title: Visualization and interpretation of eukaryotic DNA replication intermediates in vivo by electron microscopy.
abstract: The detailed understanding of the DNA replication process requires structural insight. The combination of psoralen cross-linking and electron microscopy has been extensively exploited to reveal the fine architecture of in vivo DNA replication intermediates. This approach proved instrumental to uncover the basic mechanisms of DNA duplication, as well as the perturbation of this process by various forms of replication stress. The replication structures are stabilized in vivo (by psoralen cross-linking) prior to extraction and enrichment procedures, allowing their visualization at the transmission electron microscope. This chapter outlines the procedures required to visualize and interpret in vivo replication intermediates of genomic DNA, extracted from budding yeast, Xenopus egg extracts, or cultured mammalian cells.
authors.affiliation: Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland.
authors.forename: Kai J
authors.initials: KJ
authors.lastname: Neelsen
authors.forename: Arnab Ray
authors.initials: AR
authors.lastname: Chaudhuri
authors.forename: Cindy
authors.initials: C
authors.lastname: Follonier
authors.forename: Raquel
authors.initials: R
authors.lastname: Herrador
authors.forename: Massimo
authors.initials: M
authors.lastname: Lopes
mesh_terms: D000818:Animals; D002457:Cell Extracts; D002843:Chromatin / Q000378:metabolism; D022202:Chromosomes, Artificial, Bacterial / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D004261:DNA Replication / Q000187:drug effects; D045566:DNA, Cruciform / Q000378:metabolism; D005057:Eukaryotic Cells / Q000166:cytology* / Q000648:ultrastructure*; D005363:Ficusin / Q000494:pharmacology; D016681:Genome, Fungal; D008297:Male; D008322:Mammals; D008854:Microscopy, Electron / Q000379:methods*; D009691:Nucleic Acid Denaturation / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D013094:Spermatozoa / Q000166:cytology / Q000187:drug effects; D014981:Xenopus
chemical_list: D002457:Cell Extracts; D002843:Chromatin; D003432:Cross-Linking Reagents; D045566:DNA, Cruciform; D004247:DNA; D005363:Ficusin