UVC-Induced DNA Replication Dynamics

DNA combing was performed according to our previously described protocol [12 (link)] with modifications. Briefly, cells were irradiated with 15 J/m² UVC and treated or not with C11 (0.5 µM). After 16 h of treatment, cells were pulse labeled with CldU (20 mM) for 10 min, washed twice, and incubated with IdU (200 mM) for additional 30 min (200 mM). DNA fibers were visualized using a Zeiss Axioplan confocal microscope. Images were analyzed using Zeiss LSM Image Browser software. Only bi-colored fibers were quantified to ensure that only active replication forks, but neither terminations nor recently fired origins, were analyzed.