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Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9

Author

Listed:
  • Peiwu Qin

    (University of California at Berkeley)

  • Mahmut Parlak

    (School of Medicine, University of Virginia)

  • Cem Kuscu

    (School of Medicine, University of Virginia)

  • Jigar Bandaria

    (University of California at Berkeley)

  • Mustafa Mir

    (University of California at Berkeley)

  • Karol Szlachta

    (School of Medicine, University of Virginia)

  • Ritambhara Singh

    (School of Medicine, University of Virginia
    University of Virginia)

  • Xavier Darzacq

    (University of California at Berkeley)

  • Ahmet Yildiz

    (University of California at Berkeley
    University of California at Berkeley)

  • Mazhar Adli

    (School of Medicine, University of Virginia)

Abstract

Imaging chromatin dynamics is crucial to understand genome organization and its role in transcriptional regulation. Recently, the RNA-guidable feature of CRISPR-Cas9 has been utilized for imaging of chromatin within live cells. However, these methods are mostly applicable to highly repetitive regions, whereas imaging regions with low or no repeats remains as a challenge. To address this challenge, we design single-guide RNAs (sgRNAs) integrated with up to 16 MS2 binding motifs to enable robust fluorescent signal amplification. These engineered sgRNAs enable multicolour labelling of low-repeat-containing regions using a single sgRNA and of non-repetitive regions with as few as four unique sgRNAs. We achieve tracking of native chromatin loci throughout the cell cycle and determine differential positioning of transcriptionally active and inactive regions in the nucleus. These results demonstrate the feasibility of our approach to monitor the position and dynamics of both repetitive and non-repetitive genomic regions in live cells.

Suggested Citation

  • Peiwu Qin & Mahmut Parlak & Cem Kuscu & Jigar Bandaria & Mustafa Mir & Karol Szlachta & Ritambhara Singh & Xavier Darzacq & Ahmet Yildiz & Mazhar Adli, 2017. "Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9," Nature Communications, Nature, vol. 8(1), pages 1-10, April.
    • Handle: RePEc:nat:natcom:v:8:y:2017:i:1:d:10.1038_ncomms14725
    DOI: 10.1038/ncomms14725
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    Cited by:

    1. Zhongxuan Zhang & Xiaoxiao Rong & Tianjin Xie & Zehao Li & Haozhi Song & Shujun Zhen & Haifeng Wang & Jiahui Wu & Samie R. Jaffrey & Xing Li, 2024. "Fluorogenic CRISPR for genomic DNA imaging," Nature Communications, Nature, vol. 15(1), pages 1-12, December.
    2. Qin Peng & Ziliang Huang & Kun Sun & Yahan Liu & Chi Woo Yoon & Reed E. S. Harrison & Danielle L. Schmitt & Linshan Zhu & Yiqian Wu & Ipek Tasan & Huimin Zhao & Jin Zhang & Sheng Zhong & Shu Chien & Y, 2022. "Engineering inducible biomolecular assemblies for genome imaging and manipulation in living cells," Nature Communications, Nature, vol. 13(1), pages 1-15, December.
    3. Lauren A. Blake & Leslie Watkins & Yang Liu & Takanari Inoue & Bin Wu, 2024. "A rapid inducible RNA decay system reveals fast mRNA decay in P-bodies," Nature Communications, Nature, vol. 15(1), pages 1-14, December.
    4. Yanan Li & Yonghua Wu & Ru Xu & Jialing Guo & Fenglei Quan & Yongyuan Zhang & Di Huang & Yiran Pei & Hua Gao & Wei Liu & Junjie Liu & Zhenzhong Zhang & Ruijie Deng & Jinjin Shi & Kaixiang Zhang, 2023. "In vivo imaging of mitochondrial DNA mutations using an integrated nano Cas12a sensor," Nature Communications, Nature, vol. 14(1), pages 1-16, December.
    5. Patricia A. Clow & Menghan Du & Nathaniel Jillette & Aziz Taghbalout & Jacqueline J. Zhu & Albert W. Cheng, 2022. "CRISPR-mediated multiplexed live cell imaging of nonrepetitive genomic loci with one guide RNA per locus," Nature Communications, Nature, vol. 13(1), pages 1-10, December.

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