title: GSM7310878: GP muscle from control mouse 1, 5 months of age [snRNA-seq]; Mus musculus; RNA-Seq
library_construction_protocol: Single nucleus RNA-seq libraries were generated using Single Cell 30 Reagent Kits v2 (10xGenomics) according to the manufacturer's protocol. Briefly, cells, reagents (10xGenomics, PN-120237), gel beads (10xGenomics, 220104), and partitioning oil (10xGenomics, 220088) were loaded into the Chromium Single Cell A Chip (10xGenomics, 2000019), targeting 2000$5000 nuclei to be recovered per channel. Chip was loaded to the Chromium Controller (10xGenomics) to generate Gel Bead-In-Emulsions (GEMs). After reverse tran- scription using GEM-RT incubation protocol, GEMs were broken with recovery reagent (10xGenomics, 220016), cDNA was purified with DynaBeads MyOne Silane beads (Thermo Scientific, 37002D), amplified with PCR for 14 cycles, and further purified with SPRI- select reagent (Beckman Coulter, B23318). Quality control of recovered PCR product was performed on a Tapestation (Agilent) with D5000 high sensitivity screentapes (Agilent, 5067-5592), and cDNA yield was measured by Qubit DNA high sensitivity assay (Invitro- gen, Q32854). After fragmentation, end-repair, A-tailing and size selection purification, cDNA was ligated with adaptor and subse- quently purified with SPRIselect reagents. Libraries were amplified using PCR with sample indexes from Chromium i7 Multiplex kit (10xGenomics, PN-120262), for a total of $14 sample index cycles, as estimated from original cDNA yield using Qubit assay. Final PCR products were subjected to double-sided size selection with SPRIselect reagents. Library quality control was performed on the Agilent Tapestation with D1000 screentapes (Agilent, 5067-5582), and library yield was quantified by Qubit DNA high sensitivity assay.