Density gradient analysis of newly replicated DNA from synchronized mouse lymphoma cells

Abstract

The replication of DNA in synchronous cultures of mouse lymphoma cells was investigated by use of CsCl density gradient centrifugation. We found that the buoyant density of newly replicated DNA depended upon the particular stage of S phase in which synthesis occurred. In early S phase, newly replicated DNA exhibited buoyant densities which were slightly higher, on the average, than that of pre-existing DNA. As S phase progressed, newly replicated DNA shifted to lower buoyant densities, until, near the end of S phase, densities less than pre-existing DNA were observed. These observations are discussed in terms of their possible relevance to base compositional differences between nucleotide sequences made in early as opposed to middle or late S phase. Few, if any, satellite DNA sequences were synthesized in early S phase; however, satellite replication was evident by middle S phase. Although replication of the satellite appeared to continue into late S phase, synchrony at this point in the cell cycle is probably insufficient to be certain whether the synthesis of satellite involves the later one-fifth of S phase.