bisulfite-sequencing-data-loading

bisulfite-sequencing-data-loading

Summary

Load and parse DNA methylation call files from bisulfite sequencing experiments (RRBS, WGBS, target-capture) into in-memory or disk-backed R objects, supporting both standard text formats and tabix-indexed bgzipped storage for memory-efficient batch processing.

When to use

You have raw methylation call files from Bismark, MethylDackel, or similar bisulfite alignment tools (bedGraph, cytosine report, or tabix-indexed formats) and need to import them into R as methylRaw or methylRawListDB objects for downstream differential methylation analysis, quality filtering, or annotation.

When NOT to use

• Input files are already formatted as a unified methylBase object or methylDiff object (skip directly to filtering or differential methylation analysis)
• You are working with non-bisulfite methylation data (e.g., whole-genome or targeted 5hmC without proper bisulfite conversion controls)
• Raw sequencing reads (FASTQ) are provided instead of methylation call files; align with Bismark or MethylDackel first

Inputs

• Methylation call files (cytosineReport, bedGraph, or tabix-indexed bgzipped format from Bismark/MethylDackel)
• Sample metadata (treatment/condition assignment)
• Assembly/genome identifier (e.g., 'hg19', 'mm9')

Outputs

• methylRaw object (single sample, in-memory)
• methylRawList object (multiple samples, in-memory)
• methylRawListDB object (multiple samples, disk-backed tabix storage)

How to apply

Use the methRead() function from methylKit to read methylation call files, specifying the file format (e.g., 'cytosineReport', 'bedGraph') and assembly. For memory-constrained analyses with large sample cohorts, set dbtype='tabix' to create a methylRawListDB object backed by bgzipped tabix-indexed files on disk rather than in-memory storage. By default, methRead() applies a minimum coverage threshold of 10 reads per base to ensure quality methylation percentage estimates. Verify the resulting methylRaw or methylRawListDB object structure and confirm that the dbpath slot references valid bgzipped tabix file paths when using database-mode storage. The function automatically handles CpG, CHG, and CHH context methylation calls depending on your input file's content and the bisulfite sequencing protocol (RRBS vs. WGBS).