The presence of single-stranded regions in mammalian DNA

Benzoylated naphthoylated DEAE-cellulose chromatography has been used in this study to demonstrate the presence of single-stranded regions in the DNA of mouse L cells and Chinese hamster ovary cells. In mouse L cell DNA the majority of the single-stranded regions were found to have free ends, as judged from their susceptibility to S₁ endonuclease and exonuclease VII. They were separated on average by a distance of 70 μm.The single-stranded regions fell into two classes, distinguished by whether or not they were derived from the replication fork. Pulse-label administered to Chinese hamster ovary cells was found to be closely associated with the single-stranded regions, as shearing the DNA left a greater proportion of the pulse-label than the bulk-label in the single-strand-containing fraction. However, reasonable estimates of the number of replication forks present at one time showed that the single-stranded regions occurred too frequently along the DNA for them all to be associated with replication. When DNA was isolated from Chinese hamster ovary cells synchronized in G₁ phase, it too was found to contain single-stranded regions, separated by a distance of 100 μm. The regions in the G₁ cells were demonstrated not to be the result of a low level of DNA replication nor to be an artifact of the isolation procedure. Possible functions of this non-replicative class of single-stranded regions are discussed. They may, for example, act as regulatory signals or to be pre-formed initiation points for DNA replication. They are the subject of ongoing investigation.