Programmable Live-Cell CRISPR Imaging with Toehold-Switch-Mediated Strand Displacement
- 9 November 2020
- journal article
- research article
- Published by Wiley in Angewandte Chemie-International Edition
- Vol. 59 (46), 20612-20618
- https://doi.org/10.1002/anie.202009062
Abstract
The widespread application of CRISPR-Cas9 has transformed genome engineering. Nevertheless, the precision to control the targeting activity of Cas9 requires further improvement. We report a toehold-switch-based approach to engineer the conformation of single guide RNA (sgRNA) for programmable activation of Cas9. This activation circuit is responsive to multiple inputs and can regulate the conformation of the sgRNA through toehold-switch-mediated strand displacement. We demonstrate the orthogonal suppression and activation of Cas9 with orthogonal DNA inputs. Combination of toehold switches leads to a variety of intracellular Cas9 activation programs with simultaneous and orthogonal responses, through which multiple genome loci are displayed in different colors in a controllable manner. This approach provides a new route for programing CRISPR in living cells for genome imaging and engineering.Funding Information
- National Natural Science Foundation of China (21675167, 91953106, 21904060, 9195310071, 21834007, 21991134)
- Science and Technology Commission of Shanghai Municipality (19JC1410300)
This publication has 53 references indexed in Scilit:
- Programmable base editing of A•T to G•C in genomic DNA without DNA cleavageNature, 2017
- CRISPR–Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genomeNature Biotechnology, 2017
- Live Cell Imaging Reveals the Dynamics of Telomerase Recruitment to TelomeresCell, 2016
- Epigenome editing made easyNature Biotechnology, 2015
- Engineering Complex Synthetic Transcriptional Programs with CRISPR RNA ScaffoldsCell, 2015
- Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complexNature, 2014
- CRISPR-Cas systems for editing, regulating and targeting genomesNature Biotechnology, 2014
- DNA interrogation by the CRISPR RNA-guided endonuclease Cas9Nature, 2014
- Genetic Screens in Human Cells Using the CRISPR-Cas9 SystemScience, 2014
- A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial ImmunityScience, 2012