CRISPR-Assisted DNA Detection: A Novel dCas9-Based DNA Detection Technique.
CRISPR J
; 3(6): 487-502, 2020 12.
Article
en En
| MEDLINE
| ID: mdl-33346711
ABSTRACT
Nucleic acid detection techniques are always critical to diagnosis, especially in the background of the present coronavirus disease 2019 pandemic. Simple and rapid detection techniques with high sensitivity and specificity are always urgently needed. However, current nucleic acid detection techniques are still limited by traditional amplification and hybridization. To overcome this limitation, here we developed CRISPR-Cas9-assisted DNA detection (CADD). In this detection, a DNA sample is incubated with a pair of capture single guide RNAs (sgRNAs; sgRNAa and sgRNAb) specific to a target DNA, dCas9, a signal readout-related probe, and an oligo-coated solid support beads or microplate at room temperature (RT) for 15 min. During this incubation, the dCas9-sgRNA-DNA complex is formed and captured on solid support by the capture sequence of sgRNAa, and the signal readout-related probe is captured by the capture sequence of sgRNAb. Finally, the detection result is reported by a fluorescent or colorimetric signal readout. This detection was verified by detecting DNA of bacteria, cancer cells, and viruses. In particular, by designing a set of sgRNAs specific to 15 high-risk human papillomaviruses (HPVs), the HPV infection in 64 clinical cervical samples was successfully detected by the method. All detections can be finished in 30 min at RT. This detection holds promise for rapid on-the-spot detection or point-of-care testing.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Técnicas de Amplificación de Ácido Nucleico
/
Proteína 9 Asociada a CRISPR
/
Hibridación de Ácido Nucleico
Tipo de estudio:
Diagnostic_studies
Límite:
Animals
/
Humans
Idioma:
En
Revista:
CRISPR J
Año:
2020
Tipo del documento:
Article