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Science. 2017 Apr 28;356(6336):438-442. doi: 10.1126/science.aam9321. Epub 2017 Apr 13.

Nucleic acid detection with CRISPR-Cas13a/C2c2.

Gootenberg JS1,2,3,4,5, Abudayyeh OO1,2,3,4,6, Lee JW7, Essletzbichler P1,2,3,4, Dy AJ1,4,8, Joung J1,2,3,4, Verdine V1,2,3,4, Donghia N7, Daringer NM8, Freije CA1,9, Myhrvold C1,9, Bhattacharyya RP1, Livny J1, Regev A1,10, Koonin EV11, Hung DT1, Sabeti PC1,9,12,13, Collins JJ14,4,6,7,8, Zhang F14,2,3,4.

Author information

1
Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
2
McGovern Institute for Brain Research at MIT, Cambridge, MA 02139, USA.
3
Department of Brain and Cognitive Science, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
4
Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
5
Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
6
Department of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
7
Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02115, USA.
8
Institute for Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
9
Center for Systems Biology, Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, MA 02138, USA.
10
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
11
National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894, USA.
12
Department of Immunology and Infectious Disease, Harvard School of Public Health, Boston, MA 02115, USA.
13
Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
14
Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. zhang@broadinstitute.org jimjc@mit.edu.

Abstract

Rapid, inexpensive, and sensitive nucleic acid detection may aid point-of-care pathogen detection, genotyping, and disease monitoring. The RNA-guided, RNA-targeting clustered regularly interspaced short palindromic repeats (CRISPR) effector Cas13a (previously known as C2c2) exhibits a "collateral effect" of promiscuous ribonuclease activity upon target recognition. We combine the collateral effect of Cas13a with isothermal amplification to establish a CRISPR-based diagnostic (CRISPR-Dx), providing rapid DNA or RNA detection with attomolar sensitivity and single-base mismatch specificity. We use this Cas13a-based molecular detection platform, termed Specific High-Sensitivity Enzymatic Reporter UnLOCKing (SHERLOCK), to detect specific strains of Zika and Dengue virus, distinguish pathogenic bacteria, genotype human DNA, and identify mutations in cell-free tumor DNA. Furthermore, SHERLOCK reaction reagents can be lyophilized for cold-chain independence and long-term storage and be readily reconstituted on paper for field applications.

PMID:
28408723
PMCID:
PMC5526198
DOI:
10.1126/science.aam9321
[Indexed for MEDLINE]
Free PMC Article

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