title: GSM5929219: 48h_KO_Cont; Mus musculus; RNA-Seq
library_construction_protocol: The brain hemisphere was removed after cold PBS transcardial perfusion, and tissues were cut into pieces and transferred to the gentleMACSTM C tubes (Miltenyi Biotec) containing the Adult Brain Dissociation kit mouse and rat (Miltenyi Biotec, 130-107-677) enzyme mixture. The brain tissue dissociated with gentleMACSTM program 37°C _ABDK_1 with run 30 min incubations on the MACSmix tube rotator. The supernatant was aspirated, the pellet was resuspended in 8ml of 40% Percoll (GE Healthcare) and overlaid on the top of 4ml of 80% Percoll. The myelin debris was removed using Percoll separation. The gradient is centrifuged at 2200rpm for 20 min without brake at 22°C. Cells were collected 40% to 80% interface and washed with PBS and stained with Zombie NIRTM Fixable viability kit (Biolegend, 423106). The cells were washed by PBS for two times. BD FACS Aria III (BD Biosciences) was used for sorting brain single live cells. Each cell suspension was subjected to 3′ single-cell RNA sequencing using Single Cell A Chip Kit, Single Cell 3′ Library and Gel Bead Kit V2, and i7 Multiplex Kit (10x Genomics) with a cell recovery target of 9000, following the manufacturer's instructions