title: Analysis of Single-Locus Replication Timing in Asynchronous Cycling Cells.
abstract: In higher eucaryotes, not all the genome is replicated simultaneously: there are parts of the genome that replicate at the beginning of S-phase (early S-phase), others that are replicated later. In each cell, early replicating genomic regions are alternated to late-replicating regions. In general, eucaryotic genomes are organized into structural domains where genes showing the same epigenetic state replicate at the same time.Here, we will describe the protocol that we routinely used for the analysis of replication timing of specific loci in Drosophila embryonic cell lines (S2 and S3) based on BrdU labeling and FACS sorting of different S-phase fractions (early, mid, late) of asynchronous cycling cells.
authors.affiliation: Oncogenomic and Epigenetic Unit, Regina Elena Cancer Institute, via Elio Chianesi 53, 00144, Rome, Italy. losardo@ifo.it.
authors.forename: Lo Sardo
authors.initials: LS
authors.lastname: Federica
mesh_terms: D000818:Animals; D002453:Cell Cycle / Q000235:genetics*; D004261:DNA Replication / Q000235:genetics*; D004330:Drosophila / Q000196:embryology / Q000235:genetics; D053595:Embryonic Stem Cells; D016678:Genome; D008967:Molecular Biology / Q000379:methods*; D016196:S Phase / Q000235:genetics
keywords: BrdU; Early; FACS sorting; Late; Mid; Replication timing; S-phase