GSM8062729: 2-year-old, Mxra7 KO, scRNAseq; Mus musculus; RNA-Seq
Bone marrow samples from 2-month-old and 2-year-old wild-type and Mxra7KO mice (n = 3 to 5 per group) were digested using tissue dissociation solution (RPMI 1640, Thermo Fisher Scientific) to obtain single-cell suspensions. These suspensions were subsequently filtered through 40-μm nylon sterile filters. Cell counts and viability were automatically determined and quality-controlled using a Countess® II Automated Cell Counter. In accordance with the manufacturer's protocol, Chromium Single Cell 3′ Reagent v2 kits (10X Genomics) were employed to generate barcoded single-cell RNA sequencing (scRNA-seq) libraries. The prepared cell suspension, 10X barcode gel magnetic beads, and oil were added to separate chambers of the Chromium Chip B. Subsequently, gel beads-in-emulsion (GEM) were formed using a microfluidic "double cross" system. The GEMs were then transferred to a thermal cycler for reverse transcription. The 30nt oligo-dT reverse transcription primer on the gel magnetic beads facilitated the conversion of poly-A RNA within the cells into cDNA strands containing barcode and unique molecular identifier (UMI) information. Following this, magnetic bead purification was performed to isolate the cDNA. The purified cDNA was then amplified via PCR, and its concentration and fragment size were assessed using a Qubit fluorometer and an Agilent 2100 Bioanalyzer, respectively.