Logic-Gated Iontronic Sensor with Cascaded Signal Amplification for Simultaneous Detection of m5C-DNA and m6A-RNA.

DNA and RNA methylations constitute two interconnected regulatory layers of gene expression. Simultaneous detection of DNA 5-methylcytosine (m5C-DNA) and RNA N6-methyladenosine (m6A-RNA) is essential for capturing multilayer epigenetic and epitranscriptomic information while avoiding the interpretative bias that often arises from single-modification measurements. In this study, we report an iontronic sensing method based on an anodic aluminum oxide (AAO) nanochannel, which can detect both m5C-DNA and m6A-RNA. The sensor relies on modulation of the ionic current in the confined AAO nanochannel and integrates nucleic acid amplification reactions, including catalytic hairpin assembly, hybridization chain reaction, and double-strand-specific nuclease-assisted amplification strategies to achieve high sensitivity and specificity. The developed sensor exhibited sensitive responses toward m5C-DNA and m6A-RNA, with detection limits of 1.01 and 1.32 fM, respectively. The platform shows excellent selectivity, repeatability, and long-term stability. Satisfied recoveries obtained in spiked serum samples confirm its strong anti-interference capability, which is suitable for complex biological matrices. Moreover, by introducing a Boolean logic tree framework, the sensor can decode multiple iontronic signals and reduce ambiguity in dual-methylation analysis, which allows for the flexible distinction and analysis of multiple nucleic acid modification events. By integrating high-sensitivity iontronic readout with logic-guided signal interpretation, this work establishes a versatile platform for multitarget epigenetic analysis. The proposed strategy can expand the analytical capability of nanochannel-based iontronic sensors and offer new opportunities for early disease diagnostics and precision medicine applications.