The workflow

We have written a workflow based on snakemake to cover the end-to-end analysis. Users can provide the raw methylation files and the sample sheet as a starting point, and the pipeline will automatically:

  1. Preprocess the raw files to extract the beta values with minfi.

  2. If reference samples available, call CNVs with conumee to blacklist potentially disruptive CpGs.

  3. Call fluctuating CpG sites (If no fCpG site list is provided).

  4. Create the XML file required by BEAST (alternative starting point if beta-values are already given - trees mode).

  5. Run BEAST.

  6. Automatic model selection

  7. Summarise results and draw the phylogenetic trees.

End-to-End Methylation Analysis Workflow

Overview

Welcome to our end-to-end methylation analysis workflow powered by Snakemake! This guide will walk you through processing raw methylation files to generate phylogenetic trees.

PHYFUM's workflow

Prerequisites

Ensure you have the following installed:

  • Python

  • Snakemake

  • R with minfi and conumee packages

  • BEAST

Getting Started

Input Files

You will need:

  • Raw methylation files or beta value CSV file

  • Sample sheet

Steps

  1. Preprocessing: Extract beta values from raw files using minfi.

  2. CNV Calling: Call CNVs with conumee if reference samples are available to blacklist potentially disruptive CpGs.

  3. CpG Site Calling: Identify fluctuating CpG sites. If no fCpG site list is provided, the pipeline will generate one.

  4. Generate XML for BEAST: Create the XML file required by BEAST for beta-value inputs (trees mode).

  5. Run BEAST: Execute BEAST for phylogenetic analysis.

  6. QC: mixing and convergence checks.

  7. Model Selection: The pipeline will automatically select the appropriate model.

  8. Computation of integral solution. Integrate over each number of stem cell to compute the integrate.

  9. Result Summarization: Summarize results and draw phylogenetic trees.

Running the Pipeline

To run the pipeline, execute:

phyfum run --input exampleBeta.csv --patientinfo metadata.csv --output test  --nchains 2 --stemcells 3-6-1 --workdir test --mle-ps

Support

For any issues or questions, please reach out to our support team or check the documentation.

Enjoy your analysis!

This tutorial should help you navigate our methylation analysis workflow seamlessly. Happy analyzing!

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