GSM9097140: Sample 14 Control, Ventral, 16 month; Mus musculus; RNA-Seq
Mouse dorsal and ventral hippocampus were dissected and incubated in RNAprotect Tissue Reagent (QIAGEN) at +4℃ overnight, after RNAprotect was removed samples were stored at −80°C. Nuclei were isolated separately from fresh frozen dorsal and ventral hippocampus with Chromium Nuclei Isolation Kit with RNase Inhibitor (10x Genomics) according to the Single Cell Gene Expression & Chromium Fixed RNA Profiling protocol (CG000505 Rev A). From 4 to 11 mg of tissue were used to isolate nuclei for each sample, after which nuclei were quantified using trypan blue solution 0.4% on a Countess II FL Automated Cell Counter (Thermo Fisher Scientific Inc.). snRNAseq libraries were prepared from 16 500 nuclei for each sample using Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (10x Genomics) according to the manufacturer's instructions (CG000204 Rev D, CG000315 Rev F). Quality control of prepared libraries was measured using Qubit 4.0 Flourometer (Thermo Fisher Scientific Inc.) and Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific Inc.), 4200 TapeStation System (Agilent Technologies, Inc.) and D1000 ScreenTape, D1000 Reagents (Agilent Technologies, Inc.) kits. Sequencing was performed on NovaSeq6000 and NextSeq2000 platforms (Illumina, Inc.) with a recommended coverage of 20 thousand read pairs per 1 cell, an average of 200 million read pairs per 1 sample.