A Genome-Editing Nanomachine Constructed with a Clustered Regularly Interspaced Short Palindromic Repeats System and Activated by Near-Infrared Illumination

摘要

The RNA-guided CRISPR/Cas9 system is a powerful genome-editing technology with broad applications. Improving delivery efficiency and controllable activity of the CRISPR/Cas9 system is an area of intense research. We report the design, construction, and application of a CRISPR/Cas9 nanomachine (LACM), activated by a near-infrared (NIR) laser, which enables efficient delivery of single-guide RNA (sgRNA) into living cells and achieves controlled release of the sgRNA for the CRISPR/Cas9 activity. The LACM was constructed using a gold nanorod (AuNR) as a carrier that was decorated with dozens of protector DNAs stably hybridizing with the target binding domain of sgRNA. The DNA assembly on the AuNR protected the sgRNA. Irradiation with a NIR laser generated heat on the AuNR, resulting in controlled release of sgRNA, which guided the CRISPR/Cas9 genome editing. Successful editing of the EGFP and EMXJ genes in A549 and HEK293T cells, as well as knocking down of the PLKI gene to induce apoptosis of the target cells, highlights the promising potential of the LACM for diverse applications.