cruz Biotechnology; RO‐3306 (10 μM) from Tocris; the PLK1 inhibitor BI‐2536 (100 nM) from SelleckBio; the BUB1 inhibitor BAY‐1816032 (5 μM) from MedChemExpress; the SiR‐DNA far‐red DNA probe (1:10,000) from Spirochrome.
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