not sufficient to explore previously de- scribed replication initiation zones [8,9]. The development of new methods based on deep sequencing associated with pow- erful statistical analysis should allow not only origin mapping along entire genomes, but also the search for broad zones of initiation. Such regions should have, along the zone of initiation, a signal of SNS enrichment notably higher than in do- mains devoid of origins. Finally, Sequeira-Mendes et al. found that ORIs located within promoters are References 1. Aladjem MI (2007) Replication in context: dynamic regulation of DNA replication patterns in metazoans. Nat Rev Genet 8: 588–600. 2. Farkash-Amar S, Lipson D, Polten A, Goren A, Helmstetter C, et al. (2008) Global organization of replication time zones of the mouse genome. Genome Res 18: 1562–1570. 3. MacAlpine DM, Bell SP (2005) A genomic view of eukaryotic DNA replication. Chromosome Res 13: 309–326. 4. Hiratani I, Ryba T, Itoh M, Yokochi T, Schwaiger M, et al. (2008) Global reorganization of replication domains during embryonic stem cell differentiation. PLoS Biol 6: e245. doi:10.1371/journal.pbio.0060245. 5. Cadoret JC, Meisch F, Hassan-Zadeh V, Luyten I, Guillet C, et al. (2008) Genome-wide studies highlight indirect links between human replica- significantly enriched in genes that are expressed during early development. They proposed that this marks the competence to drive replication during the rest of development. Whole-genome analyses in different cell lines and across several species will allow testing of this hypothesis with a larger dataset. This will also provide information on the dynamics of evolution of replication origins in vertebrates and enable researchers to investigate the con- straints imposed by replication on genome organization, a topic that is a matter of tion origins and gene regulation. Proc Natl Acad Sci U S A 105: 15837–15842. 6. Sequeira-Mendes J, D´ıaz-Uriarte R, Apedaile A, Huntley D, Brockdorff N, et al. (2009) Transcrip- tion initiation activity sets replication origin efficiency in mammalian cells. PLoS Genet 5(4): e1000446. doi:10.1371/journal.pgen.1000446. 7. Necsulea A, Guillet C, Cadoret JC, Prioleau MN, Duret L (2009) The relationship between DNA replication and human genome organization. Mol Biol Evol; E-pub ahead of print. doi:10.1093/ molbev/msn303. 8. Vaughn JP, Dijkwel PA, Hamlin JL (1990) Replication initiates in a broad zone in the amplified CHO dihydrofolate reductase domain. Cell 61: 1075–1087. 9. Mesner LD, Crawford EL, Hamlin JL (2006) Isolating apparently pure libraries of replication debate in the community [7]. Finally, analyses of origin firing at specific loci by in vivo labelling and single-molecule analysis by DNA combing should help to complement and confirm whole-genome approaches [10–12]. The future develop- ment of these methods will ultimately give a vision that hews closely to the true complexity of origin firing in mammalian genomes. origins from complex genomes. Mol Cell 21: 719–726. 10. Anglana M, Apiou F, Bensimon A, Debatisse M (2003) Dynamics of DNA replication in mamma- lian somatic cells: nucleotide pool modulates origin choice and interorigin spacing. Cell 114: 385–394. 11. Lebofsky R, Heilig R, Sonnleitner M, Weissenbach J, Bensimon A (2006) DNA repli- cation origin interference increases the spacing between initiation events in human cells. Mol Biol Cell 17: 5337–5345. 12. Norio P, Kosiyatrakul S, Yang Q, Guan Z, Brown NM, et al. (2005) Progressive activation of DNA replication initiation in large domains of the immunoglobulin heavy chain locus during B cell development. Mol Cell 20: 575–587. PLoS Genetics | www.plosgenetics.org 2 April 2009 | Volume 5 | Issue 4 | e1000454