An Autocatalytic Molecular Sensor Enables Rapid, Sensitive, and One-Pot Detection of Nucleic Acids

Circulating nucleic acids are emerging disease markers whose clinical applications are hindered by the lack of rapid, sensitive, convenient, and cost-effective detection assays. Inspired by the natural replication of virus, here, we developed a biomimetic one-step, one-pot, isothermal detection assay named RAPID to realize rapid and sensitive detection of nucleic acids with minimal reliance on instruments. The core element of RAPID is an autocatalytic molecular sensor that exploits the viral replication endonuclease of duck circovirus (i.e., DCV) to transform rolling circle amplification (RCA) from linear to exponential. DCV cleaves target-induced, RCA-generated amplicons into target analogs to prime secondary RCA reactions to catalytically propagate the amplification sensor. RAPID enables rapid (∼10 min), ultrasensitive (attomolar sensitivity), and direct (RNA extraction-free) detection of microRNAs and viral RNAs that is compatible with smartphone-based fluorescence detection devices. RAPID exhibited pronounced clinical translational capability by quantitatively profiling a panel of six miRNAs to achieve accurate discrimination of prostate cancer from benign prostatic hyperplasia that is exceptionally important but challenging in clinics. Furthermore, RAPID demonstrated rapid detection of influenza A viral infections of high accuracy in point-of-care settings. Simple nucleic acid detection assays like RAPID could largely promote the development and application of liquid biopsy molecular diagnostics.