Direct delivery of Cas9-sgRNA ribonucleoproteins into cells using a nanoneedle array

Abstract

The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is a powerful and widely used tool for genome editing. Recently, it was reported that direct delivery of Cas9-sgRNA ribonucleoproteins (RNPs) reduced off-target effects. Therefore, non-invasive, high-throughput methods are needed for direct delivery of RNPs into cells. Here, we report a novel method for direct delivery of RNPs into cells using a nanostructure with a high-aspect-ratio and uniform nanoneedles. This nanostructure is composed of tens of thousands of nanoneedles laid across a 2D array. Through insertion of the nanoneedle array previously adsorbed with Cas9-sgRNA, it was possible to deliver RNPs directly into mammalian cells for genome editing.