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Transient transfections

MK Menuka Karki
NK Neda Keyhaninejad
CS Charles B. Shuster
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RPE1 cells were transiently transfected with eGFP-tagged centrin-2 (a gift from Erich Nigg, Addgene plasmid # 41147) for 6 h at a final concentration of 1 μg ml−1 by using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s specifications. Transfection of small interfering RNA (siRNA) was carried out using DharmaFECT-1 transfection reagent and non-targeting control siRNA (siGENOME control siRNA #1, D001210-01-05) or ESPL1 separase siRNA (5′-GCUUGUGAUGCCAUCCUGAUU-3′) (Dharmacon, Lafayette, CO) according to the manufacturer’s recommendations. Following siRNA transfection, cells were released into media overnight followed by cell synchronization as described above.

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