A multiplexed circulating tumor DNA detection platform engineered from 3D-coded interlocked DNA rings

Circulating tumor DNA (ctDNA) is a critical biomarker not only important for the early detection of tumors but also invaluable for personalized treatments. Currently ctDNA detection relies on sequencing. Here, a platform termed three-dimensional-coded interlocked DNA rings (3D-coded ID rings) was created for multiplexed ctDNA identification. The ID rings provide a ctDNA recognition ring that is physically interlocked with a reporter ring. The specific binding of ctDNA to the recognition ring initiates target-responsive cutting via a restriction endonuclease; the cutting then triggers rolling circle amplification on the reporter ring. The signals are further integrated with internal 3D codes for multiplexed readouts. ctDNAs from non-invasive clinical specimens including plasma, feces, and urine were detected and validated at a sensitivity much higher than those obtained through sequencing. This 3D-coded ID ring platform can detect any multiple DNA fragments simultaneously without sequencing. We envision that our platform will facilitate the implementation of future personalized/precision medicine.