Thursday, 27 February 2020

Seurat for single cell RNAseq data analysis

I'm learning how to use the Seurat package for analysis of single cell RNAseq data.

Here's some notes on how I installed Seurat and made some plots, using someone else's RNAseq data set.

Installing Seurat (on a Windows laptop)
- I opened Rstudio.
- I checked the R version by going to Tools > Global options, which told me I had R 3.6.1 (Seurat requires R 3.4 or greater).
- I followed the Seurat installation instructions but with a small workaround suggested on github:
  > if (!requireNamespace("BiocManager", quietly = TRUE))
  > BiocManager::install("multtest")
  > install.packages('Seurat')
  > library(Seurat)
- This installed Seurat 3.1.0.

Reading in and plotting some single cell data
I had someone else's single cell RNAseq data as an .rds file. I read it into R as a Seurat object using:
> dat <- readRDS('file.rds')
Then I made a UMAP plot of the data using:
> DimPlot(dat, label=TRUE)
Then to make a plot showing a particular gene's expression on the UMAP, e.g. gene Smp_089320 (see examples here):
> FeaturePlot(dat, features = c("Smp-089320"))

I also learnt from a a tutorial on visualisation using Seurat that you can make a violin plot for a gene using:
> VlnPlot(dat, features = c("Smp-089320"))
You should also be able to make a plot visualising gene expression in each cluster using:
> RidgePlot(dat, features = c("Smp-089320"))
but for some reason that gave me an error message.