Precursor-product relationships between thymidine nucleotides and DNA in mammalian cells. II. Studies with dTDP and dTTP on cells partially lysed by Brij-58

Abstract

Cells from Chinese hamster ovary cell cultures were partially lysed with Brij-58 and incubated in a reaction mixture containing deoxyribonucleoside triphosphates and permitting semiconservative DNA replication. The reaction mixture was supplemented with [3H]dTTP and [14C]dTDP or, alternatively, with [14C]dTTP and [3H]dTDP, and at different times, 3H and 14C specific activities of dTTP and dTDP, as well as 3H/14C ratios in DNA were determined. Isotope ratios in DNA were in good agreement with those expected if dTTP but not dTDP was used as the proximate precursor for DNA synthesis. These data support the conclusion that the thymidine nucleotide used as proximate substrate for DNA replication is dTTP.