GSM5774609: Skeletal muscle Rosa26 #4; Mus musculus; RNA-Seq
Total RNA was isolated from snap-frozen samples of tissues, after mechanical homogenization, using RNeasy Fibrous Tissue Mini Kit (muscle, aorta; QIAGEN) according to the manufacturer's protocol. Ion Torrent sequencing libraries were prepared according to the AmpliSeq Library prep kit protocol and as previously published (Papp et al. 2018 PLoS One) 10ng of total RNA was reverse transcribed, the resulting cDNA was amplified for 12 cycles by adding PCR Master Mix, and the AmpliSeq Mouse transcriptome gene expression primer pool (targeting 20,767 well-annotated RefSeq genes + 3,163 XM and XR genes) (based on GRCm38/mm10). Amplicons were digested with the proprietary FuPa enzyme, then barcoded adapters were ligated onto the target amplicons. The library amplicons were bound to magnetic beads, and residual reaction components were washed off. Libraries were amplified, re-purified and individually quantified using Agilent TapeStation High Sensitivity tape.