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Hydrogel Microencapsulation Enables High-Sensitivity Mitochondrial RNA Profiling from Ultra-Low Input Samples

ABSTRACT: The 13 protein-coding genes encoded by the mitochondrial genome are essential markers for cellular energy metabolism and diverse physiological and pathological processes. However, conventional RNA-seq methods often suffer from poor sensitivity in detecting mitochondrial transcripts due to their low abundance, vulnerability to degradation, and loss during sample preparation. Here, we present a hydrogel-assisted RNA sequencing strategy that encapsulates individual RNA molecules within semi-permeable core-shell microgels (~80 μm in diameter), enabling ultra-low-input profiling, approximately equivalent to one RNA molecule per each of 1.2 million nanoliter-scale compartments. This spatial confinement minimizes intermolecular interactions, preserves mitochondrial RNA integrity, and enhances both reverse transcription and Tn5 tagmentation efficiency of mitochondrial RNA molecules. Compared with conventional RNA-seq, our method significantly improves the detection of all 13 protein-coding mitochondrial genes, even with minimal RNA input (~1 ng) and low sequencing depth (~1 million reads). This strategy provides a robust, cost-effective, and scalable approach for accurate and reproducible profiling of mitochondrial transcripts, establishing a powerful tool for investigating mitochondrial function in low-input or precious samples, with broad implications for biomedical research and clinical diagnostics.

ORGANISM(S): Homo sapiens

PROVIDER: GSE299609 | GEO | 2026/09/01

REPOSITORIES: GEO

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