Deeptools

Source: https://github.com/aipoch/medical-research-skills

deepTools: NGS Data Analysis Toolkit

When to Use

• Use this skill when you need ngs analysis toolkit. used for bam to bigwig conversion, quality control (correlation, pca, fingerprint plots), heatmaps/feature plots (tss, peaks), suitable for chip-seq, rna-seq, atac-seq visualization in a reproducible workflow.
• Use this skill when a data analytics task needs a packaged method instead of ad-hoc freeform output.
• Use this skill when the user expects a concrete deliverable, validation step, or file-based result.
• Use this skill when scripts/validate_files.py is the most direct path to complete the request.
• Use this skill when you need the deeptools package behavior rather than a generic answer.

Key Features

• Scope-focused workflow aligned to: NGS analysis toolkit. Used for BAM to bigWig conversion, quality control (correlation, PCA, fingerprint plots), heatmaps/feature plots (TSS, peaks), suitable for ChIP-seq, RNA-seq, ATAC-seq visualization.
• Packaged executable path(s): scripts/validate_files.py plus 1 additional script(s).
• Reference material available in references/ for task-specific guidance.
• Reusable packaged asset(s), including assets/quick_reference.md.
• Structured execution path designed to keep outputs consistent and reviewable.

Dependencies

• Python: 3.10+. Repository baseline for current packaged skills.
• Third-party packages: not explicitly version-pinned in this skill package. Add pinned versions if this skill needs stricter environment control.

Example Usage

cd "20260316/scientific-skills/Data Analytics/deeptools"
python -m py_compile scripts/validate_files.py
python scripts/validate_files.py --help

Example run plan:

1. Confirm the user input, output path, and any required config values.
2. Edit the in-file CONFIG block or documented parameters if the script uses fixed settings.
3. Run python scripts/validate_files.py with the validated inputs.
4. Review the generated output and return the final artifact with any assumptions called out.

Implementation Details

See ## Overview above for related details.

• Execution model: validate the request, choose the packaged workflow, and produce a bounded deliverable.
• Input controls: confirm the source files, scope limits, output format, and acceptance criteria before running any script.
• Primary implementation surface: scripts/validate_files.py with additional helper scripts under scripts/.
• Reference guidance: references/ contains supporting rules, prompts, or checklists.
• Packaged assets: reusable files are available under assets/.
• Parameters to clarify first: input path, output path, scope filters, thresholds, and any domain-specific constraints.
• Output discipline: keep results reproducible, identify assumptions explicitly, and avoid undocumented side effects.

Overview

deepTools is a comprehensive Python command-line toolkit designed for processing and analyzing high-throughput sequencing data. With deepTools, users can perform quality control, data normalization, sample comparison, and generate publication-quality visualizations for ChIP-seq, RNA-seq, ATAC-seq, MNase-seq, and other NGS experiments.

Core Capabilities:

• Convert BAM alignment files into normalized coverage track files (bigWig/bedGraph)
• Perform quality control assessment (fingerprint plots, correlation, coverage)
• Conduct sample comparison and correlation analysis
• Generate heatmaps and profile plots around genomic features
• Perform enrichment analysis and peak region visualization

When to Use This Skill

Use this skill in the following scenarios:

• File conversion: “Convert BAM to bigWig”, “Generate coverage tracks”, “Normalize ChIP-seq data”
• Quality control: “Check ChIP quality”, “Compare replicates”, “Evaluate sequencing depth”, “QC analysis”
• Visualization: “Create heatmap around TSS”, “Plot ChIP signal”, “Visualize enrichment”, “Generate profile plot”
• Sample comparison: “Compare treatment vs control”, “Analyze sample correlation”, “PCA analysis”
• Analysis workflows: “Analyze ChIP-seq data”, “RNA-seq coverage”, “ATAC-seq analysis”, “Complete workflow”
• Specific file types: BAM files, bigWig files, BED region files in genomic contexts

Quick Start

For new deepTools users, begin with file validation and standard workflows:

1. Validate Input Files

Before running any analysis, validate BAM, bigWig, and BED files:

python scripts/validate_files.py --bam sample1.bam sample2.bam --bed regions.bed

This checks file existence, BAM indexing, and format correctness.

