A novel method for detection of Cas9 gRNAs using a fluorophore-labeled DNA oligo

Clustered regularly interspaced short palindromic repeats (CRISPR) associated protein 9 (Cas9) has become one of the most important gene editing tools. CRISPR guide RNA (gRNA) serves as an important component in guiding Cas proteins to a target site in gene editing processes. For effective gene editing or site specific gene integration, levels of gRNA present in a target cell are critical. Especially, for prime editing, even the ratio of different gRNAs present in target cells has a major effect on gene editing efficiency. Therefore, having a convenient, highly sensitive method for direct detection of gRNAs can improve editing design and optimize targeting efficiency. Here, we have developed a convenient, highly sensitive method for direct detection of gRNAs, which may be used to improve editing design and optimize targeting efficiency.