Amplified detection of nucleic acids and proteins using an isothermal proximity CRISPR Cas12a assay.
Journal
Chemical science
ISSN: 2041-6520
Titre abrégé: Chem Sci
Pays: England
ID NLM: 101545951
Informations de publication
Date de publication:
04 Jan 2021
04 Jan 2021
Historique:
entrez:
24
6
2021
pubmed:
25
6
2021
medline:
25
6
2021
Statut:
epublish
Résumé
Herein, we describe an isothermal proximity CRISPR Cas12a assay that harnesses the target-induced indiscrimitive single-stranded DNase activity of Cas12a for the quantitative profiling of gene expression at the mRNA level and detection of proteins with high sensitivity and specificity. The target recognition is achieved through proximity binding rather than recognition by CRISPR RNA (crRNA), which allows for flexible assay design. A binding-induced primer extension reaction is used to generate a predesigned CRISPR-targetable sequence as a barcode for further signal amplification. Through this dual amplification protocol, we were able to detect as low as 1 fM target nucleic acid and 100 fM target protein isothermally. The practical applicability of this assay was successfully demonstrated for the temporal profiling of interleukin-6 gene expression during allergen-mediated mast cell activation.
Identifiants
pubmed: 34163977
doi: 10.1039/d0sc06113a
pii: d0sc06113a
pmc: PMC8179305
doi:
Types de publication
Journal Article
Langues
eng
Pagination
2133-2137Informations de copyright
This journal is © The Royal Society of Chemistry.
Déclaration de conflit d'intérêts
There are no conflicts to declare.
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