that EBNA1 could also promote genome instability by forming RFBs at FR-like sequences in the human genome (61,62). A ﬁnal cautionary note Arrays of lac or tet operator sequences bound by their cognate repressor fused to a ﬂuorescent protein are widely used as a molecular tool to tag genomic loci for live cell imaging studies. Our data show that in wild-type cells such arrays can be tolerated well as long as the level of repres- sor protein is low. However, in mutants where the machin- ery of replication fork progression/protection/restart is perturbed, even low levels of repressor protein may be sufﬁcient to induce instability at the array. This should be borne in mind when using operator sequences for live cell imaging experiments, especially when processes involved in genome stability are under investigation. SUPPLEMENTARY DATA Supplementary Data are available at NAR Online. ACKNOWLEDGEMENTS We thank Ashok Aiyar (Louisiana State University), David Sherratt (University of Oxford) and Virginia Zakian (Princeton University) for strains and plasmids. FUNDING Grants 057586/Z/99/B and 090767/Z/09/Z from the Wellcome Trust. S.S. was supported by a Skaggs-Oxford scholarship. S.C. was supported by a Dorothy Hodgkin Fellowship. J.L. was supported by grant MHIRT 2T37MD001368 from the National Center on Minority Health and Health Disparities, National Institutes of Health. Funding for open access charge: Wellcome Trust. Conﬂict of interest statement. None declared. REFERENCES 1. McGlynn,P. and Lloyd,R.G. (2002) Recombinational repair and restart of damaged replication forks. Nat. Rev. Mol. Cell. Biol., 3, 859–870. 2. Rothstein,R., Michel,B. and Gangloff,S. (2000) Replication fork pausing and recombination or ‘‘gimme a break’’. Genes Dev., 14, 1–10. 3. Branzei,D. and Foiani,M. (2007) Interplay of replication checkpoints and repair proteins at stalled replication forks. DNA Repair, 6, 994–1003. 4. Branzei,D. and Foiani,M. (2010) Maintaining genome stability at the replication fork. Nat. Rev. Mol. Cell. Biol., 11, 208–219. 5. Lambert,S., Mizuno,K., Blaisonneau,J., Martineau,S., Chanet,R., Freon,K., Murray,J.M., Carr,A.M. and Baldacci,G. (2010) Homologous recombination restarts blocked replication forks at the expense of genome rearrangements by template exchange. Mol. Cell, 39, 346–359. 6. Lambert,S., Watson,A., Sheedy,D.M., Martin,B. and Carr,A.M. (2005) Gross chromosomal rearrangements and elevated recombination at an inducible site-speciﬁc replication fork barrier. Cell, 121, 689–702. Nucleic Acids Research, 2011, Vol. 39, No. 15 6583 7. Heyer,W.D., Ehmsen,K.T. and Liu,J. (2010) Regulation of homologous recombination in eukaryotes. Annu. Rev. Genet., 44, 113–139. 8. Chen,J.M., Cooper,D.N., Ferec,C., Kehrer-Sawatzki,H. and Patrinos,G.P. (2010) Genomic rearrangements in inherited disease and cancer. Semin. Cancer Biol., 20, 222–233. 9. San Filippo,J., Sung,P. and Klein,H. (2008) Mechanism of eukaryotic homologous recombination. Annu. Rev. Biochem., 77, 229–257. 10. Atkinson,J. and McGlynn,P. (2009) Replication fork reversal and the maintenance of genome stability. Nucleic Acids Res., 37, 3475–3492. 11. Osman,F. and Whitby,M.C. (2007) Exploring the roles of Mus81-Eme1/Mms4 at perturbed replication forks. DNA Repair, 6, 1004–1017. 12. Cortes-Ledesma,F., Tous,C. and Aguilera,A. (2007) Different genetic requirements for repair of replication-born double-strand breaks by sister-chromatid recombination and break-induced replication. Nucleic Acids Res., 35, 6560–6570. 13. Aguilera,A. and Gomez-Gonzalez,B. (2008) Genome instability: a mechanistic view of its causes and consequences. Nat. Rev. Genet., 9, 204–217. 14. Ahn,J.S., Osman,F. and Whitby,M.C. (2005) Replication fork blockage by RTS1 at an ectopic site promotes recombination in ﬁssion yeast. EMBO J., 24, 2011–2023. 15. Pryce,D.W., Ramayah,S., Jaendling,A. and McFarlane,R.J. (2009) Recombination at DNA replication fork barriers is not universal and is differentially regulated by Swi1. Proc. Natl Acad. Sci. USA, 106, 4770–4775. 16. Codlin,S. and Dalgaard,J.Z. (2003) Complex mechanism of site-speciﬁc DNA replication termination in ﬁssion yeast. EMBO J., 22, 3431–3440. 17. Inagawa,T., Yamada-Inagawa,T., Eydmann,T., Mian,I.S., Wang,T.S. and Dalgaard,J.Z. (2009) Schizosaccharomyces pombe Rtf2 mediates site-speciﬁc replication termination by inhibiting replication restart. Proc. Natl Acad. Sci. USA, 106, 7927–7932. 18. Chan,K.L., North,P.S. and Hickson,I.D. (2007) BLM is required for faithful chromosome segregation and its localization deﬁnes a class of ultraﬁne anaphase bridges. EMBO J., 26, 3397–3409. 19. Chan,K.L., Palmai-Pallag,T., Ying,S. and Hickson,I.D. (2009) Replication stress induces sister-chromatid bridging at fragile site loci in mitosis. Nat. Cell Biol., 11, 753–760. 20. Moreno,S., Klar,A. and Nurse,P. (1991) Molecular genetic analysis of ﬁssion yeast Schizosaccharomyces pombe. Methods Enzymol., 194, 795–823. 21. Osman,F. and Whitby,M.C. (2009) Monitoring homologous recombination following replication fork perturbation in the ﬁssion yeast Schizosaccharomyces pombe. Methods Mol. Biol., 521, 535–552. 22. Osman,F., Adriance,M. and McCready,S. (2000) The genetic control of spontaneous and UV-induced mitotic intrachromosomal recombination in the ﬁssion yeast Schizosaccharomyces pombe. Curr. Genet., 38, 113–125. 23. Lau,I.F., Filipe,S.R., Soballe,B., Okstad,O.A., Barre,F.X. and Sherratt,D.J. (2003) Spatial and temporal organization of replicating Escherichia coli chromosomes. Mol. Microbiol., 49, 731–743. 24. Osman,F., Fortunato,E.A. and Subramani,S. (1996) Double-strand break-induced mitotic intrachromosomal recombination in the ﬁssion yeast Schizosaccharomyces pombe. Genetics, 142, 341–357. 25. Aiyar,A., Aras,S., Washington,A., Singh,G. and Luftig,R.B. (2009) Epstein-Barr Nuclear Antigen 1 modulates replication of oriP-plasmids by impeding replication and transcription fork migration through the family of repeats. Virol. J., 6, 29. 26. Maundrell,K. (1993) Thiamine-repressible expression vectors pREP and pRIP for ﬁssion yeast. Gene, 123, 127–130. 27. Huberman,J.A., Spotila,L.D., Nawotka,K.A., el-Assouli,S.M. and Davis,L.R. (1987) The in vivo replication origin of the yeast 2 microns plasmid. Cell, 51, 473–481. 28. Win,T.Z., Goodwin,A., Hickson,I.D., Norbury,C.J. and Wang,S.W. (2004) Requirement for Schizosaccharomyces pombe Top3 in the maintenance of chromosome integrity. J. Cell Sci., 117, 4769–4778.