Running scVelo on Seurat

Question

[V]

Hello,

I’m trying to run scVelo on my dataset but I’m running into some issues and was wondering if you could help me.

I have done my analysis on Seurat, and have subset and re-analysed sub-clusters and would like to run scVelo on those sub-clusters. Im trying to run it following this pipeline :

https://github.com/basilkhuder/Seurat-to-RNA-Velocity#generating-loom-files

The issue is I’ve stumbled on the first step. Is it possible to generate a loom file from Seurat that scVelo would accept and if so would you be able to give an example code of how? Trying the as.loom function in R on the subset Seurat object is being temperamental. Velocyto has been very glitchy and since this is analysis on a subcluster I’m not sure how it would need to be run as the 10x files contain all the cells and I’m only interested in a subpopulation of those.

Thank you, and sorry for the naive question.

[V]

I get this error when attempting "as.loom"

Error in loomR::create(filename = filename, data = data[feature.order,  : 
  unused arguments (feature.attrs = as.list(x = meta.feature), transpose = TRUE, calc.count = FALSE, max.size = max.size, verbose = verbose)

Have updated my Hdf5r and LoomR....

Answer 1

[Basil Khuder]

I hope this isn't too late! Seurat objects do not contain the necessary data for velocity analysis. RNA Velocity is estimated by looking at the amounts of unspliced and spliced mRNAs. Standard single cell feature counts software (such as Cell Ranger) don't provide this. If you have access to the FASTQ files used to generate the counts and subsequently your Seurat object, the Generating Loom Files portion of my tutorial will show you how you can generate files that will be accepted by scVelo. Later on in the guide, I show how you can incorporate the meta-data from your Seurat object into this RNA Velocity file.