Jurkat and MM1 cells were transduced with non-target and target sgRNA. Jurkat and MM1 dCas9 parental cells were transduced separately with individual guides (either non-targeting or Rab1a sgRNA). Selected cells (cultured in a selection media) for each condition were individually frozen. Aliquots of Rab1a and non-target sgRNA-containing cells for both Jurkat and MM1 were thawed, counted, and mixed 1:1:1:1.
Approximately 16,000 cells were loaded (9,932 cells recovered). Libraries were generated as described in the Chromium Single Cell 5' Reagent Kits v2 User Guide with Feature Barcode Technology for CRISPR Screening (CG000510) using the Chromium X. Libraries were sequenced on an Illumina NovaSeq 6000 to a read depth of approximately 20,000 mean reads per cell for gene expression libraries, 20,000 mean reads for CRISPR Screening libraries, 10,000 mean reads for TCR amplified libraries, 7,000 mean reads for BCR amplified libraries, and 4,000 for cell surface protein libraries. V(D)J warnings in web summary are expected, as the MM1S cell line does not express the Ig heavy chain.
Paired-end, dual indexing:
- Read 1: 28 cycles (16 bp barcode, 10 bp UMI)
- i5 index: 10 cycles (sample index)
- i7 index: 10 cycles (sample index)
- Read 2: 90 cycles (transcript)
Analysis parameters used: --expect-cells= 10000