title: GSM8411031: skMt_CTRL1_Pci1426_n3; Homo sapiens; RNA-Seq
library_construction_protocol: Human skMt were lysed with RLT Plus buffer (#1030963, Qiagen, Germany). Mouse triceps biopsies were lysed with bead lysis tubes (#740814.50, Macherey-Nagel, Germany) using Qiazol (#79306, Qiagen, The Netherlands) and chloroform (#102444, Sigma-Aldrich, USA) reagents. Total RNA was isolated using an RNeasy Plus Mini extraction kit (#74134, Qiagen, Germany) according to the manufacturer's instructions. DNase I digestion (Invitrogen, USA) was performed to degrade DNA in the sample.  RNA levels and quality were checked using the NanoDrop® spectrophotometer (ThermoFisher Scientific, United States). For each sample, 100 ng of total RNA was used to perform the QuantSeq 3' mRNA-Seq Library Prep FWD for Illumina (Lexogene, Vienna, Austria) resulting in NGS libraries which originate from the 3' end of polyadenylated RNA. Briefly, library generation was started by oligo(dT) priming with primers. After first strand synthesis, the RNA was removed before the second strand synthesis was initiated by random primers. Libraries were PCR amplified and barcoded in 17 cycles and were quantified using Agilent High Sensitivity DNA kit (Agilent). 2nM of pooled libraries were denatured and 1.8pM was used for cluster generation before single-end sequencing on an Illumina Nextseq550 (Illumina, US). strategy: 3'-seq