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M−CDC: Magnetic pull-down-assisted colorimetric method based on the CRISPR/Cas12a system.
- Source :
-
Methods . Jul2022, Vol. 203, p259-267. 9p. - Publication Year :
- 2022
-
Abstract
- • The M−CDC effectively integrates the advantages of CRISPR/Cas12a, magnetic beads-based separation, and AuNP bioprobes. • M−CDC is an easy-to-operated, sensitive, and specific biosensing platform for nucleic acid assay. • The M−CDC has a great potential to be used for large-scale population SARS-CoV-2 screening in resource-limited areas. The construction of a rapid, simple, and specific nucleic acid detection platform is of great significance to the control of the large-scale spread of infectious diseases. We have recently established a magnetic pull-down-assisted colorimetric method based on the CRISPR/Cas12a system (termed M−CDC), which effectively integrates the advantages of CRISPR/Cas12a, magnetic beads-based separation, and AuNP bioprobe to provide a simple and specific biosensing platform for nucleic acid assay. The M−CDC method is compatible with point-of-care testing and enables the detection of nucleic acid samples in less than an hour without relying on expensive and complex instruments. In this paper, step-by-step instructions for M−CDC assay, including recombinase polymerase amplification (RPA)/reverse transcription-polymerase chain reaction (RT-RPA) of DNA or RNA, Cas12a-mediated target recognition and cleavage, and subsequent magnetic beads-mediated colorimetric readouts are provided. In addition, the protocol for the expression and purification of Lachnospiraceae bacterium -Cas12a (LbCas12a) protein, the design and synthesis of high-efficient crRNA, and the preparation of AuNP bioprobe are also offered. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 10462023
- Volume :
- 203
- Database :
- Academic Search Index
- Journal :
- Methods
- Publication Type :
- Academic Journal
- Accession number :
- 157503600
- Full Text :
- https://doi.org/10.1016/j.ymeth.2021.11.009