Accurate targeted long-read DNA methylation and hydroxymethylation sequencing with TAPS

Area of Science:

• Epigenetics
• Genomics
• Molecular Biology

Background:

• DNA methylation and hydroxymethylation are critical epigenetic modifications.
• Accurate detection of these modifications, especially in complex genomic regions, remains challenging.
• Existing methods often lack long-range phasing capabilities.

Purpose of the Study:

• To introduce a novel long-read sequencing method for base-resolution detection of DNA methylation and hydroxymethylation.
• To enable long-range epigenetic phasing across large genomic regions.
• To analyze challenging genomic loci and viral DNA methylation.

Main Methods:

• Development of Tet-assisted pyridine borane sequencing (lrTAPS) compatible with long-read platforms.
• Targeted sequencing of genomic regions up to 10 kb with nanogram-level input DNA.
• Application of lrTAPS to mouse embryonic stem cells and hepatitis B virus genomes.

Main Results:

• lrTAPS achieves methylation detection accuracy comparable to short-read sequencing.
• The method provides long-range epigenetic phasing, revealing haplotype-specific methylation patterns.
• Successfully sequenced difficult-to-map regions and identified methylation events in integrated HBV DNA.

Conclusions:

• lrTAPS is a powerful tool for base-resolution epigenomic analysis in long reads.
• It overcomes limitations of existing methods for studying complex epigenetic landscapes.
• Enables new insights into epigenetic regulation in both host genomes and viral infections.