title: GSM4310361: young dMCAO 5d 1; Mus musculus; RNA-Seq
library_construction_protocol: Single-cell suspensions were prepared from the mouse brain. Briefly, mice were deeply anesthetized and transcardially perfused with 0.9% NaCl. The ipsilesional brain hemisphere was harvested, and cell suspensions were prepared using the Neural Tissue Dissociation Kit and gentleMACS Octo Dissociator with Heaters (Miltenyi Biotec). Suspensions were passed through a 70 μm cell strainer and fractionated on a 30% and 70% Percoll gradient at 500 g for 30 min to remove myelin and cell debris. Mononuclear cells at the interface were collected. Cells were then resuspended at 1×10^6 cells per mL, and stained with fluorophore-conjugated antibodies and the appropriate isotype controls. Fluorescence-activated cell sorting (FACS) was performed using an FACS Aria I sorter and FACS Diva software (BD Biosciences). Microglia were gated as CD11c–Ly6G–CD11b+CD45low and sorted for bulk RNA-seq. Each sample was pooled from 5 mouse brains to obtain enough cells. Total RNA was extracted from FACS-sorted microglia using RNeasy Plus micro/mini kits (Qiagen). Libraries for RNA-seq were prepared from 300 ng of total RNA with the Clonetech SMARTer Stranded Total RNA-Seq (Pico) Kit.
attributes.tag: GEO Accession
attributes.value: GSM4310361