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G2/M DNA synthesis (G-MiDS) can be observed in one in five G2/M cells in unperturbed conditions by immunofluorescence microscopy. However, little is known of the genomic sites undergoing G-MiDS. Here, we describe a protocol which allows enriching for G2/M cells and investigating the sites of G-MiDS using BrdU-seq. This method can also be used to study the role of DNA replication or transcription-associated factors in affecting G-MiDS levels in different cell lines. For complete details on the use and execution of this protocol, please refer to Wang et al. (2021).
Bibliographical noteFunding Information:
We thank the Genomics Birmingham for help with NGS library preparation and sequencing samples and Paola Pietroni and Adriana Flores-Langarica from the Flow Cytometry Facility for sorting mitotic cells, all from the University of Birmingham. This work was supported by a University of Birmingham fellowship, the Wellcome Trust ( 202115/Z/16/Z ), the Royal Society ( RG170246 ), and the BBSRC ( BB/S016155/1 ) to M.S.
- Cell Biology
- Cell isolation
- Flow Cytometry/Mass Cytometry
- Molecular Biology
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Immunology and Microbiology(all)
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- 1 Finished
1/09/19 → 31/08/23
Project: Research Councils