AbstractIn mammalian cells, distinct replication domains (RDs), corresponding to structural units of chromosomes called topologically-associating domains (TADs), replicate at different times during S-phase1–4. Further, early/late replication of RDs corresponds to active/inactive chromatin interaction compartments5,6. Although replication origins are selected stochastically, such that each cell is using a different cohort of origins to replicate their genomes7–12, replication-timing is regulated independently and upstream of origin selection13 and evidence suggests that replication timing is conserved in consecutive cell cycles14. Hence, quantifying the extent of cell-to-cell variation in replication timing is central to studies of chromosome structure and function. Here we devise a strategy to measure variation in single-cell replication timing using DNA copy number. We find that borders between replicated and un-replicated DNA are highly conserved between cells, demarcating active and inactive compartments of the nucleus. Nonetheless, measurable variation was evident. Surprisingly, we detected a similar degree of variation in replication timing from cell-to-cell, between homologues within cells, and between all domains genome-wide regardless of their replication timing. These results demonstrate that stochastic variation in replication timing is independent of elements that dictate timing or extrinsic environmental variation.
The Protective Role of Dormant Origins in Response to Replicative Stress