Human peripheral blood mononuclear cells (PBMCs) of a healthy male donor aged 27 were obtained by 10x Genomics from AllCells.
Gene Expression, Feature Barcode, BCR Enriched, and TCR Enriched libraries were generated from 1,690 cells (91% viable, expected cell recovery 922) as described in the Chromium Single Cell V(D)J Reagent Kits v1.1 User Guide with Feature Barcoding technology for Cell Surface Protein (CG000208 Rev E) and sequenced on an Illumina NovaSeq 6000.
Sequencing Configuration:
- Gene Expression: Paired-end (26X91), single-indexed sequencing. Read 1: 26 cycles, i7 index: 8 cycles, i5 index: 0 cycles, Read 2: 91 cycles
- BCR Enriched: Paired-end (28X91), single-indexed sequencing. Read 1: 28 cycles, i7 index: 8 cycles, i5 index: 0 cycles, Read 2: 91 cycles. Note: Read 1 requirement is 26 bp; 28 bp Read 1 is trimmed by Cell Ranger pipeline.
- TCR Enriched: Paired-end (28X91), single-indexed sequencing. Read 1: 28 cycles, i7 index: 8 cycles, i5 index: 0 cycles, Read 2: 91 cycles. Note: Read 1 requirement is 26 bp; 28 bp Read 1 is trimmed by Cell Ranger pipeline.
- Feature Barcoding: Paired-end (26X91), single-indexed sequencing. Read 1: 26 cycles, i7 index: 8 cycles, i5 index: 0 cycles, Read 2: 91 cycles
Analysis Settings:
- Gene Expression: --expect-cells=1000