GSM6948208: 5M_5XFAD_rep2; Mus musculus; RNA-Seq
Nuclei from frozen mouse brain were isolated according to the protocol described in a previous report. In brief, frozen mouse frontal cortex was thawed and mechanically dissociated in ST buffer (10mM Tris-HCL pH7.5, 146mM NaCl, 1mM CaCl2, 21mM MgCl2) with 0.49% CHAPS (Millipore). Single-nuclei suspensions were filtered using a 40 μm strainer, centrifuged at 500g for 5min, and resuspended in ST buffer supplemented with 0.01% BSA (Sigma). Nuclear suspensions were counted using a hemocytometer. 10x Chromium Single Cell 3' Reagent Kit v3 (10x Genomics) was used according to manufacturer's protocol. In brief, 11,000 nuclei were loaded on the Chromium Chip (10x Genomics) and processed in the Chromium Controller. Single-nuclei were partitioned into Gel Beads-in-emulsion (GEMs), followed by RNA reverse transcription with barcoding. Libraries were created by breaking GEMs and pooling barcoded fractions, cDNA amplification, fragmentation and attachment of sample index and adapter.