Designing a DNA synchronizer for compact single-rail DNA logic circuits

DNA has emerged as a robust platform for engineering molecular circuits with arbitrary logic operations. Nevertheless, implementing DNA circuits for such functions generally relies on the use of dual-rail expression that doubles the number of required gates, constraining the achievable complexity in a single solution. A fundamental limitation is that conventional single-rail circuits cannot support nonfirst-layer NOT operations. Here, we introduce the design of a DNA synchronizer (DSN), a temporal regulation module that enables time-dependent NOT function, to circumvent the fundamental limitation of conventional single-rail designs. Tuning the binding affinity between the DSN strand and an inverter strand allows for regulating the execution time of NOT gates at varying cascade depths. Single-rail NAND and NOR gates are implemented using DSNs, which are Boolean complete. We further demonstrate a 4-bit square root circuit using a minimal set of only five gates. This single-rail architecture holds promise for developing compact yet scalable DNA computing circuits while advancing applications in diagnostics and therapeutics.