Three fastq files

Question

[charms.charms]

Hi I'm trying to download data using the SRA toolkit:

prefetch SRR24516563

and then I used:

fastq-dump SRR24516563 -split-files

The output are three different files SRR24516563_1.fastq, SRR24516563_2.fastq and SRR24516563_3.fastq , this is from a paired end sequencing experiment (single-cell RNA seq), I'm planning to use Cell Ranger to align the reads, my question is: usually the output are 3 files, how do I knew which files should I use ? Should I use files _1 and _2 ?

thanks

Answer 1

[ATpoint]

It's very simple here by looking at the reads:

• _1 is only few bp so it must be the index => ignore it
• _2 is 28bp so it must be R1 with CB/UMI
• _3 is 91 so that is R2 with the gene expression

[swbarnes2]

Note that you have to rename the files in order for cellranger to understand what is what.

https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/inputs/cr-specifying-fastqs

And yes, you can simply delete the very smallest file, it's just the sample index, and cellranger doesn't actually need it to be there.

[charms.charms]

This is a very naive question, since I'm new with this type of data, but how are able to tell the reads number, I tried looking at SRA webpage, but wasn't able to get further details.

thanks again

[Ram]

What do you mean by "reads number"? ATPoint is using their knowledge of single cell RNA-seq and read lengths to deduce which files you need to use.

[GenoMax]

If you look here you can see that there are three reads involved: https://trace.ncbi.nlm.nih.gov/Traces/?view=run_browser&acc=SRR24516563&display=metadata

[charms.charms]

Thank you for your reply !

[Ram]

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