Figure 7.

Cdt1 addition in G2 induces second- and third-round replication. (A) Sperm nuclei were incubated at 5 ng DNA/μl in metaphase-arrested extracts supplemented with BrdUTP, [α-³²P]dATP and CaCl₂. After 90 min, extracts were optionally supplemented with 10 ng/μl recombinant Cdt1 and 5 mM caffeine. After a further 90 min incubation, DNA was isolated and fractionated on CsCl equilibrium density gradients. (B) As (A), except that BrdUTP, Cdt1 and caffeine were all added to extract at 120 min, and the total incubation time before DNA isolation was 300 min.