CellRanger Workflow Not Able to Assign Cells to Samples Using 10x Genomics' Cell Multiplexing Oligos

[Amanda Hornick]

Good Afternoon, 

 

I used 10x Genomics’ 3’ CellPlex Kit Set A Cell Multiplexing Oligos (CMOs) with the Chromium Next GEM Single Cell 3’ Reagent Kit v3.1 (Dual Index) with Feature Barcode technology for Cell Multiplexing for my recent single cell RNA-sequencing experiment, and I uploaded the BCL files to a Google Bucket and ran the cumulus/cellranger_workflow/26 on Terra following the documentation here, but I have been unable to demultiplex the samples using this workflow. In particular, the output files generated by the workflow, namely “assignment_confidence_table.csv” and “tag_calls_summary.csv”, show that none of the cells in my samples can be assigned to a specific CMO, and thus, a specific sample. 

 

Have other CMO users encountered this issue? Is it likely to be an issue upstream of the computational analysis, or are there parameters I could change to help assign cells to samples? 

 

I have attached the sample sheet, one of the FeatureBarcode files, and three of the files outputted by cellranger multi for reference. 

 

Thank you,

Amanda Hornick