GSM6797789: 5xFAD Inpp5dfl/flCx3cr1ERT2Cre (5xBOAT); Mus musculus; RNA-Seq
Mice were euthanized using CO2 asphyxiation and transcardially perfused with ~25 mL of ice cold 1xPhosphate Buffered Saline. Brains were dissected out and the cortex from the right half of the brain was microdissected and flash-frozen on dry ice and stored at -80°C. Brain cortices from three individual mice were pooled based on genotype (5xFAD Inpp5dfl/flCx3cr1ERT2Cre or 5xFAD) and submitted to GENEWIZ (Azenta Life Sciences) for nuclei extraction and sequencing. Nuclei extraction was performed using the Miltenyi Nuclei Extraction Buffer (Miltenyi Biotec) following the manufacturer's guidelines with gentle MACS Dissociation and C tubes (Miltenyi Biotec). Following isolation, nuclei were counted using a Countess 3 Automated Cell Counter (Thermo Fisher). Single-nuclei 3' RNA libraries were generated using the Chromium Single Cell 3' kit (10X Genomics). For each sample, a target capture of ~3,000 Gel Beads-in-Emulsion per sample was loaded onto the Chromium Controller (10X Genomics) and processed following the standard manufacturer's specifications. Sequencing libraries were evaluated for quality using the Agilent TapeStation (Agilent Technologies) and quantified using a Qubit 2.0 Fluorometer (Invitrogen). Pooled libraries were quantified using qPCR (Applied Biosystems) and sequenced using a HiSeq 4000 (Illumina). Samples were sequenced at a configuration compatible with the recommended guidelines as outlined by 10X Genomics and raw sequence data were converted into fastq files.