GSM7716385: PBMC of CS3392 on 12Aug20; Homo sapiens; RNA-Seq
Single-cell RNA-seq (scRNAseq) libraries were prepared from tissue samples from Twin-CS (lymph node and bone marrow), as well as from peripheral blood mononuclear cells (PBMC) from the affected twins and unaffected sister. Tissues were dissociated into single cells using the Tumour Dissociation Kit, human (Miltenyi Biotec, Bergisch Gladbach, Germany) as per manufacturer's protocol. Each cell was captured and uniquely-barcoded using the 10X Chromium Next GEM Single Cell 5' Kit v2 (PN1000265) and 10X Chromium Controller according to manufacturer's protocol (10X Genomics, CA, USA). Following Gel Beads-in-emulsion (GEMs) generation, cell lysis and dissolution of the Gel Bead within each reaction vesicle enabled reverse transcription of polyadenylated mRNA, producing cDNA tagged with both a universal cell barcode and unique molecular index (UMI). The reaction vesicles were then lysed for cDNA amplification followed by purification using Dynabeads MyOne SILANE magnetic beads. Quantification of cDNA was performed using Agilent Bioanalyzer High Sensitivity DNA Kit (Agilent Technologies, CA, USA). The generated cDNA from each sample was used for 5' gene expression (5'GEX) scRNA-seq library generation. Enzymatic fragmentation of the cDNA transcripts was carried out, followed by End-Repair and A-tailing, adaptor ligation and final library amplification PCR with unique sample indices for each sample. Final library quality was determined using the Agilent Bioanalyzer High Sensitivity DNA Kit. scRNA-seq 10X 5' kit v2