GSM8752931: Ntsr1; Mus musculus; RNA-Seq
Brains were harvested after rapid decapitation to avoid stress- or anesthesia-related changes in gene expression, cooled in ice-cold Hibernate-A for 2min, placed into a chilled stainless steel brain matrix and then sectioned coronally into 1mm thick slices containing the sSC (Bregma -3.0 mm to -5.0 mm). The sSC was then visualized under a dissecting microscope, micro-dissected approximately by knife cuts. Next, we homogenized the sSC in lysis buffer and separated nuclei from cellular debris using density gradient centrifugation. After resuspending the nuclei pellet, the concentration of nuclei was counted with propidium iodide by a CellDrop automated cell counter (DeNovix, Delaware, U.S.A.). We used 10X Chromium Next GEM 3' reagent kit v3.1 according to the manufacturer's protocol (user guide revision D) to capture each nucleus's poly-adenylated RNA in a droplet, process it into cDNA sequencing libraries, and sequenced to an average of 35,998 mean reads per cell.