An aM-level sensitive cascade CRISPR-Dx system (ASCas) for rapid detection of RNA without pre-amplification.
Biosens Bioelectron
; 230: 115248, 2023 Jun 15.
Article
in English
| MEDLINE | ID: covidwho-2288684
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.
Keywords
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
Nucleic Acids
/
Biosensing Techniques
/
COVID-19
Type of study:
Diagnostic study
Limits:
Humans
Language:
English
Journal:
Biosens Bioelectron
Journal subject:
Biotechnology
Year:
2023
Document Type:
Article
Similar
MEDLINE
...
LILACS
LIS