berguner/nf-core-methylseq

Methylation (Bisulfite-Sequencing) analysis pipeline using Bismark or bwa-meth + MethylDackel

Introduction

nf-core/methylseq is a bioinformatics analysis pipeline used for Methylation (Bisulfite) sequencing data. It pre-processes raw data from FastQ inputs, aligns the reads and performs extensive quality-control on the results.

The pipeline is built using Nextflow, a workflow tool to run tasks across multiple compute infrastructures in a very portable manner. It uses Docker / Singularity containers making installation trivial and results highly reproducible.

On release, automated continuous integration tests run the pipeline on a full-sized dataset on the AWS cloud infrastructure. This ensures that the pipeline runs on AWS, has sensible resource allocation defaults set to run on real-world datasets, and permits the persistent storage of results to benchmark between pipeline releases and other analysis sources.The results obtained from the full-sized test can be viewed on the nf-core website.

Pipeline Summary

The pipeline allows you to choose between running either Bismark or bwa-meth / MethylDackel. Choose between workflows by using --aligner bismark (default, uses bowtie2 for alignment), --aligner bismark_hisat or --aligner bwameth.

Step	Bismark workflow	bwa-meth workflow
Generate Reference Genome Index (optional)	Bismark	bwa-meth
Merge re-sequenced FastQ files	cat	cat
Raw data QC	FastQC	FastQC
Adapter sequence trimming	Trim Galore!	Trim Galore!
Align Reads	Bismark	bwa-meth
Deduplicate Alignments	Bismark	Picard MarkDuplicates
Extract methylation calls	Bismark	MethylDackel
Sample report	Bismark	-
Summary Report	Bismark	-
Alignment QC	Qualimap	Qualimap
Sample complexity	Preseq	Preseq
Project Report	MultiQC	MultiQC

Quick Start

1. Install Nextflow (>=22.10.1)

2. Install any of Docker, Singularity (you can follow this tutorial), Podman, Shifter or Charliecloud for full pipeline reproducibility (you can use Conda both to install Nextflow itself and also to manage software within pipelines. Please only use it within pipelines as a last resort; see docs).

3. Download the pipeline and test it on a minimal dataset with a single command:

   nextflow run nf-core/methylseq -profile test,YOURPROFILE --outdir <OUTDIR>

• Please check nf-core/configs to see if a custom config file to run nf-core pipelines already exists for your Institute. If so, you can simply use -profile <institute> in your command. This will enable either docker or singularity and set the appropriate execution settings for your local compute environment.
• If you are using singularity then the pipeline will auto-detect this and attempt to download the Singularity images directly as opposed to performing a conversion from Docker images. If you are persistently observing issues downloading Singularity images directly due to timeout or network issues then please use the --singularity_pull_docker_container parameter to pull and convert the Docker image instead. It is also highly recommended to use the NXF_SINGULARITY_CACHEDIR or singularity.cacheDir settings to store the images in a central location for future pipeline runs.
• If you are using conda, it is highly recommended to use the NXF_CONDA_CACHEDIR or conda.cacheDir settings to store the environments in a central location for future pipeline runs.

4. Start running your own analysis!

   nextflow run nf-core/methylseq --input samplesheet.csv --outdir <OUTDIR> --genome GRCh37 -profile <docker/singularity/podman/shifter/charliecloud/conda/institute>

Documentation

The nf-core/methylseq pipeline comes with documentation about the pipeline: usage and output.

Credits

These scripts were originally written for use at the National Genomics Infrastructure at SciLifeLab in Stockholm, Sweden.

• Main author:
  • Phil Ewels (@ewels)
• Contributors:
  • Rickard Hammarén (@Hammarn)
  • Alexander Peltzer (@apeltzer)
  • Patrick Hüther (@phue)

Contributions and Support

If you would like to contribute to this pipeline, please see the contributing guidelines.

For further information or help, don't hesitate to get in touch on the Slack #methylseq channel (you can join with this invite).

Citations

If you use nf-core/methylseq for your analysis, please cite it using the following doi: 10.5281/zenodo.1343417

An extensive list of references for the tools used by the pipeline can be found in the CITATIONS.md file.

You can cite the nf-core publication as follows:

The nf-core framework for community-curated bioinformatics pipelines.

Philip Ewels, Alexander Peltzer, Sven Fillinger, Harshil Patel, Johannes Alneberg, Andreas Wilm, Maxime Ulysse Garcia, Paolo Di Tommaso & Sven Nahnsen.

Nat Biotechnol. 2020 Feb 13. doi: 10.1038/s41587-020-0439-x.