GSM8262821: Male microglia, Sham, Day 3; Mus musculus; RNA-Seq
Mice were intubated, anesthesized with isoflurane and transcardially perfused. Brains were immediately harvested and only the ipsilateral hemisphere was proccessed. Brain tissue was dissociated by mechanical dissociation, filtered (70 um). A percoll gradient  were used to isolate enriched microglial cells yield (pellet) after centrifugation . Samples were resuspended in FASC buffer (RPMI1640, 2% FBS, and 2 mM EDTA). Four samples per condition were combined at this step. Next, live ZsGreen+ cells were sorted using fluorescence-activated cell sorting (BD FACSAria II) and immediately after sequenced (Novaseq 6000). 10x Genomics Chromium Controller Single-Cell Instrument (10x Genomics, Pleasanton, CA, USA) was used to generate single-cell gel beads with reverse transcription (RT) reagents in emulsions (GEMs). After GEM-RT was performed, the single-strand cDNA was purified with DynaBeads MyOne Silane Beads (Thermo Fisher Scientific). The cDNA was then amplified and purified. Indexed sequencing libraries were constructed using the Chromium Single-Cell 3' v3.1 Library Kit following the manufacture's instruction. The barcoded sequencing libraries were quantified and loaded on a Novaseq 6000 (Illumina, San Diego, CA, USA) for sequencing.