Setup

Data

All data for this lesson are included in the lesson repository under episodes/data/. No separate download is required if you have cloned or downloaded the repository.

Key data directories:

Path	Contents
`episodes/data/ssrnaseq_data/EO/`	10X Cell Ranger filtered matrices for four electric organ samples (BB48, BB49INJ, BB50, BB50INJ)
`episodes/data/ssrnaseq_data/genes.gtf.gz`	B. brachyistius RefSeq GTF for gene annotation
`episodes/data/ssrnaseq_data/gene_ontology.gaf.gz`	GO annotation file for B. brachyistius
`episodes/data/eod_duration/`	JSON files of electric organ discharge recordings

Software Setup

This lesson requires R (≥ 4.3) and RStudio (≥ 2023.06). All R packages are managed with renv; most will install automatically when you open the project. Two packages need to be installed manually before running the final episode.

Step 1 — Install R and RStudio

Download R for your operating system
Download RStudio Desktop

Step 2 — Open the project and restore the renv environment

Open the ss_rnaseq_nsb2026.Rproj file in RStudio. When prompted, run:

renv::restore()

This installs all packages recorded in the lockfile. It may take several minutes the first time.

Step 3 — Install harmony and speckle (for the final episode)

Two packages used in the “Hormone-Driven Gene Expression Changes” episode are not yet in the default lockfile and must be installed once before running that episode:

# harmony — batch correction (CRAN)
install.packages("harmony")

# speckle — differential cell-type abundance / propeller (Bioconductor)
if (!requireNamespace("BiocManager", quietly = TRUE))
  install.packages("BiocManager")
BiocManager::install("speckle")

# Record in renv after installing
renv::snapshot()

If speckle does not install cleanly in your environment, the differential abundance section of the episode includes an automatic fallback to limma (already installed), so the episode will still render without it.

Step 4 — Verify the installation

Run the following in the R console to confirm key packages are available:

required_pkgs <- c(
  "Seurat", "DESeq2", "harmony", "limma",
  "clusterProfiler", "GO.db", "tidyverse", "patchwork"
)
missing_pkgs <- required_pkgs[!sapply(required_pkgs, requireNamespace,
                                       quietly = TRUE)]
if (length(missing_pkgs) == 0) {
  message("All required packages are installed.")
} else {
  message("Missing packages: ", paste(missing_pkgs, collapse = ", "))
}

Package version reference

Package	Version used	Source
Seurat	5.4.0	CRAN
DESeq2	1.46.0	Bioconductor
harmony	≥ 1.2.0	CRAN
speckle	≥ 1.0.0	Bioconductor
limma	3.62.2	Bioconductor
clusterProfiler	≥ 4.10	Bioconductor
GO.db	≥ 3.18	Bioconductor
tidyverse	≥ 2.0	CRAN
patchwork	≥ 1.2	CRAN
glmGamPoi	≥ 1.14	Bioconductor
presto	≥ 1.0	GitHub (immunogenomics/presto)

--- title: Setup --- ## Data All data for this lesson are included in the lesson repository under `episodes/data/`. No separate download is required if you have cloned or downloaded the repository. Key data directories: | Path | Contents | |------|----------| | `episodes/data/ssrnaseq_data/EO/` | 10X Cell Ranger filtered matrices for four electric organ samples (BB48, BB49INJ, BB50, BB50INJ) | | `episodes/data/ssrnaseq_data/genes.gtf.gz` | *B. brachyistius* RefSeq GTF for gene annotation | | `episodes/data/ssrnaseq_data/gene_ontology.gaf.gz` | GO annotation file for *B. brachyistius* | | `episodes/data/eod_duration/` | JSON files of electric organ discharge recordings | ## Software Setup This lesson requires **R** (≥ 4.3) and **RStudio** (≥ 2023.06). All R packages are managed with `renv`; most will install automatically when you open the project. Two packages need to be installed manually before running the final episode. ### Step 1 — Install R and RStudio - [Download R](https://cran.r-project.org) for your operating system - [Download RStudio Desktop](https://posit.co/download/rstudio-desktop/) ### Step 2 — Open the project and restore the renv environment Open the `ss_rnaseq_nsb2026.Rproj` file in RStudio. When prompted, run: ```r renv::restore() ``` This installs all packages recorded in the lockfile. It may take several minutes the first time. ### Step 3 — Install harmony and speckle (for the final episode) Two packages used in the "Hormone-Driven Gene Expression Changes" episode are not yet in the default lockfile and must be installed once before running that episode: ```r # harmony — batch correction (CRAN) install.packages("harmony") # speckle — differential cell-type abundance / propeller (Bioconductor) if (!requireNamespace("BiocManager", quietly = TRUE)) install.packages("BiocManager") BiocManager::install("speckle") # Record in renv after installing renv::snapshot() ``` If `speckle` does not install cleanly in your environment, the differential abundance section of the episode includes an automatic fallback to `limma` (already installed), so the episode will still render without it. ### Step 4 — Verify the installation Run the following in the R console to confirm key packages are available: ```r required_pkgs <- c( "Seurat", "DESeq2", "harmony", "limma", "clusterProfiler", "GO.db", "tidyverse", "patchwork" ) missing_pkgs <- required_pkgs[!sapply(required_pkgs, requireNamespace, quietly = TRUE)] if (length(missing_pkgs) == 0) { message("All required packages are installed.") } else { message("Missing packages: ", paste(missing_pkgs, collapse = ", ")) } ``` ### Package version reference | Package | Version used | Source | |---------|-------------|--------| | Seurat | 5.4.0 | CRAN | | DESeq2 | 1.46.0 | Bioconductor | | harmony | ≥ 1.2.0 | CRAN | | speckle | ≥ 1.0.0 | Bioconductor | | limma | 3.62.2 | Bioconductor | | clusterProfiler | ≥ 4.10 | Bioconductor | | GO.db | ≥ 3.18 | Bioconductor | | tidyverse | ≥ 2.0 | CRAN | | patchwork | ≥ 1.2 | CRAN | | glmGamPoi | ≥ 1.14 | Bioconductor | | presto | ≥ 1.0 | GitHub (immunogenomics/presto) |