Whole extracts were prepared for 1 million cells after flow cytometry sorting. Cells were pelleted and resuspended in 50 μl of PBS and 50 μl of 2× Laemmli buffer [0.1 M tris (pH 6.8), 2% SDS, and 5% glycerol] supplemented with protease and phosphatase inhibitors [1× EDTA-free inhibitor cocktail (Roche), 1 mM phenylmethylsulfonyl fluoride, 5 mM NaF, and 2 mM Na3VO4]. Alternatively, 1 million cell cycle–sorted FUCCI wild-type cells subjected to cell fractionation as previously described in (23). Equivalent amount of cells was loaded for Western blots. Western blotting was carried out using standard procedures. Quantification of band intensity and normalization with LaminB was carried out using ImageJ.

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