title: GSM9191278: scRNA-seq-Control T; Homo sapiens; RNA-Seq
library_construction_protocol: Single-cell RNA sequencing was performed using the sorted CAR-T cells and control T cells (GFP⁺) cells on day 12 of culture. After sorting, dead cells were removed using a Dead Cell Removal kit (Miltenyi Biotec, #130-090-101). The cells were then resuspended using PBS with 0.04% BSA. The scRNA-Seq assay was performed according to the manufacturer's instructions (Chromium GEM-X Single Cell 3' Reagent Kits v4 (Dual Index), 10x Genomics). Briefly cells were resuspended in the reverse transcription master mix and loaded together with partitioning oil and gel beads into the chip to generate a gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every GEM was reverse transcribed into cDNA, adding an Illumina TruSeq R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The cell barcoded molecules were then cleaned up with Silane DynaBeads and amplified using 11 PCR cycles.  Next, full-length, barcoded cDNA was enzymatically fragmented, sized-selected, adapter-ligated, and amplified for library construction. During the library construction, P5, P7, i7 and i5 sample indexes, and TruSeq Read 2 were added. Samples were pooled and run on the NovaSeq X Plus sequencer using one lane of the 10B flow cell using the following sequencing parameters: R1: 28 cycles, i7: 10 cycles, i5: 10 cycles, R2: 90 cycles.
identifiers.namespace: GEO
identifiers.id: GSM9191278_r1