Protocol to isolate cells in four stages of S phase for high-resolution replication-timing sequencing

Johnathan R. Whetstine, Capucine Van Rechem

Research output: Contribution to journalArticlepeer-review

Abstract

Traditional replication timing (RT) experiments divide S phase into two phases: early and late. However, there is an increasing awareness that variation in RT can occur during the course of S phase and impact our understanding of RT patterns and regulation. Here, we describe a RT protocol in RPE-1 cells for collecting four phases within S and the library preparation that takes advantage of a commercial kit for methyl-DNA. This step allows BrdU-labeled DNA sequencing and assessment of RT genome wide. For complete details on the use and execution of this protocol, please refer to Van Rechem et al. (2021).

Original languageEnglish
Article number101209
JournalSTAR Protocols
Volume3
Issue number1
DOIs
StatePublished - Mar 18 2022

Keywords

  • Cell isolation
  • Flow Cytometry/Mass Cytometry
  • Genomics
  • Molecular Biology
  • Sequencing

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