GEOapplication/xmlftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE320nnn/GSE320256/primaryOK200OtherHomo sapiensOtherhttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE320256GEOGSEfalseSingle-molecule DNA replication dynamics under WEE1 inhibition in wild-type and TRESLIN-stabilized HCT116 cellsWe profiled single-molecule DNA replication dynamics in human HCT116 cells to determine how WEE1 inhibition and stabilization of the origin-firing factor TRESLIN (TICRR) influence replication initiation, termination, and fork progression. We analyzed two engineered HCT116 lines expressing mClover-tagged TRESLIN from the endogenous locus: a wild-type TRESLIN allele and a stabilized ΔSBI TRESLIN mutant. Cells were treated with the WEE1 inhibitor MK-1775 (1 µM) or vehicle and sequentially pulse-labeled with EdU and BrdU, followed by a thymidine chase, to generate replication tracts for long-read nanopore sequencing. High–molecular weight genomic DNA was prepared and sequenced on an Oxford Nanopore PromethION platform, and reads were aligned to the human reference genome (hg38/GRCh38). Replication features (leftward/rightward forks, initiation events, and termination events) and tract lengths were inferred from EdU/BrdU segmentation using DNAscent forkSense, enabling quantification of fork directionality and fork rates across conditions. Associated preprint: bioRxiv [Preprint]. 2025 Jun 11;2025.06.10.657920. doi:10.1101/2025.06.10.657920.2026/04/17GSE320256GSM9538414GSM9538413GSM9538405GSM9538416GSM9538415GSM9538410GSM9538412GSM9538411GSM9538407GSM9538406GSM9538409GSM953840826167320256Homo sapiens10.1101/2025.06.10.657920