CRISPR technologies for detecting DNA and RNA methylation: Mechanisms, platforms, and translational opportunities

DNA and RNA methylation are key epigenetic and epitranscriptomic modifications involved in gene regulation, genome stability, RNA metabolism, and disease progression. Aberrant methylation patterns in cell-free DNA and RNA have emerged as valuable biomarkers for cancer detection, disease monitoring, and therapeutic stratification. However, conventional methods such as bisulfite sequencing, methylation-specific PCR, MeRIP-seq, SCARLET, and LC-MS/MS often require harsh processing, high sample input, complex instrumentation, or lack site-specific resolution, limiting their clinical and point-of-care applications. CRISPR-based diagnostics provide a promising alternative by combining programmable nucleic acid recognition with collateral cleavage-mediated signal amplification. This review summarizes recent CRISPR strategies for detecting DNA and RNA methylation, including chemical conversion-assisted assays, restriction enzyme-mediated detection, direct amplification-free sensing based on methylation-modulated Cas activity, detection of oxidized cytosine derivatives, reverse transcription-mediated Cas12 detection of m6A, and structure-sensitive Cas13 sensing. We highlight how methylation-dependent sequence conversion, enzyme accessibility, polymerase behavior, and nucleic acid structure can be translated into CRISPR-readable signals. Finally, we discuss current translational challenges and emerging opportunities in point-of-care methylation diagnostics, integrated DNA-RNA profiling, engineered Cas effectors, AI-guided assay design, and CRISPR-compatible methylome analysis.