Researchers at the New York Genome Center and New York University have developed a new kind of CRISPR screen technology to target RNA. Many CRISPR-based genetic screens can only edit or target DNA, and cannot target RNA viruses like coronavirus or flu. Using a recently characterized CRISPR enzyme called Cas13 that targets RNA instead of DNA, the researchers engineered a platform for massively parallel genetic screens targeting messenger RNAs of a green fluorescent protein transgene and other cell-surface proteins in human cells. By targeting thousands of different sites in human RNA transcripts, the researchers developed a machine learning-based predictive model to expedite identification of the most effective Cas13 guide RNAs. Using the model, they predicted optimized Cas13 guide RNAs for all protein-coding transcripts in the human genome. “We tiled guide RNAs across many different transcripts, including several human genes where we could easily measure transcript knock-down via antibody staining and flow cytometry,” researcher Hans-Hermann Wessels said. Their findings led them to identify a critical “seed” region that is highly sensitive to mismatches between the CRISPR guide and the target. This discovery could aid scientists in designing guide RNAs to avoid off-target activity on unintended target RNAs. Since a typical human cell expresses approximately 100,000 RNAs, accurate targeting is vital for screening and therapeutic applications. In addition to furthering scientists’ understanding of Cas13 off-targets, the “seed” region could be used for next-generation biosensors to more precisely discriminate between closely related RNA species. Cas13 enzymes traveling along an RNA landscape. Researcher Hans-Hermann Wessels, researcher Alejandro Méndez-Mancilla, and study senior author Neville Sanjana (bottom, left to right). Courtesy of Christian Stolte and New York Genome Center. Using the model derived from their massively parallel screens, the researchers identified optimal guide RNAs that could be used for future detection and therapeutic applications for the COVID-19 public health emergency caused by a coronavirus that contains an RNA genome. Predictions for Cas13 guide RNAs for a strain of SARS-CoV-2 isolated in New York have been made available online at http://bit.ly/coronavirus-guides. The research was published in Nature Biotechnology (www.doi.org/10.1038/s41587-020-0456-9). The web tool for predictive scoring of Cas13 guide RNAs can be found at http://cas13design.nygenome.org.