Summary and Setup

Slides available here.

Author: Manveer Chauhan, Clark Lab, The University of Melbourne.
Contributors: Vini Salazar, Susan (Jiayi) Si, Raymond (Bibo) Zhao, Emma Gail, Melbourne Bioinformatics.

Last updated November 2025

Overview

Topic

Skill level

Content:
Section 1: Setup, Quality Control and Sample Integration.
Section 2: Differential Gene Expression when dealing with two treatment conditions.
Section 3: Differential Expression using a pseudobulk approach and DESeq2.

Prerequisite

Some knowledge of R and scRNA-seq analysis is assumed. We recommend reviewing the following materials before the starting the materials: - Introduction to R - 10X single-cell RNA-seq analysis in R

Please see our Intro to R and RNA-seq: counts to genes workshops if you need a refresher.

Data Sets

Click here to download the RData object that will be used for this workshop.

Software Setup

RStudio Setup

We use RStudio for coding in R.

Click here and follow the instructions to install RStudio Desktop in your system.

R packages

Run the code block below to install the packages needed for this workshop.

To check if installed properly, load each package in one at a time using the library() function.

R 

install.packages('Seurat')

if (!requireNamespace("remotes", quietly = TRUE)) {
  install.packages("remotes")
}
remotes::install_github("satijalab/seurat-data", quiet = TRUE)

if (!require("BiocManager", quietly = TRUE)) {
  install.packages("BiocManager")
}
BiocManager::install("DESeq2")
BiocManager::install("multtest") # dependency commonly missing
BiocManager::install(c("SingleR", "celldex"))
install.packages("harmony") # dependency needed for harmony analysis

install.packages("tidyverse")
install.packages("pheatmap")
install.packages("metap")
install.packages("ggplot2")