Possible effect of the cell synchronization method on the pattern of 5-bromodeoxyuridine incorporation into early synthesized DNA.

Synchronously and asynchronously growing chick embryo fibroblasts have been used to study the pattern of 5-bromodeoxyuridine (BrdU) incorporation into DNA. In the synchronous cell system, the density of unifilarly substituted DNA is about 0.010 g/ml higher during first half of S phase than during second half of S phase. The density of unifilarly substituted DNA isolated from asynchronously growing cells is similar to that of DNA synthesized during the second half of S phase of synchronously growing cells for a given concentration of analogue in culture medium. Reassociation kinetics experiments have shown the oversubstitution to occur at the level of early synthesized repeated and/or intermediate DNA sequences. It is then assumed that the oversubstitution is due to some metabolic changes caused by the synchronization procedure itself. As BrdU incorporation into early replicating DNA is known to induce alterations of the cell metabolism, the implication of this phenomenon is discussed at the level of the inhibition of transformation which takes place when chick embryo fibroblasts are infected with Rous sarcoma virus during G1 and subsequently treated with BrdU during early S phase.