GSM9097226: ATL-CD4, scRNA-seq; Homo sapiens; RNA-Seq
Approximately 0.12 x 106 cells, viability >80%, were used.Sample was suspended in 50 μl of ice - chilled phosphate-buffered saline + 0.04% bovine serum albumin (Thermo Fisher Scientific) and processed to yield scRNA-seq libraries using 10X Chromium Next Gem Single Cell 3' Reagent Kits v 3.1 (Dual Index) (10x Genomics, Pleasanton, CA) following standard manufacturer protocols. A Bio-Rad T100 thermal cycler (Bio-Rad, Hercules, CA) was used for PCR. After library quality was assessed using Bioanalyzer 2100 (Agilent, Tokyo, Japan), samples were sequenced (Macrogen, Seoul, South Korea) on the HiseqX (Illumina, San Diego, CA) platform at a depth of 2,000,000 pair reads/sample. Library was prepared using Chromium Single Cell 3ʹ v3.1 protocol (10x Genomics). Cellular mRNA and antibody-derived oligos were reverse-transcribed and indexed with a shared cellular barcode. Indexed cDNAs were then pooled and amplified by PCR according to the Chromium Single Cell 3' v3.1 protocol (10x Genomics) and with specific primers amplifying the antibody-derived tags. SPRI bead size selection was performed in order to separate both the mRNA-derived cDNA (>300bp) and the antibody-derived tagged cDNAs (180bp). For the mRNA derived cDNA library preparation, we further proceeded according to the manufacturer's instructions for Single Cell 3' v3.1 protocol (10x Genomics).