title: GSM8682340: Control_scRNAseq_1; Mus musculus; RNA-Seq
library_construction_protocol: Brains of neonatal mice were extracted and transferred into 60mm dishes with 1X Lo Ovo buffer (1% BSA in DPBS, Sigma-Aldrich #A3803) and pial membranes were carefully removed. All procedures were done at room temperature, unless otherwise stated. Forebrains from individual neonatal mice were cut into chunks and placed in papain (20U/mL in EBSS, Worthington #LK003176，Worthington #LK003188) and DNase (200U/mL in ddH2O, Worthington #LK003170) in 1X Lo Ovo. Tissue chunks were digested for 30 min at 37 °C, then washed by 1X Lo Ovo for 2-3 times and triturated by 5 mL pipette and P-1000 pipetman. The cells were centrifuged for 15 min at 300 x g, then discarded supernatant and resuspended cell pellets with 1X Lo Ovo. Cells were kept on ice before sorting. For fluorescence-activated cell sorting (FACS), the cells were incubated with DAPI (1μg/mL, Sigma-Aldrich #D9542) for 10 min and then sorted using the BD FACSAria Fusion Flow Cytometer (BD Biosciences). After gating on morphology and singlets, DAPI-negative (live) and tdTomato+ cells were collected into a 5mL tube (Eppendorf #30119401) containing 1X Lo Ovo. The scRNA-seq libraries were constructed according to the protocol of the GEXSCOPER® Single-Cell RNA Library Kit (Singleron 4180011). Briefly, single cell suspension loaded onto the GEXSCOPE microchip which integrates single cell capture, cell lysis, capture of the cellular mRNA and molecular labeling. The barcoded cDNA was then amplified and used for the construction of single cell NGS libraries. The loading of the GEXSCOPE microchip was performed using the Singeron Matrix instrument. Individual libraries were diluted to 4 nM, pooled, and sequenced on Illumina NovaSeq 6000 with 150 bp paired end reads.