CRISPR-Cas12a Target Binding Unleashes Indiscriminate Single-stranded DNase Activity

Overview

Journal Science

Specialty Science

Date 2018 Feb 17

PMID 29449511

Citations 1100

Authors

Janice S Chen

Enbo Ma

Lucas B Harrington

Maria Da Costa

Xinran Tian

Joel M Palefsky

Jennifer A Doudna

Affiliations

Soon will be listed here.

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.

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