Methylation analysis presents an unusual measurement problem: many established methods do not retain the modification they are intended to study. In these workflows, methylation must instead be inferred via chemical treatment. Chemical conversion adds a time-consuming, potentially variable step that requires hazardous reagents and can fragment the sample, making some regions more difficult to map and represent evenly.
Direct sequencing of native DNA preserves methylation over large genomic distances and on individual DNA strands. With this technology, researchers can detect base modifications alongside nucleotide sequences without PCR or chemical conversion. This guide compares direct detection with short-read bisulfite sequencing and examines the decisions involved in designing targeted or whole-genome studies.
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- Compare direct native-DNA and short-read bisulfite sequencing across read depth, mapping, CpG detection, and analysis time
- Determine how read-length goals, sample throughput, and data requirements inform library preparation and sequencing setup
- Learn how to detect methylation in sequencing data, distinguish 5mC from 5hmC, and resolve modifications at single-base resolution
- Explore how brain and cancer studies integrated methylation profiling with haplotype resolution and genetic variant analysis
