title: GSM4760616: Donor 3  HC35 total nuclei; Homo sapiens; RNA-Seq
library_construction_protocol: Nuclei were isolated from post-mortem frozen or flash-frozen human brain tissue. Frozen tissue (50 to 100 mg) was homogenized in 15 ml of ice-cold nuclei homogenization buffer [2 M sucrose, 10 mM Hepes (pH 7.9), 25 mM KCl, 1 mM EDTA (pH 8.0), 10% glycerol, and ribonuclease (RNase) inhibitors freshly added (80 U/ml)] with Dounce tissue grinder (10 strokes with loose pestle and 10 strokes with tight pestle). The homogenate was transferred into an ultracentrifuge tube on top of 10 ml of fresh nuclei homogenization buffer and centrifuged at 24,000 rpm for 60 min at 4°C on high-speed centrifuge. The supernatant was removed, and the pellet was resuspended in 1 ml of nuclei resuspension buffer [15 mM Hepes (pH 7.4), 15 mM NaCl, 60 mM KCl, 2 mM MgCl2, 3 mM CaCl2, and RNase inhibitors freshly added (80 U/ml)] and counted on a hemocytometer. The snRNA-seq libraries were prepared by the 10x Genomics Chromium Single Cell 3′ Reagent Kit v3 chemistry according to the manufacturer's instructions and sequenced using Illumina HiSeq 4000.