title: High-Throughput Live-Cell Microscopy Analysis of Association Between Chromosome Domains and the Nucleolus in S. cerevisiae.
abstract: Spatial organization of the genome has important impacts on all aspects of chromosome biology, including transcription, replication, and DNA repair. Frequent interactions of some chromosome domains with specific nuclear compartments, such as the nucleolus, are now well documented using genome-scale methods. However, direct measurement of distance and interaction frequency between loci requires microscopic observation of specific genomic domains and the nucleolus, followed by image analysis to allow quantification. The fluorescent repressor operator system (FROS) is an invaluable method to fluorescently tag DNA sequences and investigate chromosome position and dynamics in living cells. This chapter describes a combination of methods to define motion and region of confinement of a locus relative to the nucleolus in cell's nucleus, from fluorescence acquisition to automated image analysis using two dedicated pipelines.
authors: Laboratoire de Biologie Moléculaire Eucaryote, Centre de Biologie Intégrative (CBI), Université de Toulouse, CNRS, UPS, 31000, Toulouse, France.
authors: Renjie
authors: R
authors: Wang
authors: Laboratoire de Biologie Moléculaire Eucaryote, Centre de Biologie Intégrative (CBI), Université de Toulouse, CNRS, UPS, 31000, Toulouse, France.
authors: Christophe
authors: C
authors: Normand
authors: Laboratoire de Biologie Moléculaire Eucaryote, Centre de Biologie Intégrative (CBI), Université de Toulouse, CNRS, UPS, 31000, Toulouse, France. gadal@ibcg.biotoul.fr.
authors: Olivier
authors: O
authors: Gadal
mesh_terms: D002466:Cell Nucleolus / Q000235:genetics*; D015825:Chromosomes, Fungal; D017930:Genes, Reporter; D056426:Genetic Loci; D057166:High-Throughput Screening Assays; D007091:Image Processing, Computer-Assisted; D008856:Microscopy, Fluorescence / Q000379:methods; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D059629:Signal-To-Noise Ratio; D059008:Time-Lapse Imaging
keywords: Chromosome domain dynamics; FROS; Fluorescence microscopy; Live-cell imaging; Nuclear organization; Nucleolus; Saccharomyces cerevisiae; Yeast