Running Cite-Seq-count with next GEM protocol

[yeroslaviz]

I'm trying to understand how to run the cite-seq-count with data made with the 10x protocol for Next GEM Single Cell 3' HT v3.1.

Reading the user manual for the tool, I should run the command with -R1, which containing the barcodes/UMIs and with -R2, which contain the antibody barcodes.

When I download the fastq files of the 10x protocol from here and untar them, I get two separate output folders, both folders are in paired-end mode, as is the usual for 10x data. One folder contains I1, R1 and R2 for two lanes for the RNA data, the second folders contains also I1, R1 and R2 but for the antibody results.

$ ls -1  -R 5k_pbmc_protein_v3_nextgem_fastqs/
5k_pbmc_protein_v3_nextgem_fastqs/:
5k_pbmc_protein_v3_nextgem_antibody_fastqs
5k_pbmc_protein_v3_nextgem_gex_fastqs

5k_pbmc_protein_v3_nextgem_fastqs/5k_pbmc_protein_v3_nextgem_antibody_fastqs:
5k_pbmc_protein_v3_nextgem_antibody_S2_L001_I1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_antibody_S2_L001_R1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_antibody_S2_L001_R2_001.fastq.gz
5k_pbmc_protein_v3_nextgem_antibody_S2_L002_I1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_antibody_S2_L002_R1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_antibody_S2_L002_R2_001.fastq.gz

5k_pbmc_protein_v3_nextgem_fastqs/5k_pbmc_protein_v3_nextgem_gex_fastqs:
5k_pbmc_protein_v3_nextgem_gex_S1_L001_I1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_gex_S1_L001_R1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_gex_S1_L001_R2_001.fastq.gz
5k_pbmc_protein_v3_nextgem_gex_S1_L002_I1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_gex_S1_L002_R1_001.fastq.gz
5k_pbmc_protein_v3_nextgem_gex_S1_L002_R2_001.fastq.gz

Can you please explain to me, how I use these files with the cite-seq-count command?

Do I use only the R2.fastq files as input?

thanks Assa

[Hoohm]

Hello @yeroslaviz,

you will need to run both R1 and R2 files as they are both necessary to run the command.

Have you had a look at the documentation?

It should answer most of your questions. Let me know if you still need help.

[yingyonghui]

Hi @yeroslaviz , have you solved the problem? I've just download the same files and feel very puzzled about the cite-seq-count command. Any suggestions would be appreciated!

Best yonghui