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Cell. 2013 Dec 19;155(7):1479-91. doi: 10.1016/j.cell.2013.12.001.

Dynamic imaging of genomic loci in living human cells by an optimized CRISPR/Cas system.

Author information

  • 1Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158, USA.
  • 2Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, Berkeley, CA 94720, USA.
  • 3Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA; Tetrad Graduate Program, University of California, San Francisco, San Francisco, CA 94158, USA.
  • 4Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; UCSF Center for Systems and Synthetic Biology, University of California, San Francisco, San Francisco, CA 94158, USA.
  • 5Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA.
  • 6Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, Berkeley, CA 94720, USA; California Institute for Quantitative Biomedical Research (QB3), San Francisco, CA 94158, USA.
  • 7Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; UCSF Center for Systems and Synthetic Biology, University of California, San Francisco, San Francisco, CA 94158, USA; California Institute for Quantitative Biomedical Research (QB3), San Francisco, CA 94158, USA. Electronic address: stanley.qi@ucsf.edu.
  • 8Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158, USA; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA; California Institute for Quantitative Biomedical Research (QB3), San Francisco, CA 94158, USA. Electronic address: bo.huang@ucsf.edu.

Erratum in

  • Cell. 2014 Jan 16;156(1-2):373.

Abstract

The spatiotemporal organization and dynamics of chromatin play critical roles in regulating genome function. However, visualizing specific, endogenous genomic loci remains challenging in living cells. Here, we demonstrate such an imaging technique by repurposing the bacterial CRISPR/Cas system. Using an EGFP-tagged endonuclease-deficient Cas9 protein and a structurally optimized small guide (sg) RNA, we show robust imaging of repetitive elements in telomeres and coding genes in living cells. Furthermore, an array of sgRNAs tiling along the target locus enables the visualization of nonrepetitive genomic sequences. Using this method, we have studied telomere dynamics during elongation or disruption, the subnuclear localization of the MUC4 loci, the cohesion of replicated MUC4 loci on sister chromatids, and their dynamic behaviors during mitosis. This CRISPR imaging tool has potential to significantly improve the capacity to study the conformation and dynamics of native chromosomes in living human cells.

Copyright © 2013 Elsevier Inc. All rights reserved.

PMID:
24360272
[PubMed - indexed for MEDLINE]
PMCID:
PMC3918502
Free PMC Article
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