Journal
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
Volume 57, Issue 10, Pages 2657-2661Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/anie.201713082
Keywords
CRISPR; Cas9; GFP knockout; intracellular delivery; nanomotors; nanowires
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Funding
- Defense Threat Reduction Agency Joint Science and Technology Office for Chemical and Biological Defense [HDTRA1-13-1-0002, HDTRA1-14-1-0064]
- Innovation Fund Denmark [4107-00020B]
- China Scholarship Council (CSC) [201706895002]
- Consejo Nacional de Ciencia y Tecnologia (CONACyT)
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Direct and rapid intracellular delivery of a functional Cas9/sgRNA complex using ultrasound-powered nanomotors is reported. The Cas9/sgRNA complex is loaded onto the nanomotor surface through a reversible disulfide linkage. A 5min ultrasound treatment enables the Cas9/sgRNA-loaded nanomotors to directly penetrate through the plasma membrane of GFP-expressing B16F10 cells. The Cas9/sgRNA is released inside the cells to achieve highly effective GFP gene knockout. The acoustic Cas9/sgRNA-loaded nanomotors display more than 80% GFP knockout within 2h of cell incubation compared to 30% knockout using static nanowires. More impressively, the nanomotors enable highly efficient knockout with just 0.6nm of the Cas9/sgRNA complex. This nanomotor-based intracellular delivery method thus offers an attractive route to overcome physiological barriers for intracellular delivery of functional proteins and RNAs, thus indicating considerable promise for highly efficient therapeutic applications.
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