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Science. 2019 Sep 20;365(6459):1301-1305. doi: 10.1126/science.aax7852. Epub 2019 Sep 5.

CRISPR-mediated live imaging of genome editing and transcription.

Author information

1
Department of Bioengineering, Stanford University, Stanford, CA 94305, USA.
2
ZJU-UOE Institute, Zhejiang University, School of Medicine, Haining, China.
3
Castilleja School, Palo Alto, CA 94301, USA.
4
Department of Statistics, Stanford University, Stanford, CA 94305, USA.
5
Department of Bioengineering, Stanford University, Stanford, CA 94305, USA. stanley.qi@stanford.edu.
6
Department of Chemical and Systems Biology, Stanford University, Stanford, CA 94305, USA.
7
ChEM-H Institute, Stanford University, Stanford, CA 94305, USA.

Abstract

We report a robust, versatile approach called CRISPR live-cell fluorescent in situ hybridization (LiveFISH) using fluorescent oligonucleotides for genome tracking in a broad range of cell types, including primary cells. An intrinsic stability switch of CRISPR guide RNAs enables LiveFISH to accurately detect chromosomal disorders such as Patau syndrome in prenatal amniotic fluid cells and track multiple loci in human T lymphocytes. In addition, LiveFISH tracks the real-time movement of DNA double-strand breaks induced by CRISPR-Cas9-mediated editing and consequent chromosome translocations. Finally, by combining Cas9 and Cas13 systems, LiveFISH allows for simultaneous visualization of genomic DNA and RNA transcripts in living cells. The LiveFISH approach enables real-time live imaging of DNA and RNA during genome editing, transcription, and rearrangements in single cells.

PMID:
31488703
DOI:
10.1126/science.aax7852

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