Preparation of large quantities of synchronized mammalian cells in late G1 in the pre-DNA replicative phase of the cell cycle

Chinese hamster cells, line CHO, can be synchronized in late G1 by growth in isoleucine-deficient medium for 30 h followed by resuspension in fresh, complete medium containing either hydroxyurea (to 10 −3 M) or cytosine arabinoside (to 5 μg/ml) for 10 h. Cells which are then washed and resuspended in fresh, complete medium without drugs initiate DNA synthesis shortly thereafter, then commence dividing within 7 h. The technique yields large quantities of cells suitable for biochemical studies of events associated with initiation of genome replication and completion of interphase.