CRISPR/Cas12a DTR system: a topology-guided Cas12a assay for specific dual detection of RNA and DNA targets.

ABSTRACT: The CRISPR/Cas12a technology has revolutionized molecular diagnostics. However, existing Cas12a systems depend on continuous target DNA activation, which limits them to single-target detection. In this study, we developed a novel topology-guided Cas12a system, the double-target responsive (DTR) system, capable of being activated by noncontiguous dual RNA/DNA targets. The DTR system employs two split CRISPR RNA (crRNA) fragments and two Cas12a proteins that cooperatively reconstitute upon recognizing two nucleic acid activators. We demonstrated the DTR system's ability to specifically detect dual nucleic acid substrates in a single readout, achieving a detection limit of 78 fM for RNA and exceptional specificity for single-nucleotide variations. Additionally, we successfully applied the DTR

SUBMITTER: Jiang Q 

PROVIDER: S-EPMC12421382 | biostudies-literature | 2025 Sep

REPOSITORIES: biostudies-literature