DRD_2025_Project

Comprehensive Analysis of DNA Methylation Using the Illumina HumanMethylation450 BeadChip Platform

This repository contains the code, data, and report for a DNA methylation analysis project completed as part of the final examination for the course DNA/RNA Dynamics at the University of Bologna (2025).

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Project Overview

This project explores genome-wide DNA methylation patterns using the Illumina HumanMethylation450 BeadChip platform. The main goal is to identify differentially methylated regions (DMRs) and evaluate epigenetic modifications relevant to biological processes or disease states, using real data and a reproducible R-based pipeline.

The analysis includes:

• Data import and preprocessing of raw .idat files
• Quality control and normalization (e.g., preprocessNoob)
• Probe annotation and filtering
• Calculation of beta and M values
• Principal Component Analysis (PCA)
• Differential methylation analysis (t-test, multiple testing correction)
• Visualization and interpretation of results

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Workflow Summary

The main steps of the analysis pipeline are:

1. Project Setup: Run scripts/00setup_project_structure.R to create the required folder structure and project files.
2. Data Import: Load raw data and sample sheet using minfi.
3. Quality Control: Assess data quality using signal intensities, detection p-values, and control probes.
4. Normalization: Apply normalization (e.g., preprocessNoob) to correct technical variation.
5. Exploratory Analysis: Calculate and visualize beta/M values, perform PCA, and check for batch effects.
6. Differential Methylation: Identify differentially methylated probes between groups using statistical tests and multiple testing correction.
7. Reporting: All code and results are documented in docs/FinalReport.Rmd and rendered as HTML/PDF.

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Repository Structure

DRD_2025_Project/
├── data/                # Raw and processed data files
│   ├── raw/             # Raw .idat files and sample sheet
│   └── processed/       # RData files generated during analysis
├── docs/                # Project report, documentation, and [README](docs/README.md)
├── scripts/             # R scripts for setup and analysis
├── results/             # Output results, figures, and [README](results/README.md)
├── lib/                 # Local R package tarballs (e.g., SummarizedExperiment)
├── .gitignore           # Git ignore rules
├── LICENSE              # License file
├── CITATION.cff         # Citation file
└── README.md            # This file

• See docs/README.md for details on documentation and reports.
• See results/README.md for details on results and figures.

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How to Run the Analysis

1. Clone this repository:

   git clone https://github.com/kianinsilico/DRD_2025_Project.git
   cd DRD_2025_Project

2. Set up the project structure (optional, if not already present):

   source("scripts/00setup_project_structure.R")

3. Install required R packages:

   install.packages(c("BiocManager", "gplots", "qqman", "tinytex"))
   BiocManager::install(c(
     "minfi", "minfiData", "sva", "shinyMethyl",
     "IlluminaHumanMethylation450kmanifest",
     "IlluminaHumanMethylation450kanno.ilmn12.hg19",
     "AnnotationDbi"
   ))
   # Install SummarizedExperiment from local tarball
   install.packages("./lib/SummarizedExperiment_1.38.0.tar.gz", repos = NULL)

4. Run the analysis:

   • Follow the steps in docs/FinalReport.Rmd or run the scripts in scripts/ as needed.
   • Output and figures will be saved in results/ and docs/.

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Report & Results

• 📄 HTML Report: docs/FinalReport.html
• 📄 PDF Report: docs/FinalReport.pdf
• 📊 Results & Figures: results/

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Authors

• Keshani, K.
• Castro Vargas, P.
• Gustini, B.
• Aynede, A.
• Dobrokhotov, I.
• Otoijamun, F.

As part of the course DNA/RNA Dynamics, University of Bologna (2025).

License and Use

This project is licensed under the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License (CC BY-NC-SA 4.0).
See the full terms in the LICENSE.txt file.

Summary:

• You may use, adapt, and redistribute the material for academic purposes.
• You may not use it commercially.
• You must give appropriate credit and cite the authors.

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Citation Requirement

If you use this project (in whole or in part), you must cite it as follows:

Keshani K, Castro Vargas P, Gustini B, MohammadNejad Aynede A, Dobrokhotov I, Otoijamun F.
DRD_2025_Project: Educational pipeline for DNA methylation analysis.
University of Bologna, 2025.
GitHub: https://github.com/kianinsilico/DRD_2025_Project

If you're unsure how to cite this work, please contact the authors via GitHub Issues.

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Academic Integrity Statement

This project was created by the above authors as part of a university-assigned team effort.
Any reposting, modification, or reuse of this repository without proper attribution is considered a breach of the license and may constitute academic plagiarism.

We actively monitor the project’s use online. Reposting without citation, especially under a different name, will be reported to GitHub and the University of Bologna.

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Related Analyses by Other Groups

As part of a collaborative project for the DNA/RNA Dynamics course at the University of Bologna (2025), each group was assigned different methods or parameters for DNA methylation analysis.

To explore alternative approaches and complementary results, see the repositories below:

• 🔗 Group 4 – Martina Castellucci’s Team
  Applied: t-test and preprocessFunnorm normalization

• 🔗 Group 5 – Luca Cagnini’s Team
  Applied: Mann-Whitney test and preprocessNoob normalization

More links to other groups’ repositories will be added here as they become available.

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Contact

For questions or feedback, please open an issue or contact any of the contributors.