Chromosome replication timing is biphasic (early-late) in the cell cycle of vertebrates and of most (possibly all) eukaryotes. In the present work we have compared the extended, detailed replication timing maps that are available, namely those of human chromosomes 6, 11q, and 21q, with chromosomal bands as visualized at low (400 bands), high (850 bands), and highest (3,200 isochores) resolution. We have observed that the replicons located in a given isochore practically always show either all early or all late replication timing and that early-replicating isochores are short and GC-rich and late-replicating isochores are long and GC-poor. In the vast majority of cases, replicons are clustered in isochores, which are themselves most often clustered in early-or late-replication timing zones and may often reach the size of high-resolution bands and, very rarely, even that of low-resolution bands. Finally, we show that our results should be representative for the whole human genome and thus help to predict replication timing zones in all chromosomes.replication origins ͉ replicon clusters ͉ replicons F orty years ago Huberman and Riggs (1) showed, by combining the techniques of pulse-labeling and DNA autoradiography, that mammalian chromosomes are replicated from many replication origins and that adjacent starting points initiate replication at the same time. In other words, replicons, the DNA sequences that start their replication from a given origin, are organized in clusters that fire simultaneously. By using the incorporation of BrdU into human DNA, and thus changing the appearance of metaphase chromatids, it was shown, at the very low resolution of 277 bands, that reverse (R) bands replicate early, whereas quinacrine (Q) or Giemsa (G) bands replicate late (2-4), a result that was widely accepted for many years. Additional investigations indicated the existence of nine replication timings within each one of these two groups (5, 6).The heterogeneity in replication timing within the early and late classes was shown to be associated with different band structures. Indeed, at the low resolution of 400 bands, the R bands containing the GC-richest, gene-richest isochores from the H3 family (the H3 ϩ bands; for a general review on isochores see ref. 7) replicated at the onset of the S phase, whereas the bands not containing H3 isochores (H3 Ϫ bands) replicated later (8). Likewise, among low-resolution G bands, those predominantly composed by the GC-poorest, gene-poorest L1 isochores (L1 ϩ bands), which corresponded to the darkest G bands of Francke (9), replicated at the latest times, whereas the L1 Ϫ bands were the earliest-replicating among the G bands (10). These experiments indicated the existence of a correlation between the replication timings and the fine structure of chromosomal bands, namely the isochore composition of bands.A further step was made by assessing replication timing along chromosomal DNA sequences as done by Watanabe et al. (11) for human chromosomes 11q and 21q. These authors found that replic...