Programmable Nucleic Acid Sensing in Human Cells Using Circularizable ssDNA.

Programmable technologies that sense nucleic acid signatures in living cells and trigger cellular functions hold promise for biotechnology and medicine. Here, we develop SONAR (Sensing Of Nucleic acids using ASOs and Reverse-transcriptases), a platform that detects target DNA and RNA sequences and triggers controlled gene expression in human cells. SONAR operates through circularizable single-stranded DNA (ssDNA) sensors that, upon hybridization with complementary DNA or reverse-transcribed RNA, undergo target-dependent ligation via cellular ligases, subsequently driving expression of genetic payloads. For RNA sensing, we employ antisense oligonucleotides (ASOs) to prime targeted reverse transcription, generating complementary DNA that promotes ssDNA circularization. We demonstrate SONAR's ability to detect ssDNA, exogenous and endogenous RNA, couple sensing to programmable expression of diverse protein payloads, including reporters, recombinases, and genome editors, and enable enrichment and clonal recovery of target-positive cells from mixed populations. This platform establishes a versatile framework for targeted nucleic acid detection and inducible gene expression, with broad potential applications in diagnostics, therapeutics, and synthetic biology.