GSM9029987: AML3: scATACseq from CD34+ sorted bone marrow cells from AML patient; Homo sapiens; ATAC-seq
Cryopreserved bone marrow mononuclear cells were thawed in prewarmed FACS buffer (PBS + fetal bovine serum). Cells were pelleted and washed, then blocked with Human TruStain FcX™ (Biolegend #422301) for 15 minutes at 4°C. Antibodies against CD34 (Clone 561; FITC Biolegend 343603) and CD133 (Clone 7; PE Biolegend 372803) were added and incubated for 15 minutes at 4°C. Cells were washed, stained with DAPI, and sorted on a Sony SH800 cell sorter to enrich for immature populations (e.g., CD34+ or PROM+). Unsorted mononuclear cells were also used for sequencing. All samples and sorted populations were profiled using 10X Genomics Chromium Single Cell 3' v3 (for scRNA-seq) and the 10X Genomics Single Cell ATAC Reagent Kits v1.1 (for scATAC-seq). For scATAC-seq, nuclei were isolated and subjected to a transposition reaction at 37°C for 60 minutes before encapsulation with the 10X Chromium Controller targeting ~5,000 nuclei. Libraries were constructed per manufacturer instructions (10X Genomics; CG000168 RevA), assessed via Bioanalyzer High-Sensitivity DNA Kit (Agilent), and sequenced on Illumina HiSeq 2500 and NovaSeq platforms.