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Topological barrier to Cas12a activation by circular DNA nanostructures facilitates autocatalysis and transforms DNA/RNA sensing.
Deng, Fei; Li, Yi; Yang, Biyao; Sang, Rui; Deng, Wei; Kansara, Maya; Lin, Frank; Thavaneswaran, Subotheni; Thomas, David M; Goldys, Ewa M.
Afiliación
  • Deng F; Graduate School of Biomedical Engineering, Faculty of Engineering, University of New South Wales, Sydney, NSW, 2052, Australia.
  • Li Y; ARC Centre of Excellence for Nanoscale Biophotonics, University of New South Wales, Sydney, NSW, 2052, Australia.
  • Yang B; Graduate School of Biomedical Engineering, Faculty of Engineering, University of New South Wales, Sydney, NSW, 2052, Australia. yi.li6@unsw.edu.au.
  • Sang R; ARC Centre of Excellence for Nanoscale Biophotonics, University of New South Wales, Sydney, NSW, 2052, Australia. yi.li6@unsw.edu.au.
  • Deng W; Graduate School of Biomedical Engineering, Faculty of Engineering, University of New South Wales, Sydney, NSW, 2052, Australia.
  • Kansara M; ARC Centre of Excellence for Nanoscale Biophotonics, University of New South Wales, Sydney, NSW, 2052, Australia.
  • Lin F; Graduate School of Biomedical Engineering, Faculty of Engineering, University of New South Wales, Sydney, NSW, 2052, Australia.
  • Thavaneswaran S; ARC Centre of Excellence for Nanoscale Biophotonics, University of New South Wales, Sydney, NSW, 2052, Australia.
  • Thomas DM; School of Biomedical Engineering, University of Technology Sydney, Sydney, NSW, 2007, Australia.
  • Goldys EM; Garvan Institute of Medical Research, Darlinghurst, Sydney, NSW, 2011, Australia.
Nat Commun ; 15(1): 1818, 2024 Mar 05.
Article en En | MEDLINE | ID: mdl-38443394
ABSTRACT
Control of CRISPR/Cas12a trans-cleavage is crucial for biosensor development. Here, we show that small circular DNA nanostructures which partially match guide RNA sequences only minimally activate Cas12a ribonucleoproteins. However, linearizing these structures restores activation. Building on this finding, an Autocatalytic Cas12a Circular DNA Amplification Reaction (AutoCAR) system is established which allows a single nucleic acid target to activate multiple ribonucleoproteins, and greatly increases the achievable reporter cleavage rates per target. A rate-equation-based model explains the observed near-exponential rate trends. Autocatalysis is also sustained with DNA nanostructures modified with fluorophore-quencher pairs achieving 1 aM level (<1 copy/µL) DNA detection (106 times improvement), without additional amplification, within 15 min, at room temperature. The detection range is tuneable, spanning 3 to 11 orders of magnitude. We demonstrate 1 aM level detection of SNP mutations in circulating tumor DNA from blood plasma, genomic DNA (H. Pylori) and RNA (SARS-CoV-2) without reverse transcription as well as colorimetric lateral flow tests of cancer mutations with ~100 aM sensitivity.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Helicobacter pylori / Nanoestructuras Idioma: En Revista: Nat Commun Asunto de la revista: BIOLOGIA / CIENCIA Año: 2024 Tipo del documento: Article País de afiliación: Australia Pais de publicación: Reino Unido
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Helicobacter pylori / Nanoestructuras Idioma: En Revista: Nat Commun Asunto de la revista: BIOLOGIA / CIENCIA Año: 2024 Tipo del documento: Article País de afiliación: Australia Pais de publicación: Reino Unido