Background-Free Nanopore Decoding of microRNA Expression Patterns Using Circular Diagnostic DNA

This paper describes a background-free nanopore decoding method to identify microRNA (miRNA) expression patterns that are both upregulated and downregulated. This method uses a circular DNA construct on MinION, a commercially available nanopore device. This system uses a pair of complementary diagnostic DNAs (dgDNAs) that form a closed structure too large to pass through the nanopore. This effectively prevents translocation and eliminates background signals. When the target miRNAs bind to dgDNAs, they open the circular structure, and the resulting dgDNA-miRNA complexes generate detectable events, improving the signal-to-noise ratio and enabling reliable identification of specific miRNA expression patterns. This strategy addresses a major challenge of conventional nanopore sensing, where nontarget DNA molecules generate signals that complicate target analysis. We implemented this system on the MinION device, demonstrating its potential for portable and accessible cancer diagnostics, including point-of-care testing.