CRISPR-Cas9 genome editing utilizing chemically synthesized RNA
The CRISPR-Cas9 system allows researchers to quickly edit genes for functional gene knockout in mammalian, fish, and plant genomes, among others. Consequently, this has dramatically transformed biological research. Cas9 nuclease and a guide RNA are required for CRISPR-Cas9 genome engineering; however, these components can be utilized in different reagent formats, depending upon the application. Vector-based guide RNA reagents utilize an expressed chimeric single guide RNA (sgRNA), while synthetic reagents can be either sgRNA or a two-RNA system of CRISPR RNA (crRNA) and tracrRNA as the guide RNA component. Chemical RNA synthesis has been applied for the rapid generation of either crRNA and tracrRNA or sgRNA. This allows for direct delivery into cells for unique gene editing applications such as high throughput arrayed screening or experiments that benefit from DNA-free components.