title: GSM8728815: Control treated parental tumor 5; Mus musculus; RNA-Seq
library_name: GSM8728815
library_construction_protocol: Tumors were collected in PureZOL Reagent (BioRad) and stored at -80°C. For processing, tumors were dissociated with 5mm Stainless steel beads (Qiagen) using the TissueLyser II (QIAGEN). Total RNA was extracted using the Direct-zol RNA Mini Prep Kit (Zymo Research) following the manufacturer's recommendations and included a DNase digestion step. RNA was quantified using a NanoDrop One (ThermoFisher) or a Bioanalyzer (Agilent Technologies) for RNA-seq. For qPCR, total RNA was extracted from cells using RLT lysis buffer (QIAGEN) and purified using RNeasy RNA isolation kit (QIAGEN). RNA was quantified using a NanoDrop One (ThermoFisher) and 2mg cDNA was synthesized by reverse transcription using High Capacity cDNA reverse transcription kit (Applied Biosystems). For bulk RNA-seq, mRNA libraries were prepared using QuantSeq 3' mRNA-Seq Library Prep Kit FWD with UDI (Lexogen) from 500 ng total RNA, quality checked by Fragment Analyzer, quantified by qPCR using the KAPA Library Quantification Kit for Illumina (Roche), and sequenced on the Illumina NovaSeq 6000. RNA-Seq reads were quality checked using MultiQC v1.4, which aggregated metrics from FastQC v 0.11.7 (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/), trimmed using Trim Galore version 0.6.5, and aligned in single-end mode to the mouse genome assembly (GRCm38.75) using the STAR aligner version 2.6.1d with default parameters. Mapped data were converted to gene level integer read counts (expression) using featureCounts and Ensemble GTF annotation (Mus_musculus.GRCm38.75.gtf).
identifiers.namespace: GEO
identifiers.id: GSM8728815_r1