DNA synthesis and proliferation of human lymphocytes in vitro. II. Characterization of the DNA newly synthesized after phytohemagglutinin stimulation.

DNA newly synthesized by phytohemagglutinin-(PHA) stimulated human lymphocytes has been analyzed for the possibility that all DNA synthesis may not represent premitotic genome duplication. Equilibrium density gradient characterization of bromodeoxyuridine-(BUdR) substituted DNA demonstrates semi-conservative DNA replication, without evidence for repair synthesis. Experiments to detect selective replication (amplification) of a portion of the DNA involved measurement of reassociation kinetics as well as measurement of the kinetics of BUdR appearance in prelabeled DNA. The presence of large quantities of amplified DNA has been excluded, although amplification cannot be ruled out completely by these techniques. Finally, the kinetics of DNA release from dead lymphocytes is characterized, and factors tending to reduce the complexity of released, labeled DNA are identified.