2. Generate Workflow Templates

For standard analyses, use the workflow generator to create customized scripts:

List available workflows

python scripts/workflow_generator.py --list

Generate a ChIP-seq QC workflow

python scripts/workflow_generator.py chipseq_qc -o qc_workflow.sh
--input-bam Input.bam --chip-bams "ChIP1.bam ChIP2.bam"
--genome-size 2913022398

Grant execution permission and run

chmod +x qc_workflow.sh ./qc_workflow.sh 3. Most Common Operations

Refer to assets/quick_reference.md for commonly used commands and parameters.

Installation uv pip install deeptools Core Workflow Pattern

deepTools workflows typically follow this pattern: QC → Normalization → Comparison/Visualization

ChIP-seq Quality Control Workflow

When users request ChIP-seq QC or quality assessment:

Use scripts/workflow_generator.py chipseq_qc to generate a workflow script

Key QC steps:

Sample correlation (multiBamSummary + plotCorrelation)

PCA analysis (plotPCA)

Coverage assessment (plotCoverage)

Fragment size validation (bamPEFragmentSize)

ChIP enrichment strength (plotFingerprint)

Result Interpretation:

Correlation: Replicates should cluster together with high correlation (>0.9)

Fingerprint plot: Strong ChIP signal shows a steep curve; a flat diagonal indicates poor enrichment

Coverage: Evaluate whether sequencing depth is sufficient

See references/workflows.md → "ChIP-seq Quality Control Workflow" for details.

Complete ChIP-seq Analysis Workflow

For full ChIP-seq analysis from BAM to visualization:

Generate normalized coverage tracks (bamCoverage)

Create comparison tracks (bamCompare to calculate log2 ratio)

Compute signal matrices around features (computeMatrix)

Generate visualizations (plotHeatmap, plotProfile)

Perform enrichment analysis at peaks (plotEnrichment)

Use scripts/workflow_generator.py chipseq_analysis to generate a template.

Full command sequence: references/workflows.md → "ChIP-seq Analysis Workflow"

RNA-seq Coverage Workflow

For strand-specific RNA-seq coverage tracks:

Use bamCoverage with --filterRNAstrand to separate forward and reverse strands.

Important: Never use --extendReads for RNA-seq (it extends across splice junctions).

Normalization:

Fixed bins: CPM

Gene-level analysis: RPKM

Template: scripts/workflow_generator.py rnaseq_coverage

Details: references/workflows.md → "RNA-seq Coverage Workflow"

ATAC-seq Analysis Workflow

ATAC-seq requires Tn5 shift correction:

Use alignmentSieve with --ATACshift to shift reads

Use bamCoverage to generate coverage tracks

Analyze fragment size (expect nucleosome ladder pattern)

Visualize signal at peaks (if peak files available)

Template: scripts/workflow_generator.py atacseq

Full workflow: references/workflows.md → "ATAC-seq Workflow"

Tool Categories and Common Tasks BAM/bigWig Processing

Convert BAM to normalized coverage file:

bamCoverage --bam input.bam --outFileName output.bw
--normalizeUsing RPGC --effectiveGenomeSize 2913022398
--binSize 10 --numberOfProcessors 8

Compare two samples (log2 ratio):

bamCompare -b1 treatment.bam -b2 control.bam -o ratio.bw
--operation log2 --scaleFactorsMethod readCount

Core tools: bamCoverage, bamCompare, multiBamSummary, multiBigwigSummary, correctGCBias, alignmentSieve

Full reference: references/tools_reference.md → "BAM and bigWig File Processing Tools"

Quality Control

Check ChIP enrichment:

plotFingerprint -b input.bam chip.bam -o fingerprint.png
--extendReads 200 --ignoreDuplicates

Sample correlation:

multiBamSummary bins --bamfiles *.bam -o counts.npz plotCorrelation -in counts.npz --corMethod pearson
--whatToShow heatmap -o correlation.png

Core tools: plotFingerprint, plotCoverage, plotCorrelation, plotPCA, bamPEFragmentSize

Full reference: references/tools_reference.md → "Quality Control Tools"

Visualization

Create heatmap around TSS: