GSM5605521: ApoE4-104-loxlox; Mus musculus; RNA-Seq
Once the mice were ready to be sacrificed for tissue extraction, animals were anesthetized intraperitoneally and transcardially perfused. The brain was collected and the brainstem and cerebellum were removed. For isolation of nuclei, the right cortical mantle was separated from the right hemisphere after the removal of the hippocampus and the visible white matter, and subsequently flash frozen in liquid nitrogen. Flash frozen cortical mantles were homogenized in a Dounce homogenizer in Lysis Buffer. Single nuclei isolated from the mouse cortical mantles were loaded onto the Chromium platform from 10X Genomics for droplet-based library preparation. The Chromium 3' Reagent Kits v3 was used to capture RNA molecules for amplification. Integrity and quality of the libraries were assessed using a BioAnalyzer (Agilent) at the Eli and Edythe Broad CIRM Center for Regenerative Medicine and Stem Cell Research at University of Southern California. Single nuclei isolated from the young and aged mouse cortex were loaded onto the Chromium platform from 10X Genomics for droplet-based library preparation. The Chromium 3' Reagent Kits v3 was used to capture RNA molecules for amplification. Further cDNA fragment sizes were analyzed before QC sequencing runs using an Illumina MiSeq sequencer to examine sample multiplexing and mapping rate to the reference mouse genome. Production sequencing runs were carried out on Illumina HiSeq platform to acquire more than 400 million read pairs per sample.