CRISPR-Cas9 is best known for genome editing, yet it derives from immune systems in bacteria. We discovered that the RNAs that guide DNA cutting by Cas9 can come from other RNAs in the cell. We learned how to dictate which RNAs become guide RNAs, thereby allowing us to connect an RNA-of-interest to cleavage of a matching DNA sequence. We converted this concept into a first-in-class diagnostic platform called LEOPARD (Leveraging Engineered tracrRNAs and On-target DNAs for PArallel RNA Detection). LEOPARD can detect a large number of RNA sequences in a single test, offering simple and rapid detection of many disease-related biomarkers that has the potential to revolutionize medical diagnostics for infectious disease, cancer, and human health.

Cellular RNAs guide CRISPR-Cas: The Cas9 nuclease widely used for genome editing is derived from natural bacterial defense systems that protect against invading viruses. Cas9 is directed by RNA guides to cut matching viral DNA. While analyzing Cas9-RNA complexes in the bacterial pathogen Campylobacter jejuni, we discovered that RNA guides can also originate from cellular RNAs unassociated with viral defense (DOI: 10.1126/science.abe7106; Perspective: 10.1126/science.abi9335). We rendered this process programmable, linking the presence of virtually any RNA to cutting of matching DNA by Cas9. This capability is the basis of our new CRISPR diagnostic method, LEOPARD (leveraging engineered tracrRNAs and on-target DNAs for parallel RNA detection), that can detect many biomarkers at once. LEOPARD can detect, e.g., RNAs from severe acute respiratory syndrome coronavirus 2 and other viruses with single-base resolution, thereby translating a new CRISPR discovery into a powerful diagnostic tool. This capability offers a first-in-class platform for multiplexed diagnostics that could disrupt how we detect infectious disease and cancer and how we evaluate our own health.

Tags: CRISPR, medical diagnostics, Covid-19, LEOPARD, RNA

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