GSM3900313: PDX4 C420R; Homo sapiens; RNA-Seq
tissue samples were harvested with the aid of a fluorescence microscope from end stage tumors, dissecting out the GFP-positive tumor tissue, minced into ~30mg pieces, snap frozen on dry ice, and stored at -80°C. Total RNA was isolated using the RNeasy Plus Mini Kit (Qiagen, #74134) according to the manufacturer's protocol. Samples were prepared in biological replicates, n ≥ 3 per genotype. RNA integrity (RIN ≥ 8.0) was confirmed using the High Sensitivity RNA Analysis Kit (AATI, DNF-472-0500) on a 12-Capillary Fragment Analyzer. Illumina sequencing libraries with 6-bp single indices were constructed from 1 μg total RNA using the TruSeq Stranded mRNA LT kit (Illumina, RS-122-2101). The resulting library was validated using the Standard Sensitivity NGS Fragment Analysis Kit (AATI, DNF-473-0500) on a 12-Capillary Fragment Analyzer. Equal concentrations (2nM) of libraries were pooled and subjected to sequencing of approximately 20 million reads per sample using the Mid Output v2 kit (Illumina, FC-404-2001) on a Illumina NextSeq550 following the manufacturer's instructions.