Single Cell workshop
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Welcome to the NCGR/INBRE single cell RNA-seq analysis workshop. We will familiarize you with commonly useful R commands, the SingleCellExperiment and Seurat R packages, the Loupe Browser desktop application, and an example of a ShinyCell web application. These tools will give you basic experience in performing statistical analyses and data visualization for a variety of bioinformatic approaches to single cell sequencing.
Acknowledgments
The structure of our workshop owes a great deal to other courses and resources available online.
Bioconductor (SingleCellExperiment) and Seurat vignettes are the definitive resources for basic package functionality and examples.
This Sanger Institute course, Analysis of single cell RNA-seq data, is especially well organized.
The book by Amezquita et al., Orchestrating Single-Cell Analysis with Bioconductor (2020), goes into more detail, especially in the advanced analysis sections.
Heumos and Schaar, Single-cell best practices, in progress.
The Loupe Browser online tutorial.
Our single-cell nucleus RNA-seq dataset is a subset (sample #5) of the data from Farmer et al., Single-nucleus RNA and ATAC sequencing reveals the impact of chromatin accessibility on gene expression in Arabidopsis roots at the single-cell level, Molecular Plant 14:3, 372-383 (2021).
This workshop was developed with funding from the Institutional Development Award (IDeA) of the National Institutes of Health (NIH) & National Institute of General Medical Sciences (NIGMS) Grant # P20GM103451.
0.1 Installs
Make sure the following software is installed on your local computer before the workshop. Please contact Ethan Price at inbre@ncgr.org if you have questions.
Computer requirements:
Windows 7, 8, 10 or later
Mac Sierra or later
(No Chromebooks)
Install the correct version for your OS. Don’t plan to use the web version, as this does not have enough features.
- MobaXterm (Windows Users Only. Mac or Linux users will use their native terminal)
Download the “Home” “Installer” edition. Be careful not to install the portable or professional editions.
Before installing, extract the installer zip folder to any location on your computer.
Now run the installer and follow the prompts.
R (Latest Version for your OS)
Download and install the Loupe Browser
0.2 Agenda
ID: 82648563997
Passcode: 054689
Sessions are from 9-12 and 1-4 mountain time (M-F). Please let us know (inbre@ncgr.org or during the workshop) if you have to miss part of a day.
Zoom recordings: Please do not share these (see the Wrap Up chapter for more information).
Monday
MORNING
Introductions
Linux (Adam)
AFTERNOON
Linux (cont) (Adam)
Tuesday
MORNING
Linux Review (Adam)
Module 1: Biological context: (Adam)
AFTERNOON
Module 2: R basics (Joann)
Wednesday
MORNING
Bioconductor & SingleCellExperiment (Sven)
Quality Control (Sven)
AFTERNOON
Quality Control (continued) (Sven)
Dimensionality Reduction (Sven)
Thursday
MORNING
Seurat (Sven)
AFTERNOON
Seurat (continued) (Sven)
Integration (Sven)
Loupe Browser setup (Sven)
Friday
MORNING
Practice on other datasets (Sven)
AFTERNOON
Loupe Browser (Sven)
ShinyCell (Sven)
Catch-up and questions (Sven and Adam)
Surveys and Wrap-up (Adam)
Green text indicates hands-on sessions. Please note that the timing might shift slightly.