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Eukaryotic genomes are replicated under the control of a highly sophisticated program during the restricted time period corresponding to S phase. The most widely used replication timing assays, which are performed on populations of millions of cells, suggest that most of the genome is synchronously replicated on homologous chromosomes. We investigated the stochastic nature of this temporal program, by comparing the precise replication times of allelic loci within single vertebrate cells progressing through S phase at six loci replicated from very early to very late. We show that replication timing is strictly controlled for the three loci replicated in the first half of S phase. Out of the three loci replicated in the second part of S phase, two present a significantly more stochastic pattern. Surprisingly, we find that the locus replicated at the very end of S phase, presents stochasticity similar to those replicated in early S phase. We suggest that the richness of loci in efficient origins of replication, which decreases from early- to late-replicating regions, and the strength of interaction with the nuclear lamina may underlie the variation of timing control during S phase.

Replication dynamics of individual loci in single living cells reveal changes in the degree of replication stochasticity through S phase