title: GSM9240374: 436-BM_GP2_GEX; Mus musculus; RNA-Seq
library_construction_protocol: Isolated bone marrow mononuclear cells were processed and re-suspended in PBS+0.04% BSA at a concentration of 1000 cells/uL. Single-cell transcriptomic libraries were generated using the 10x Genomics Chromium Next GEM Single Cell 3' v3.1 chemistry. Prior to library construction, single-cell suspensions were assessed for quality using the ViaStain™ AOPI Staining Solution in combination with a Cellometer K2 automated cell counter (Nexcelom Bioscience) to determine cell concentration, viability, and to confirm the absence of aggregates or debris that might interfere with droplet encapsulation. Cells were loaded onto the Chromium X Controller (10x Genomics), where they were partitioned into nanoliter-scale Gel Beads-in-Emulsion (GEMs), each containing a single cell and a barcoded bead. Within each GEM, reverse transcription was performed to produce uniquely barcoded, full-length cDNA. Following emulsion breakage, the barcoded cDNA was purified and amplified via PCR. Amplified cDNA was enzymatically fragmented, followed by end-repair, A-tailing, adaptor ligation, and sample indexing via PCR to generate Illumina-compatible libraries. The final libraries were assessed for size distribution and integrity using the D1000 ScreenTape assay on a TapeStation 4200 (Agilent Technologies) and quantified using the KAPA Library Quantification Kit for Illumina platforms (Roche). Sequencing libraries were pooled at equimolar concentrations, denatured, and diluted to a final loading concentration of 300 pM with 1% PhiX control library (Illumina). Pooled libraries were sequenced on an Illumina NovaSeq 6000 system, following the manufacturer's protocol. An average sequencing depth of ~20,000 paired-end reads per cell was achieved for downstream analysis.