Indexed by:
Abstract:
CRISPR/Cas12a, as a powerful and programmable biosensing tool, has brought great convenience in visual detection of the target in a sequence-specific mode. Though fluorescent output is the predominant way of present visual methods, this strategy highly depends on cleaving fluorophore-/quencher-labeled oligonucleotides. Few research focuses on realizing fluorescent visual detection without DNA-labeling. Herein, we have proposed a CRISPR/Cas12a-based label-free fluorescent visual detection strategy. The basic principle of this strategy is that G-quadruplex can enhance the fluorescent emission of fluorescent ligands while dsDNA target-activated Cas12a make them impotent by disrupting the higher-ordered structure. After comparing four kinds of G-quadruplexspecific ligands, we selected Thioflavin T (ThT) to achieve the visual observation goal. The application feasibility was confirmed by combining with loop-mediated isothermal amplification (LAMP) for detection of Vibrio parahaemolyticus (V. parahaemolyticus), and it demonstrated a high sensitivity (1.36 x 10(2) copies) and specificity (among 12 kinds of bacteria). The method displayed similar performance as the real-time PCR for detection of real samples yet had lower requirement on the instrument. The signal output format could also distinguish Salmo salar from other seven kinds of sequence-similar Salmonidae. The findings and strategy proposed in this manuscript will expand the application of CRISPR/Cas12a-based label-free fluorescent analysis.
Keyword:
Reprint 's Address:
Email:
Version:
Source :
SENSORS AND ACTUATORS B-CHEMICAL
ISSN: 0925-4005
Year: 2022
Volume: 370
8 . 4
JCR@2022
8 . 0 0 0
JCR@2023
ESI Discipline: CHEMISTRY;
ESI HC Threshold:74
JCR Journal Grade:1
CAS Journal Grade:1
Cited Count:
WoS CC Cited Count: 6
SCOPUS Cited Count: 7
ESI Highly Cited Papers on the List: 0 Unfold All
WanFang Cited Count:
Chinese Cited Count:
30 Days PV: 3
Affiliated Colleges: