CRISPR Cas9 is a gene editing tool that has increased in popularity due to its simplicity to use. It allows researchers to seamlessly edit DNA sequences by combining a sequence identifying guide RNA (gRNA) with the Cas9 endonuclease enzyme. However, its applicability as a gene-editing and therapeutic tool is impeded due to undesired off-target binding of Cas9.

Conventional bulk in vitro assays provide information on the DNA sequences but do not give insights into the changes in the local DNA structure. In this application note, dynamic single molecule (DSM) analysis is shown to be a novel and a more powerful investigative tool to directly visualize and manipulate CRISPR Cas9 – DNA interactions, and accelerate gene-editing researchand therapy development.