Repli-Seq

Repli-Seq

Repli-Seq maps the sequences of nascent DNA replication strands throughout the whole genome during each of the six cell cycle phases. This is achieved by growing cells in bromouridine triphosphate (BrdU) media to replace thymidine. The cells are then sorted to their current state in cell division using Fluorescent-activated cell sorting (FACS). BrdU-labelled DNA strands are immunoprecipitated by anti-BrdU antibodies on magnetic beads. These immunoprecipitated strands can be prepared for sequencing following TruSeq DNA library preparation protocol.

Pros:

• Maps sequences of newly replicated DNA to the phases of cell division
• Low sample input required (5000 cells) makes it suitable for studying rare cell populations
• Streamlined DNA library prep step

Cons:

• The BrdU labeling requirement limits this approach to cultured cells

• Repli-Seq: Hansen R. S., Thomas S., Sandstrom R., Canfield T. K., Thurman R. E., et al. (2010) Sequencing newly replicated DNA reveals widespread plasticity in human replication timing. Proc Natl Acad Sci U S A 107: 139-144

1. Deyle D. R., Hansen R. S., Cornea A. M., Li L. B., Burt A. A., et al. (2014) A genome-wide map of adeno-associated virus-mediated human gene targeting. Nat Struct Mol Biol 21: 969-975
2. Barlow J. H., Faryabi R. B., Callen E., Wong N., Malhowski A., et al. (2013) Identification of early replicating fragile sites that contribute to genome instability. Cell 152: 620-632