An RNA-Guided Cas9 Nickase-Based Method for Universal Isothermal DNA Amplification
© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim..
We have developed an ingenious method, termed Cas9 nickase-based amplification reaction (Cas9nAR), to amplify a target fragment from genomic DNA at a constant temperature of 37 °C. Cas9nAR employs a sgRNA:Cas9n complex with a single-strand nicking property, a strand-displacing DNA polymerase, and two primers bearing the cleavage sequence of Cas9n, to promote cycles of DNA replication through priming, extension, nicking, and displacement reaction steps. Cas9nAR exhibits a zeptomolar limit of detection (2 copies in 20 μL of reaction system) within 60 min and a single-base discrimination capability. More importantly, the underlying principle of Cas9nAR offers simplicity in primer design and universality in application. Considering the superior sensitivity and specificity, as well as the simple-to-implement, rapid, and isothermal features, Cas9nAR holds great potential to become a routine assay for the quantitative detection of nucleic acids in basic and applied studies.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2019 |
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| Erschienen: |
2019 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:58 |
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| Enthalten in: |
Angewandte Chemie (International ed. in English) - 58(2019), 16 vom: 08. Apr., Seite 5382-5386 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Wang, Ting [VerfasserIn] |
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| Links: |
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| Themen: |
CRISPR-Associated Protein 9 |
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| Anmerkungen: |
Date Completed 08.09.2020 Date Revised 21.02.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1002/anie.201901292 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| 520 | |a © 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. | ||
| 520 | |a We have developed an ingenious method, termed Cas9 nickase-based amplification reaction (Cas9nAR), to amplify a target fragment from genomic DNA at a constant temperature of 37 °C. Cas9nAR employs a sgRNA:Cas9n complex with a single-strand nicking property, a strand-displacing DNA polymerase, and two primers bearing the cleavage sequence of Cas9n, to promote cycles of DNA replication through priming, extension, nicking, and displacement reaction steps. Cas9nAR exhibits a zeptomolar limit of detection (2 copies in 20 μL of reaction system) within 60 min and a single-base discrimination capability. More importantly, the underlying principle of Cas9nAR offers simplicity in primer design and universality in application. Considering the superior sensitivity and specificity, as well as the simple-to-implement, rapid, and isothermal features, Cas9nAR holds great potential to become a routine assay for the quantitative detection of nucleic acids in basic and applied studies | ||
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| 650 | 4 | |a isothermal amplification | |
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| 700 | 1 | |a Sun, Huan-Huan |e verfasserin |4 aut | |
| 700 | 1 | |a Yin, Bin-Cheng |e verfasserin |4 aut | |
| 700 | 1 | |a Ye, Bang-Ce |e verfasserin |4 aut | |
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