Personal-Genome-Pipeline

Step 16b: mosdepth Coverage Statistics

Fast per-base and per-region depth statistics from BAM. Produces coverage distributions, threshold reports, and a genome-wide summary.

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What It Does

Reads the BAM file and computes:

1. Mean coverage per chromosome and genome-wide
2. Coverage distribution — cumulative fraction of bases at each depth level
3. Region coverage — mean depth in 500bp windows across the genome
4. Threshold report — fraction of bases at 1x, 5x, 10x, 15x, 20x, 30x, 50x depth

Why

• Validates sequencing depth — confirms your WGS actually has 30X (or whatever depth was ordered)
• Identifies low-coverage regions — gaps in coverage cause missed variant calls
• Detects sex chromosome anomalies — chrX depth relative to autosomes reveals XX vs XY
• More precise than indexcov — indexcov (step 16) reads only the BAM index for a quick estimate; mosdepth reads actual alignments

Tool

mosdepth v0.3.13 — fast BAM/CRAM depth calculation.

• Paper: Pedersen & Quinlan, Bioinformatics 2018 (doi:10.1093/bioinformatics/btx699)
• Source: github.com/brentp/mosdepth

Docker Image

quay.io/biocontainers/mosdepth:0.3.13--hba6dcaf_0

Command

export GENOME_DIR=/path/to/data
./scripts/16b-mosdepth.sh <sample_name>

Output

File	Description
mosdepth/<sample>.mosdepth.summary.txt	Per-chromosome mean coverage + genome total
mosdepth/<sample>.mosdepth.global.dist.txt	Cumulative coverage distribution (for plotting)
mosdepth/<sample>.regions.bed.gz	Mean depth per 500bp window
mosdepth/<sample>.thresholds.bed.gz	Fraction of bases at 1x/5x/10x/15x/20x/30x/50x

Interpreting Results

Summary file

chrom   length      bases          mean    min  max
chr1    248956422   7394254892     29.70   0    312
chr2    242193529   7185433827     29.67   0    290
...
total   3088286401  91784821430    29.72   0    312

• mean ~30: You have 30X whole-genome coverage (good for germline calling)
• mean < 15: Variant calling accuracy degrades significantly
• chrX mean ~15 (male) or ~30 (female): Expected sex-linked depth

Threshold report

Shows what fraction of each region is covered at key depths:

• >95% at 10x: Adequate for most germline variant callers
• >90% at 20x: Good quality WGS
• >80% at 30x: High-quality WGS

Runtime

Dataset	Threads	Time	Memory
30X WGS (~100 GB BAM)	4	~5-10 min	< 2 GB
chr22 BAM	2	< 30 sec	< 1 GB

MultiQC Integration

mosdepth output is automatically detected by MultiQC. The mosdepth.global.dist.txt and mosdepth.summary.txt files are consumed to generate coverage distribution plots and summary statistics in the aggregated report.

Notes

• --fast-mode skips per-base output (saves ~10 GB of disk for 30X WGS) while keeping all distributions and thresholds
• Supports ALIGN_DIR variable to use alternative alignments: ALIGN_DIR=aligned_bwamem2 ./scripts/16b-mosdepth.sh sample
• Supports CAPTURE_BED for WES on-target coverage: CAPTURE_BED=${GENOME_DIR}/captures/my_panel.bed ./scripts/16b-mosdepth.sh sample. The BED file must be inside GENOME_DIR — the Docker container only mounts GENOME_DIR as /genome/, so paths outside it are invisible to the container
• mosdepth uses 1 main thread + N decompression threads, so --threads 4 actually uses 5 threads total

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