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A CRISPR-Cas9-triggered strand displacement amplification method for ultrasensitive DNA detection.


ABSTRACT: Although polymerase chain reaction (PCR) is the most widely used method for DNA amplification, the requirement of thermocycling limits its non-laboratory applications. Isothermal DNA amplification techniques are hence valuable for on-site diagnostic applications in place of traditional PCR. Here we describe a true isothermal approach for amplifying and detecting double-stranded DNA based on a CRISPR-Cas9-triggered nicking endonuclease-mediated Strand Displacement Amplification method (namely CRISDA). CRISDA takes advantage of the high sensitivity/specificity and unique conformational rearrangements of CRISPR effectors in recognizing the target DNA. In combination with a peptide nucleic acid (PNA) invasion-mediated endpoint measurement, the method exhibits attomolar sensitivity and single-nucleotide specificity in detection of various DNA targets under a complex sample background. Additionally, by integrating the technique with a Cas9-mediated target enrichment approach, CRISDA exhibits sub-attomolar sensitivity. In summary, CRISDA is a powerful isothermal tool for ultrasensitive and specific detection of nucleic acids in point-of-care diagnostics and field analyses.

SUBMITTER: Zhou W 

PROVIDER: S-EPMC6258682 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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A CRISPR-Cas9-triggered strand displacement amplification method for ultrasensitive DNA detection.

Zhou Wenhua W   Hu Li L   Ying Liming L   Zhao Zhen Z   Chu Paul K PK   Yu Xue-Feng XF  

Nature communications 20181127 1


Although polymerase chain reaction (PCR) is the most widely used method for DNA amplification, the requirement of thermocycling limits its non-laboratory applications. Isothermal DNA amplification techniques are hence valuable for on-site diagnostic applications in place of traditional PCR. Here we describe a true isothermal approach for amplifying and detecting double-stranded DNA based on a CRISPR-Cas9-triggered nicking endonuclease-mediated Strand Displacement Amplification method (namely CRI  ...[more]

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