Limited pre-RC re-assembly occurs in the absence of geminin.
(A) Chromatin from control RNAi (pUC), G1/S arrested (HU), and geminin depleted re-replicating cells (Gem) was fractionated biochemically and immunoblotted for MCMs and ORC2. Levels of ORC and MCMs are also shown for whole cell extracts (WCE). (B) Fraction of control or geminin depleted cells exhibiting active DNA synthesis (EdU incorporation), with error bars indicating standard error. DNA synthesis was determined by pulsing the cells for 30 minutes with EdU at the given time points. At least 200 cells were counted in three independent experiments. (C) Time course of the fraction of control (open diamond) or geminin depleted (open square) cells exhibiting nuclear MCM localization following RNAi treatment for the indicated duration, with error bars indicating standard error. At least 200 cells were counted in three independent experiments.