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An aM-level sensitive cascade CRISPR-Dx system (ASCas) for rapid detection of RNA without pre-amplification.
- Source :
-
Biosensors & Bioelectronics . Jun2023, Vol. 230, pN.PAG-N.PAG. 1p. - Publication Year :
- 2023
-
Abstract
- The CRISPR/Cas system is known as one of the directions of the next generation of mainstream molecular diagnostic technology. However, most current CRISPR/Cas molecular diagnostics still rely on the pre-amplification of nucleic acid due to the limited sensitivity of CRISPR/Cas alone, which has no significant advantage over commercial Taqman-PCR and TwistAmp® Exo kits. Herein, we report an aM-level sensitive cascade CRISPR-Dx system (ASCas) that eliminates nucleic acid pre-amplification, thus avoiding aerosol contamination and greatly reducing the testing environment and personnel skill requirements for molecular diagnostics. Most importantly, the Cas13a nucleases with high sensitivity and trans -cleavage efficiency can rapidly cleaved RNA bubbles on the hybridized cascade probe at low concentration target RNA detection, which results in the destruction of the cascade probe and releases a large amount of trigger DNA for further signal amplification of secondary Cas12a reactions. Therefore, the ASCas system achieves amplification-free, ultra-sensitivity (1 aM), and ultra-fast (20 min) RNA detection. In addition, the ASCas system replaces the complicated screening process of primers and probes with the programmed Cas13a-crRNA design so that a suitable detection system can be constructed more quickly and straightforwardly for the mutation-prone SARS-CoV-2 virus. • A novel cascade CRISPR-Dx system achieves aM-level sensitivity and detects SARS-CoV-2 standard within 20 min. • The cascade probe superimposes the Cas12a and Cas13a reactions, enabling detection without nucleic acid amplification. • This system reduces specialized equipment requirements and is suitable for developing countries. • The ASCas system is suitable for detecting other RNA targets and has potential applications in cancer diagnosis. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 09565663
- Volume :
- 230
- Database :
- Academic Search Index
- Journal :
- Biosensors & Bioelectronics
- Publication Type :
- Academic Journal
- Accession number :
- 163018180
- Full Text :
- https://doi.org/10.1016/j.bios.2023.115248