Nucleic acid detection with CRISPR-Cas13a/C2c2.
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.
|Year of Publication||
2017 04 28
|PubMed Central ID||
R01 MH110049 / MH / NIMH NIH HHS / United States
HHMI / Howard Hughes Medical Institute / United States
R01 AI117043 / AI / NIAID NIH HHS / United States
DP1 MH100706 / MH / NIMH NIH HHS / United States
K08 AI119157 / AI / NIAID NIH HHS / United States
F30 CA210382 / CA / NCI NIH HHS / United States