How Cell ranger count UMI from intronic region in the latest version

Question

[tien]

Hello all,

I'm trying to understand how the latest version of cell ranger takes intronic reads into account when performing UMI counting.

They have documented about how UMI counting works but it's only for Cell ranger 3 (https://www.10xgenomics.com/resources/analysis-guides/tutorial-navigating-10x-barcoded-bam-files):

For a UMI to be counted it must meet specific criteria:

• Have a MAPQ score of 255 (see the STAR manual, section 4.2).
• Maps to exactly one gene (as shown in the GX tag of the BAM file)
• Overlaps an exon by at least 50% in a way consistent with annotated splice junctions and strand annotation.
• Records that align to exons will have an RE:A:E tag.
• Remove any records with matching UMI and Barcode values that map to different genes.

Then what is condition for intronic reads to be counted?

Thanks for your help.

[GenoMax]

Please ask 10x tech support. They are very responsive. This is a new feature and only they can provide an authoritative answer.

Post their answer here once you hear back from them.

Answer 1

[Friederike]

This link here: https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/algorithms/overview seems to suggest that intronic reads are part of the UMI matrix unless you specify that you want to exclude them. That being said, genomax is right and it's always best to check with 10X Genomics Tech Support.