varlociraptor/varlociraptor-methylation-evaluation

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Overview

Latest release: v1.0.1, Last update: 2025-11-20

Linting: linting: passed, Formatting: formatting: failed

Wrappers: bio/bismark/bismark bio/bismark/deduplicate_bismark bio/bwameth/index bio/mosdepth bio/picard/markduplicates bio/reference/ensembl-sequence bio/samtools/merge bio/samtools/sort bio/sra-tools/fasterq-dump

Deployment

Step 1: Install Snakemake and Snakedeploy

Snakemake and Snakedeploy are best installed via the Conda. It is recommended to install conda via Miniforge. Run

conda create -c conda-forge -c bioconda -c nodefaults --name snakemake snakemake snakedeploy

to install both Snakemake and Snakedeploy in an isolated environment. For all following commands ensure that this environment is activated via

conda activate snakemake

For other installation methods, refer to the Snakemake and Snakedeploy documentation.

Step 2: Deploy workflow

With Snakemake and Snakedeploy installed, the workflow can be deployed as follows. First, create an appropriate project working directory on your system and enter it:

mkdir -p path/to/project-workdir
cd path/to/project-workdir

In all following steps, we will assume that you are inside of that directory. Then run

snakedeploy deploy-workflow https://github.com/varlociraptor/varlociraptor-methylation-evaluation . --tag v1.0.1

Snakedeploy will create two folders, workflow and config. The former contains the deployment of the chosen workflow as a Snakemake module, the latter contains configuration files which will be modified in the next step in order to configure the workflow to your needs.

Step 3: Configure workflow

To configure the workflow, adapt config/config.yml to your needs following the instructions below.

Step 4: Run workflow

The deployment method is controlled using the --software-deployment-method (short --sdm) argument.

To run the workflow with automatic deployment of all required software via conda/mamba, use

snakemake --cores all --sdm conda

Snakemake will automatically detect the main Snakefile in the workflow subfolder and execute the workflow module that has been defined by the deployment in step 2.

For further options such as cluster and cloud execution, see the docs.

Step 5: Generate report

After finalizing your data analysis, you can automatically generate an interactive visual HTML report for inspection of results together with parameters and code inside of the browser using

snakemake --report report.zip

Configuration

The following section is imported from the workflow’s config/README.md.

Configuration Overview

Genome Settings

Defines which genome build and version to use during alignment and reference file retrieval.

sample:
  species: "homo_sapiens"
  datatype: "dna"
  build: "GRCh38"
  release: "110"

Sequencing Platforms and Chromosome Selection

We focus our analysis on chromosome 21 for all sequencing platforms in order to avoid increasing the runtime and memory consumption. By commenting out a sequencing platform, the results for this platform will not be calculated.

seq_platforms: 
  Illumina_pe: 21
  PacBio: 21
  Nanopore: 21

Reference Methylation Callers for Benchmarking

  • Illumina bisulfite data can be compared against: bismark, bsMap, methylDackel and bisSNP

  • PacBio and Nanopore long-read platforms can be compared against: modkit, pb_CpG_tools

  • We evaluated Varlociraptor on multiple samples at the same time. To run the evaluation you have to define multi_sample and leave intentionally empty (required by downstream workflow)

ref_tools:
  Illumina_pe: [bismark, bsMap, methylDackel]
  PacBio: [modkit, pb_CpG_tools]
  Nanopore: [modkit, pb_CpG_tools]
  multi_sample: []

Input Data (Accessions and URLs)

We download the Illumina data using SRR accession numbers from the EpiQC study. To keep the structure for every platform the same we use dummy names for PacBio and Nanopore.

data:
  Illumina_pe:
    TruSeq_HG002_LAB01_REP01: [SRR13050995, SRR13050996, SRR13050998]
    TruSeq_HG002_LAB01_REP02: [SRR13050992, SRR13050993, SRR13050994]
    SPLAT_HG002_LAB01_REP01: [SRR13051055, SRR13051056, SRR13051057, SRR13051058]
    SPLAT_HG002_LAB01_REP02:  [SRR13051050, SRR13051051, SRR13051052, SRR13051054]
    MethylSeq_HG002_LAB01_REP01: [SRR13051104, SRR13051105, SRR13051106]
    MethylSeq_HG002_LAB01_REP02: [SRR13051101, SRR13051102, SRR13051103]
    EMSeq_HG002_LAB02_REP02: [SRR13051139]
    EMSeq_HG002_LAB02_REP01: [SRR13051140]
    EMSeq_HG002_LAB01_REP02: [SRR13051141]
    EMSeq_HG002_LAB01_REP01: [SRR13051142]
    TrueMethylOX_HG002_LAB01_REP02: [SRR13051230]
    TrueMethylOX_HG002_LAB01_REP01: [SRR13051231, SRR13051232]
    TrueMethylBS_HG002_LAB01_REP02: [SRR13051250, SRR13051251] 
    TrueMethylBS_HG002_LAB01_REP01: [SRR13051253] 
  PacBio:
    REP01: [pb_rep1]
    REP02: [pb_rep2]
  Nanopore:
    REP01: [np_rep1]
    REP02: [np_rep2]
  multi_sample:
    REP01: [dummy_rep1]
    REP02: [dummy_rep2]

The real PacBio and Nanopore data comes from direct download URLs:

pb_rep1: https://downloads.pacbcloud.com/public/revio/2022Q4/HG002-rep1/analysis/HG002.m84011_220902_175841_s1.GRCh38.bam
pb_rep2: https://downloads.pacbcloud.com/public/revio/2022Q4/HG002-rep2/analysis/HG002.m84005_220919_232112_s2.GRCh38.bam

np_rep1: https://42basepairs.com/download/s3/ont-open-data/giab_2025.01/basecalling/hac/HG002/PAW70337/calls.sorted.bam
np_rep2: https://42basepairs.com/download/s3/ont-open-data/giab_2025.01/basecalling/hac/HG002/PAW71238/calls.sorted.bam

Sample Definitions

Names of the individual samples under which the workflow merges the two replicates.

samples: 
  Illumina_pe: [TruSeq_HG002_LAB01, EMSeq_HG002_LAB02, EMSeq_HG002_LAB01, TrueMethylOX_HG002_LAB01, TrueMethylBS_HG002_LAB01, SPLAT_HG002_LAB01, MethylSeq_HG002_LAB01]
  PacBio: [REP]
  Nanopore: [REP]
  multi_sample: [REP]

Methylation Calling Settings

  • Filters reads below mapping quality 10.

  • Enables parallelization using scatter-gather (20 tasks).

  • Performs methylation calling at multiple FDR thresholds.

min_mapping_quality: 10
scatter_number: 20
fdr_alpha: [0.01, 1.0]

Plotting Settings

heatmap_bin_size: 5
plot_type: svg

  • Binning resolution for heatmaps (bp).

  • Output format for figures (png, svg, or html).

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