Extended Data Fig. 8.

a, Cdc7^AID/AID/Tir1 ESC (upper panel) or Cdk1^AS/AS ESC (lower panel) were synchronized in mitosis by nocodazole and released as in Fig. 3i and Extended Data Fig. 7h, i. Two hours after the release, Cdc7^AID/AID/Tir1 ESC were treated for 2 h with vehicle (Control), auxin (IAA), Cdk1 inhibitor Ro-3306, or with both compounds. Cdk1^AS/AS ESC were treated for 2 h with vehicle, Cdc7 inhibitor XL-413, 3MB-PP1 (to inhibit Cdk1), or with both compounds. b, Upper panel: asynchronously growing Cdc7^AID/AID/Tir1 ESC were treated for 2 h with vehicle, auxin, Ro-3306 or with both compounds. Lower panel: Cdk1^AS/AS ESC were treated for 2 h with vehicle, XL-413, 3MB-PP1, or with both compounds. c, Cdc7^AID/AID/Tir1 MEFs were treated for 2 h with vehicle, auxin, Ro-3306 or with both compounds. d, MCF10A, HMEC, BJ fibroblasts and primary human dermal fibroblasts (HDF) were treated for 2 h with vehicle, XL-413, Ro-3306 or with both compounds. a-d, The endogenous Cdk2 was immunoprecipitated (IP-Cdk2) from treated cells and used for kinase reactions with [γ³²P]-ATP and histone H1 as a substrate. IgG, control immunoprecipitation with IgG. Note that treatment of cells with any of these inhibitors (either singly, or in combination) did not decrease the activity of the endogenous Cdk2. a-d, representative results (out of 2).