Studies on the replication of Escherichia coli phage λ DNA I. The kinetics of DNA replication and requirements for the generation of rolling circles

Escherichia coli phage λ DNA has been isolated from infected bacteria using a new technique by which virtually all phage DNA is recovered. Isolated DNA is examined by electron microscopy. Addition of φX174 RF1 molecules as a counting standard enables us to determine the average number of λ DNA molecules present in an infected cell. In this study, we have followed the kinetics of λ DNA replication and examined rolling circle replication. The most important findings are the following: (1) Rolling circle replication is initiated at roughly the same time as is θ replication, indicating that the rolling circle is not solely a late-replicating form. (2) θ replication stops at about 16 min after infection. (3) Early in infection the number of DNA molecules per cell doubles every 2–3 min until θ replication stops, at which point most DNA synthesis consists of growth of the tails of about three rolling circles per cell. (4) Neither the timing of rolling circle replication nor the number of molecules is affected by the activity of the λ red genes. (5). The red genes are responsible for the production of oligomeric circles late in infection.