A minimal transcription template-based amplification-free CRISPR-Cas13a strategy for DNA detection

CRISPR-Cas13a has shown great potential for the rapid and accurate detection of pathogen nucleic acids. However, conventional CRISPR-Cas13a-based assays typically require pre-amplification, which can introduce aerosol contamination and operational complexities. In this study, we developed a Minimali...

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Veröffentlicht in:Biosensors & bioelectronics 2025-02, Vol.270, p.116918, Article 116918
Hauptverfasser: Zhuang, Tianchi, Gao, Chang, Zhao, Wenwu, Yu, Hairong, Liu, Yun, Zhang, Ning, Li, Ning, Ji, Minghui
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Sprache:eng
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Zusammenfassung:CRISPR-Cas13a has shown great potential for the rapid and accurate detection of pathogen nucleic acids. However, conventional CRISPR-Cas13a-based assays typically require pre-amplification, which can introduce aerosol contamination and operational complexities. In this study, we developed a Minimalist transcription template-based Amplification-free CRISPR-Cas13a strategy for DNA detection (MAD). This strategy facilitates the release of pathogen DNA and its annealing with primers from nasopharyngeal swab samples in a straightforward manner, followed by T7 transcription and CRISPR-Cas13a detection, completing the entire process within 40 min. MAD eliminates the need for DNA extraction and pre-amplification while maintaining high sensitivity after optimization, allowing for result visualization via lateral flow strips. Furthermore, evaluation of 167 clinical pediatric samples identified 18 positive cases of human adenovirus, demonstrating a 99.4% concordance in detection compared to standard qPCR. We believe that MAD offers new insights into CRISPR-Cas diagnostics and, due to its simplicity, rapidity, and safety, is poised for widespread application in clinical practice.
ISSN:0956-5663
1873-4235
1873-4235
DOI:10.1016/j.bios.2024.116918