Figure 5. SV40 DNA replication in Hepes-containing permeabilization medium.

SV40-infected CV1 cells were permeabilized either in standard permeabilization medium containing glutamate or in a medium containing Hepes instead of glutamate; the cells were incubated normoxically thereafter. Succinate (20 mM) was added to one (Hepes) culture, 15 min after permeabilization. All cultures were labelled for 5 min with [α-³³P]dATP (10 μCi/ml, 0.1 μCi/nmol dATP), 30 min after permeabilization, and processed for alkaline sedimentation on a 15–30% sucrose gradient. Sedimentation was from left to right; the top and bottom of the gradients are indicated. Total incorporation (fractions 5–20) was 2137 (glutamate buffer), 1863 (Hepes buffer) and 2870 c.p.m. (Hepes buffer+succinate).