"Challenges to the stable maintenance of the human genome can come from both endogenous and exogenoussources. However, one of the major threats to genome stability occurs during normal DNA metabolism. Thegenome is particularly susceptible to perturbation during the S-phase of the cell cycle when DNA replicationoccurs. This is because DNA replication forks can encounter chemical adducts, DNA secondary structures,topological constraints or bound proteins that hinder their progression. In actively proliferating cells, such asstem cells, replication perturbation can lead to fork stalling, breakage or collapse. These scenarios can, inturn, generate deleterious chromosomal rearrangements that have the potential to initiate human disease.Despite recent advances in our understanding of the biochemical process of DNA replication, the precisedetails of the events occurring at sites where replication forks have been perturbed remain poorlycharacterised. This is because in-depth analysis of the perturbation of replication represents a major technicalchallenge, principally because adducts and lesions generated by DNA damaging agents are randomlydistributed throughout the genome at sites that cannot be controlled or predicted. To overcome this technicallimitation, we have developed systems for site-specific perturbation of DNA replication that can betransferred to any locus in any cell type. The aim is to define how replication fork perturbation is detectedand engaged by cellular stress-response factors, and then tolerated or repaired. This highly integratedproposal, and the pioneering technologies that will be used to fulfil our ambitious aims, will have significantimplications for the understanding not only of replication perturbation and its effects on chromosomedynamics in mitosis, but also of the role of replication stress in the aetiology of cancer and premature ageing.It will open up new horizons both in and across these fields of research."