scRNA-seq analysis

Question

[kayah]

I think that I already prepare all data sets but it didn't work. Please help me. I don't know what's wrong.

(ps) barcodes.tsv.gz & genes.tsv.gz & matrix.mtx.gz are unzip files

[Ram]

Please write professionally on professional forums. Avoid IM/SMS jargon such as "plz" and multiple dots (Example: "plz help me..." should be "Please help me.") I've fixed things for you this time.

Answer 1

[GenoMax]

Files need to be named barcodes.tsv.gz, genes.tsv.gz, matrix.mtx.gz. Remove the additional stuff (GSM...) you have in the file names.

[kayah]

I changed the file names but it didn't work. But, thank you so much

[GenoMax]

What version of cellranger is this data from?

As the seurat manual says:

# For output from CellRanger < 3.0
data_dir <- 'path/to/data/directory'
list.files(data_dir) # Should show barcodes.tsv, genes.tsv, and matrix.mtx
expression_matrix <- Read10X(data.dir = data_dir)
seurat_object = CreateSeuratObject(counts = expression_matrix)

# For output from CellRanger >= 3.0 with multiple data types
data_dir <- 'path/to/data/directory'
list.files(data_dir) # Should show barcodes.tsv.gz, features.tsv.gz, and matrix.mtx.gz
data <- Read10X(data.dir = data_dir)
seurat_object = CreateSeuratObject(counts = data$`Gene Expression`)
seurat_object[['Protein']] = CreateAssayObject(counts = data$`Antibody Capture`)

You appear to have files named for older cellranger data but they are not from that version?

[kayah]

CellRanger version is 2.2.1 and I solved it by change genes to features

Thank you!! Have a good day

[jared.andrews07]

Also unless you have a very good reason to be using that old a version of CellRanger, I'd really recommend a newer one. Newer versions will do a much better job with cell filtering and generally utilize more reads in counts (as CellRanger 7+ will include intronic reads by default), resulting in more usable cells recovered.

[Ram]

A small educational note: if an answer was helpful, you should upvote it; if the answer resolved your question, you should mark it as accepted. You can accept more than one answer if they work. This will help future users that might find this post find the right answer.