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CRISPR-induced DNA reorganization for multiplexed nucleic acid detection

Author

Listed:
  • Margot Karlikow

    (University of Toronto)

  • Evan Amalfitano

    (University of Toronto)

  • Xiaolong Yang

    (University of Toronto)

  • Jennifer Doucet

    (University of Toronto)

  • Abigail Chapman

    (University of British Columbia)

  • Peivand Sadat Mousavi

    (University of Toronto)

  • Paige Homme

    (University of Toronto)

  • Polina Sutyrina

    (University of Toronto)

  • Winston Chan

    (University of Toronto)

  • Sofia Lemak

    (University of Toronto)

  • Alexander F. Yakunin

    (University of Toronto
    Bangor University)

  • Adam G. Dolezal

    (University of Illinois at Urbana–Champaign)

  • Shana Kelley

    (University of Toronto
    University of Toronto
    University of Toronto
    Northwestern University)

  • Leonard J. Foster

    (University of British Columbia)

  • Brock A. Harpur

    (Purdue University)

  • Keith Pardee

    (University of Toronto
    University of Toronto)

Abstract

Nucleic acid sensing powered by the sequence recognition of CRIPSR technologies has enabled major advancement toward rapid, accurate and deployable diagnostics. While exciting, there are still many challenges facing their practical implementation, such as the widespread need for a PAM sequence in the targeted nucleic acid, labile RNA inputs, and limited multiplexing. Here we report FACT (Functionalized Amplification CRISPR Tracing), a CRISPR-based nucleic acid barcoding technology compatible with Cas12a and Cas13a, enabling diagnostic outputs based on cis- and trans-cleavage from any sequence. Furthermore, we link the activation of CRISPR-Cas12a to the expression of proteins through a Reprogrammable PAIRing system (RePAIR). We then combine FACT and RePAIR to create FACTOR (FACT on RePAIR), a CRISPR-based diagnostic, that we use to detect infectious disease in an agricultural use case: honey bee viral infection. With high specificity and accuracy, we demonstrate the potential of FACTOR to be applied to the sensing of any nucleic acid of interest.

Suggested Citation

  • Margot Karlikow & Evan Amalfitano & Xiaolong Yang & Jennifer Doucet & Abigail Chapman & Peivand Sadat Mousavi & Paige Homme & Polina Sutyrina & Winston Chan & Sofia Lemak & Alexander F. Yakunin & Adam, 2023. "CRISPR-induced DNA reorganization for multiplexed nucleic acid detection," Nature Communications, Nature, vol. 14(1), pages 1-11, December.
    • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-36874-6
    DOI: 10.1038/s41467-023-36874-6
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    References listed on IDEAS

    as
    1. Evan Amalfitano & Margot Karlikow & Masoud Norouzi & Katariina Jaenes & Seray Cicek & Fahim Masum & Peivand Sadat Mousavi & Yuxiu Guo & Laura Tang & Andrew Sydor & Duo Ma & Joel D. Pearson & Daniel Tr, 2021. "A glucose meter interface for point-of-care gene circuit-based diagnostics," Nature Communications, Nature, vol. 12(1), pages 1-10, December.
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    5. Daphne Collias & Chase L. Beisel, 2021. "CRISPR technologies and the search for the PAM-free nuclease," Nature Communications, Nature, vol. 12(1), pages 1-12, December.
    6. Benjamin P. Kleinstiver & Michelle S. Prew & Shengdar Q. Tsai & Ved V. Topkar & Nhu T. Nguyen & Zongli Zheng & Andrew P. W. Gonzales & Zhuyun Li & Randall T. Peterson & Jing-Ruey Joanna Yeh & Martin J, 2015. "Engineered CRISPR-Cas9 nucleases with altered PAM specificities," Nature, Nature, vol. 523(7561), pages 481-485, July.
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    8. Gallai, Nicola & Salles, Jean-Michel & Settele, Josef & Vaissière, Bernard E., 2009. "Economic valuation of the vulnerability of world agriculture confronted with pollinator decline," Ecological Economics, Elsevier, vol. 68(3), pages 810-821, January.
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    Cited by:

    1. Tao Hu & Xinxin Ke & Yingying Yu & Hongmei Feng & Senfeng Zhang & Yinuo Cui & Boyang Zhang & Min He & Yinbing Tang & Lei Liu & Yu Lin & Quanquan Ji & Chuanxia Chen & Chunlong Xu & Chunyi Hu, 2025. "NAPTUNE: nucleic acids and protein biomarkers testing via ultra-sensitive nucleases escalation," Nature Communications, Nature, vol. 16(1), pages 1-16, December.
    2. Jeong Moon & Changchun Liu, 2023. "Asymmetric CRISPR enabling cascade signal amplification for nucleic acid detection by competitive crRNA," Nature Communications, Nature, vol. 14(1), pages 1-11, December.

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