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Science. 2018 Apr 27;360(6387):436-439. doi: 10.1126/science.aar6245. Epub 2018 Feb 15.

CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity.

Author information

1
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
2
Department of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
3
Department of Chemistry, University of California, Berkeley, Berkeley, CA 94720, USA.
4
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA. doudna@berkeley.edu.
5
Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94704, USA.
6
Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
7
Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.

Abstract

CRISPR-Cas12a (Cpf1) proteins are RNA-guided enzymes that bind and cut DNA as components of bacterial adaptive immune systems. Like CRISPR-Cas9, Cas12a has been harnessed for genome editing on the basis of its ability to generate targeted, double-stranded DNA breaks. Here we show that RNA-guided DNA binding unleashes indiscriminate single-stranded DNA (ssDNA) cleavage activity by Cas12a that completely degrades ssDNA molecules. We find that target-activated, nonspecific single-stranded deoxyribonuclease (ssDNase) cleavage is also a property of other type V CRISPR-Cas12 enzymes. By combining Cas12a ssDNase activation with isothermal amplification, we create a method termed DNA endonuclease-targeted CRISPR trans reporter (DETECTR), which achieves attomolar sensitivity for DNA detection. DETECTR enables rapid and specific detection of human papillomavirus in patient samples, thereby providing a simple platform for molecular diagnostics.

PMID:
29449511
DOI:
10.1126/science.aar6245
[Indexed for MEDLINE]

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