Differential Expression Analysis on Optical Pooled Screens (Feldman et al., 2019)

An end-to-end bulk RNA-Seq pipeline evaluating temporal transcriptomic responses ($1,\text{h}$ vs. $4,\text{h}$) in MED12-KO and MED24-KO HeLa cell lines under cytokine stimulation ($\text{IL-1}\beta$ / $\text{TNF}-\alpha$).

This repository contains the full computational workflow developed as part of a university course project, recreating and extending the differential expression analysis framework from the dataset published by Feldman et al. (2019) (Optical Pooled Screens in Human Cells).

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Biological Background & Objective

• Biological Context: The Mediator complex subunits MED12 and MED24 act as negative regulators of NF-$\kappa\text{B}$ nuclear translocation dynamics, causing prolonged retention of the p65 subunit in the nucleus at timepoints $> 2,\text{hours}$.
• Primary Objective: Quantify transcriptomic alterations between $1,\text{hour}$ and $4,\text{hour}$ post-stimulation timepoints to isolate high-confidence consensus Differentially Expressed Genes (DEGs) and perturbed downstream inflammatory pathways.

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Repository Structure

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├── Dataset.rds                   # Processed SummarizedExperiment dataset (GEO: GSE132704)
├── Differential_Expression.Rmd   # Complete RMarkdown analytical pipeline
├── report.pdf                    # Compiled final technical report
└── README.md                     # Project documentation