Sequence-Unconstrained DNA Computing: DSN cycling and PER circuitry for dynamic miRNAs analysis and multifunctional logic operations

Abstract

DNA strand displacement and enzyme-driven catalysis are pivotal in regulating molecular interactions and constructing molecular logic circuits. However, their application in intricate hierarchical cascading networks has been hindered by sequence crosstalk restrictions on the multiple inputs and outputs. In this study, we present a sequence-unconstrained strategy to implement versatile molecular logic circuits, relying on duplex-specific nuclease (DSN)-assisted cycling and primer exchange reaction (PER) amplification circuitry (DSN-PER). Employing linear capture probe (CP) conjugated with magnetic beads (MBs) significantly minimizes crosstalk reactions compared to the previous molecular beacon probe (MBP) for DNA barcoding. Importantly, the sequence design of “barcodes” is fully independent of target binding sites on MBs, allowing for the sensitive detection of a broad range of microRNAs (miRNAs) using the DSN-PER method. Specifically, this platform supports the programming of common logic computations (YES, NOT, OR, INHIBIT, AND, and NOR) and hierarchical circuit operation (OR-AND) in response to a specific pattern of varying miRNAs. These versatile logic circuits offer unique features with flexible sequence design, potentially broadening the applications of dynamic nucleic acid nanotechnologies in multiplexed bio-computation, intelligent diagnostics, and sophisticated DNA nanodevices.