title: GSM6604365: Ad-lib-sleep; Mus musculus; RNA-Seq
library_construction_protocol: Whole brains were quickly dissected after cervical dislocation, submerged in ice-cold PBS, embedded in OCT compound (Tissue-Tek, SAKURA Finetek), and frozen in a bath of isopentane chilled with dry ice. Coronal sections with 100-µm thickness were prepared and then the primary motor areas were further dissected. Frozen samples were homogenized using SingulatorTM 100 System (singulator 100, genomics) with Standard Nuclei Isolation according to the manufacturer's specifications. The nuclei were isolated with sucrose density gradient cenrifugation. After removing cell debris with 40 μm Flowmi™ Cell Strainer (H13680-0040, Bel-Art), the cell concentration was calculated with cell counter (CellDrop FL, DeNovix). snRNA-seq library was constructed using the Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (Dual index). Approximately 11,000-17,000 nuclei were added to reverse transcription mix targeting recovery of 10,000 nuclei.