Chromium single cell platform with a single cell 5 library and gel bead kit

Libraries for scRNA-seq were prepared using the Chromium Single Cell Platform with a Single-Cell 5′ Library and Gel Bead Kit (10X Genomics, PE-1000006). Thawed PBMC samples were evaluated for viability prior to scRNA-seq analysis, and all were ≥95% viable. Cells were resuspended in a volume equivalent to 10,000 target cells for each sample, and were individually loaded onto a Chromium single-cell controller (10X Genomics) to generate single-cell gel beads-in-emulsion (GEMs). Captured cells were then lysed and the released RNA was barcoded through reverse transcription in individual GEMs. Complementary DNAs (cDNA) were generated and split to generate additional libraries γδ scTCR-seq amplicons. To this end, gene-specific primers^{25 (link)} were used within the 5′ regions of the TRGC and TRDC segments for the enrichment of TCR transcripts.
Complementary DNAs were amplified, and the quality was assessed using an Agilent 4200 Tapestation.
The scRNA-seq libraries were sequenced using an Illumina Novaseq 6000 sequencer with a paired-end 150-bp (PE150) reading strategy (performed by CapitalBio Technology) and the scTCR-seq libraries were sequenced on Illumina NextSeq 550 platform.

Libraries for scRNA-seq were prepared using the Chromium Single Cell Platform with a Single Cell 5′ Library and Gel Bead Kit (10X Genomics, 1000006). Thawed PBMC samples were evaluated for viability prior to scRNA-seq analysis, and all were ≥95% viable. Cells were resuspended in a volume equivalent to 10,000 target cells for each sample and were individually loaded onto a Chromium single-cell controller (10X Genomics) to generate single-cell gel beads-in-emulsion (GEMs). Captured cells were then lysed and the released RNA was barcoded through reverse transcription in individual GEMs. Complementary DNAs (cDNA) were generated and split to generate additional libraries of αβ TCR amplicons. Complementary DNAs were amplified, and the quality was assessed using an Agilent 4200 Tapestation.
The scRNA and scTCR libraries were sequenced using an Illumina Novaseq6000 sequencer with a paired-end 150-bp (PE150) reading strategy (performed by CapitalBio Technology).