Unable to distinguish between [SC5P-R2, SC3Pv2] chemistries based on the R2 read mapping for Sample

[CadenZhao]

Fastqs downloaded from SRA:

My shell scripts to iteratively run cellranger count:

#!/usr/bin/bash

ref_dir=../../input/ref/refdata-gex-mm10-2020-A
fq_dir=../../input/SRA/fastq

for i in `ls ${fq_dir}`
do
    sample="${fq_dir}/${i}"
    cellranger count --id=${i} --fastqs=${sample} --transcriptome=${ref_dir} --localcores=32
done

ERROR I got:

Among 14 samples, 7 can be processed successfully:

One ERROR is different from others: I fount this file size is different from the file in SRA, I'll re-download it.

But file sizes in the other 6 ERRORs are same as files in SRA. How should I debug it?

Thank you very much!

[CadenZhao]

BTW, software version: cellranger 4.0.0

[dadkhahk]

@CadenZhao I wondered if you got any response from 10x to this issue as I got the same error message for several samples recently. Thanks! Kimia

[evolvedmicrobe]

Please contact support@10xgenomics.com for help with this issue.

[longrw]

@CadenZhao @evolvedmicrobe @dadkhahk i wonder if you got any response from 10X to this issue as i got the same error message. Thinks! longrw

[dadkhahk]

Not here. 10x usually doesn't respond here and they are more responsive through email. They will ask for some files to be provided so they can figure out the issue: support@10xgenomics.com

[longrw]

In order to distinguish between the 3' vs 5' assay configuration the following conditions need to be satisfied:

• A minimum of 1000 confidently mapped reads
• A minimum of 5.0% of the total reads considered needs to be confidently mapped
• The number of sense reads need to be at least 2x compared to the antisense reads or vice versa

Please validate the inputs and/or specify the chemistry via the --chemistry argument.