GSM8412996: BM mononuclear cells from patient 2, enriched in the CD34+ HSPC fraction, scRNAseq; Homo sapiens; RNA-Seq
Cell suspensions were diluted in PBS with 0.04% BSA to a final concentration of 1 × 106 cells/mL. Vitality was assessed for each individual sample. Approximately 20,000 cells were loaded onto a single-cell chip for gel bead-in-emulsion (GEM) generation using the 10x Genomics Chromium Controller (10× Genomics). 3′mRNA-seq gene expression libraries were constructed using the Chromium Single Cell 3′ Library & Gel Bead Kit v2 (10x Genomics) according to the manufacturer's guidelines. All reactions were performed in a C1000 Touch Thermal Cycler (Bio-Rad Laboratories). Twelve cycles were used for cDNA amplification, while the number of total cycles for the sample index PCR was calculated based on the cDNA concentration. Amplified cDNA and final libraries were evaluated using a Tape Station 4200 (Agilent Technologies) with a high sensitivity chip. scRNA-seq libraries were sequenced on an Illumina NovaSeq6000 with the standard sequencing protocol of R1 28; I1 10; I2 10; R2 90 nt read length.