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Tunable control of Cas12 activity promotes universal and fast one-pot nucleic acid detection

Author

Listed:
  • Zhou-Hua Cheng

    (University of Science and Technology of China)

  • Xi-Yan Luo

    (University of Science and Technology of China)

  • Sheng-Song Yu

    (University of Science and Technology of China)

  • Di Min

    (University of Science and Technology of China)

  • Shu-Xia Zhang

    (Fujian Medical University Union Hospital)

  • Xiao-Fan Li

    (Fujian Medical University Union Hospital)

  • Jie-Jie Chen

    (University of Science and Technology of China)

  • Dong-Feng Liu

    (University of Science and Technology of China)

  • Han-Qing Yu

    (University of Science and Technology of China)

Abstract

The CRISPR-based detection methods have been widely applied, yet they remain limited by the non-universal nature of one-pot diagnostic approaches. Here, we report a universal one-pot fluorescent method for the detection of epidemic pathogens, delivering results within 15-20 min. This method uses heparin sodium to precisely tunes the cis-cleavage capability of Cas12 via interference with the Cas12a-crRNA binding process, thereby generating significant fluorescence due to the accumulation of isothermal amplification products. Additionally, this universal assay accommodates both classic and suboptimal PAMs, as well as various Cas12a subtypes such as LbCas12a, AsCas12a, and AapCas12b. Such a robust method demonstrates sensitivity and specificity exceeding 95% in the detection of monkeypox pseudovirus, influenza A virus, and SARS-CoV-2 from saliva or wastewater samples, when compared with qPCR or RT-qPCR. Moreover, the cost of heparin sodium per thousand uses is $0.01 to $0.04 only. Collectively, this universal and fast one-pot approach based on heparin sodium offers potential possibilities for point-of-care testing.

Suggested Citation

  • Zhou-Hua Cheng & Xi-Yan Luo & Sheng-Song Yu & Di Min & Shu-Xia Zhang & Xiao-Fan Li & Jie-Jie Chen & Dong-Feng Liu & Han-Qing Yu, 2025. "Tunable control of Cas12 activity promotes universal and fast one-pot nucleic acid detection," Nature Communications, Nature, vol. 16(1), pages 1-16, December.
    • Handle: RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-56516-3
    DOI: 10.1038/s41467-025-56516-3
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    References listed on IDEAS

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    1. Xiong Ding & Kun Yin & Ziyue Li & Rajesh V. Lalla & Enrique Ballesteros & Maroun M. Sfeir & Changchun Liu, 2020. "Ultrasensitive and visual detection of SARS-CoV-2 using all-in-one dual CRISPR-Cas12a assay," Nature Communications, Nature, vol. 11(1), pages 1-10, December.
    2. Xuan Li & Huan Liu & Li Gao & Samendra P. Sherchan & Ting Zhou & Stuart J. Khan & Mark C. M. Loosdrecht & Qilin Wang, 2023. "Wastewater-based epidemiology predicts COVID-19-induced weekly new hospital admissions in over 150 USA counties," Nature Communications, Nature, vol. 14(1), pages 1-14, December.
    3. Daphne Collias & Elena Vialetto & Jiaqi Yu & Khoa Co & Éva d. H. Almási & Ann-Sophie Rüttiger & Tatjana Achmedov & Till Strowig & Chase L. Beisel, 2023. "Systematically attenuating DNA targeting enables CRISPR-driven editing in bacteria," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
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