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Oligonucleotide microarrays were used to map the detailed topography of chromosome replication in the budding yeast Saccharomyces cerevisiae. The times of replication of thousands of sites across the genome were determined by hybridizing replicated and unreplicated DNAs, isolated at different times in S phase, to the microarrays. Origin activations take place continuously throughout S phase but with most firings near mid-S phase. Rates of replication fork movement vary greatly from region to region in the genome. The two ends of each of the 16 chromosomes are highly correlated in their times of replication. This microarray approach is readily applicable to other organisms, including humans.
The replication of eukaryotic chromosomes is highly regulated. Replication is limited to the S phase of the cell cycle; and within S phase, initiation of replication is controlled with respect to both location and time. The sites of initiation, called replication origins, have been best characterized in the budding yeast Saccharomyces cerevisiae, in which a functional assay based on plasmid maintenance has allowed identification of potential origins of replication [autonomous replication sequence elements (ARSs)]. There are estimated to be -200 to 400 ARSs in the yeast genome (1, 2), and most, but not all, function as chromosomal origins (3). The few origins investigated at the sequence level usually encompass -200 base pairs (bp); most contain a perfect match or a one-base mismatch to an 11-bp ARS consensus sequence (ACS) (4, 5). However, the presence of an ACS is not sufficient to predict an origin of replication: There are many more ARS consensus sequences in the genome than origins and the -200 bp of sequence that flank the ACS, although essential, share no obvious sequence similarities (3).
From the analysis of a few origins, the general features of origin control in yeast appear to be as follows (6-8). Most (perhaps all) origins are found in intergenic regions, but only a subset are chosen as replication origins in any given S phase. Furthermore, origins range in efficiency from those that are active in almost every cell cycle to those that are used in only a small proportion of cell cycles. Origins are activated in a reproducible temporal sequence through S phase. The time of origin activation appears not to be an intrinsic...