GSM9287002: Whole skeletal muscle sample of a mouse-APM chimera in the Pax7Cre/ERT2; Rosa26lsl-DTA host; Mus musculus; RNA-Seq
Single cell RNA sequencing (scRNA-Seq) was performed as previously described (Lenardic et al., 2024). Skeletal muscles were isolated, and 1g of muscle was homogenized, resuspended and digested in HBSS (Thermo Fisher Scientific, 14025050), containing 1.5% bovine serum albumin (BSA; AppliChem, A1391) and 0.2% Collagenase II (Thermo Fisher Scientific, 17101015) for 60-90min at 37°C in a shaking water bath. After digestion, medium consisting of low glucose DMEM (Thermo Fisher Scientific, 31885049) supplemented with 10% FBS was added to the sample. The sample was vortexed and filtered through a 100μm cell strainer. After straining, the sample was centrifuged at 350g for 5min and subsequently resuspended in 2ml ACK lysis buffer (Thermo Fisher Scientific, A1049201) and incubated on ice for 1min. After incubation, cold PBS was added to a final volume of 25ml, after the sample was strained through a 40μm cell strainer and centrifuged for 8min at 650 x g. The pellet was resuspended in FACS buffer containing 0.5% BSA and placed on ice until a live staining was performed with eBioscience™ Calcein Violet 450 AM Viability Dye (1:10000, Thermo Fisher Scientific, 65-0854-39). Live cells were sorted, and 20K cells were used for the library preparation according to the manufacturer's instruction of the Next GEM Single Cell 3' Reagent Kits v3.1 (Dual Index, 10X Genomics platform). Single cell sequencing of the libraries was performed by Novogene GmbH (Germany). According to the manufacture's protocol of the Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (Dual Index, 10X Genomics platform, PN-1000268)