High-resolution mapping of mitotic DNA synthesis under conditions of replication stress in cultured cells

Abstract

Summary: Cells experiencing DNA replication stress enter mitosis with under-replicated DNA, which activates a repair mechanism known as mitotic DNA synthesis (MiDAS). Here we describe a protocol to identify at genome wide and at high resolution the genomic sites where MiDAS occurs in cells exposed to aphidicolin. We use EdU incorporation to label nascent DNA in mitotic cells, followed by isolation of the EdU-labeled DNA and next-generation sequencing.For complete details on the use and execution of this protocol, please refer to Groelly et al. (2022)1 and Macheret et al. (2020).2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell Biology
• Cell culture
• Sequencing
• ChIPseq
• Molecular Biology