title: GSM7074917: B_#3 scTCR-Seq; Homo sapiens; OTHER
library_construction_protocol: PBMCs were isolated from peripheral venous blood via standard Ficoll-Paque (GE Healthcare, Uppsala, Sweden) density gradient centrifugation, frozen in freezing media, and stored in liquid nitrogen until use. All samples showed a high viability of about 94% on average after thawing. LUNA-FL™ Automated Fluorescence Cell Counter (logos biosystems) was used to consult the 10x Genomics Single Cell Protocols Cell Preparation Guide with the Guidelines for Optimal Sample Preparation flowchart (Documents CG00053 and CG000126 respectively) for more information on preparing cells. Libraries were prepared using the Chromium controller according to the 10x Chromium Next GEM Single Cell V(D)J User Guide (10x Genomics). Briefly, the cell suspensions were diluted in nuclease-free water to achieve a targeted cell count of 10,000. Cell suspension was mixed with master mix and loaded with Single Cell 5′ Gel Beads and Partitioning Oil into a Next GEM Chip G. RNA transcripts from single cells were uniquely barcoded and reverse-transcribed within droplets. cDNA molecules were pooled and enriched with PCR. For V(D)J Enriched Library, the enriched cDNA pool was first amplified using T Cell Mix1 primer and second amplified using T Cell Mix2 primer. The purified libraries were quantified using qPCR according to the qPCR Quantification Protocol Guide (KAPA) and qualified using the Agilent Technologies 4200 TapeStation (Agilent technologies). And then the libraries were sequenced using HiSeq platform (Illumina) according to the read length in the user guide. scTCR-seq