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Split crRNA with CRISPR-Cas12a enabling highly sensitive and multiplexed detection of RNA and DNA

Author

Listed:
  • Yichuan Chen

    (Wuhan Polytechnic University
    Hubei University)

  • Xinping Wang

    (Hubei University)

  • Junqi Zhang

    (Hubei University)

  • Qingyuan Jiang

    (Hubei University)

  • Bin Qiao

    (Zhengzhou University)

  • Baoxia He

    (Affiliated Cancer Hospital of Zhengzhou University and Henan cancer hospital)

  • Wenhao Yin

    (BravoVax Co. Ltd.)

  • Jie Qiao

    (Wuhan Polytechnic University
    Zhengzhou University)

  • Yi Liu

    (Hubei University
    BravoVax Co. Ltd.)

Abstract

The CRISPR-Cas12a system has revolutionized nucleic acid testing (NAT) with its rapid and precise capabilities, yet it traditionally required RNA pre-amplification. Here we develop rapid fluorescence and lateral flow NAT assays utilizing a split Cas12a system (SCas12a), consisting of a Cas12a enzyme and a split crRNA. The SCas12a assay enables highly sensitive, amplification-free, and multiplexed detection of miRNAs and long RNAs without complex secondary structures. It can differentiate between mature miRNA and its precursor (pre-miRNA), a critical distinction for precise biomarker identification and cancer progression monitoring. The system’s specificity is further highlighted by its ability to detect DNA and miRNA point mutations. Notably, the SCas12a system can quantify the miR-21 biomarker in plasma from cervical cancer patients and, when combined with RPA, detect HPV at attomole levels in clinical samples. Together, our work presents a simple and cost-effective SCas12a-based NAT platform for various diagnostic settings.

Suggested Citation

  • Yichuan Chen & Xinping Wang & Junqi Zhang & Qingyuan Jiang & Bin Qiao & Baoxia He & Wenhao Yin & Jie Qiao & Yi Liu, 2024. "Split crRNA with CRISPR-Cas12a enabling highly sensitive and multiplexed detection of RNA and DNA," Nature Communications, Nature, vol. 15(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:15:y:2024:i:1:d:10.1038_s41467-024-52691-x
    DOI: 10.1038/s41467-024-52691-x
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    1. Santosh R. Rananaware & Emma K. Vesco & Grace M. Shoemaker & Swapnil S. Anekar & Luke Samuel W. Sandoval & Katelyn S. Meister & Nicolas C. Macaluso & Long T. Nguyen & Piyush K. Jain, 2023. "Programmable RNA detection with CRISPR-Cas12a," Nature Communications, Nature, vol. 14(1), pages 1-14, December.
    2. Zhichen Xu & Dongjuan Chen & Tao Li & Jiayu Yan & Jiang Zhu & Ting He & Rui Hu & Ying Li & Yunhuang Yang & Maili Liu, 2022. "Microfluidic space coding for multiplexed nucleic acid detection via CRISPR-Cas12a and recombinase polymerase amplification," Nature Communications, Nature, vol. 13(1), pages 1-14, December.
    3. Xiaoyi Fu & Guoliang Ke & Fangqi Peng & Xue Hu & Jiaqi Li & Yuyan Shi & Gezhi Kong & Xiao-Bing Zhang & Weihong Tan, 2020. "Size-selective molecular recognition based on a confined DNA molecular sieve using cavity-tunable framework nucleic acids," Nature Communications, Nature, vol. 11(1), pages 1-11, December.
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