Cockayne syndrome (CS) is a rare recessive genetic disease characterized in part by premature ageing and photosensitive skin. Because of the latter characteristic, this syndrome was considered to be an example of a UV-sensitive DNA repair-defective human disorder. We demonstrated normal levels of UV-induced unscheduled DNA synthesis (UDS) in four unrelated CS patients that show hypersensitivity to both UV and Mitomycin C (MMC). At low UV exposure, CS DNA shows a dose-dependent decrease in size. By contrast, heterozygotes appear to have a threshold below which there is little change in size of single strand DNA. Immediately following UV or MMC treatment, CS DNA is deficient in high molecular weight species, but undergoes a normal transition to larger DNA during a chase interval in the presence or absence of caffeine. This suggests a defect in replication or excision repair and no defect in post-replication repair (PRR). Pulse studies performed in the presence of hydroxyurea (HU) also reveal a deficient production of large DNA, suggesting the defect is in repair. As these cells have normal UDS and normal PRR, the basis for their UV sensitivity must be distinct from that observed in xeroderma pigmentosum (XP).