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A double thymidine block is a synchronization strategy in which two thymidine-induced interruptions of DNA synthesis, separated by release, enrich a cultured-cell biological population near the transition into S phase. Excess thymidine perturbs nucleotide balance rather than acting as a physical barrier. Cells resume progression after release with a distribution of timings, so synchronization is enrichment rather than perfect identity of cell states. The perturbation can itself affect replication and stress responses; measurements after release must account for this changed starting condition. ^[ref]