Fractional S-phase sorting using bromodeoxyuridine uptake to study gene replication timing

The determination of the percentage of cells in S-phase is dependent upon the detection of a thymidine analogue, bromodeoxyuridine (BrdU), which when added to culture medium is incorporated into DNA during DNA replication is a well known technique in flow cytometry.

Gene replication timing studies uses the BrdU labelling technique, S phase cells are sorted into 3 or more fractions. Collected fractions are analysed by PCR for genetic analysis after immunoprecipitation of DNA via incorporated BrdU.

• S phase sorted fractions [new window]

Publications.

P Takousis, P Johonnett, J Williamson, P Sasieni, G Warnes, T Forshew, V Azuara, A Fisher, PJ Wu, PJ Jones, R Vatcheva, S Beck, D Sheer. Replication timing profile reflects the distinct functional and genomic features of the MHC Class II Region. Cell Cycle Oct 6:19, 2393-2398, 2007.

V Azuara, P Perry, S Sauer, M Spivakov, HF Jorgensen, RM John, M Gouti, M Casanova, G Warnes, M Merkenschlager, AG Fisher. Chromatin signatures of pluripotent cell lines. Nature Cell Biol. 8, 532-538, 2006.

P Perry, S Saur, N Billion, WD Richardson, M Spivakov, G Warnes, FJ Livesey, M Merkenschlager, AG Fisher, V Auara. A dynamic switch in the replication timing of key regulator genes in embryonic stem cells upon neural induction. Cell Cycle 3, (12), 1645-50, 2004.