Mayo Clinic GBM PDX
RNAseq was performed using RNA isolated from a single flash frozen orthotopic tumor specimen. For each PDX line, a single tumor-bearing brain from a moribund mouse was serial sectioned and flash frozen in Tissue-Tek Optimal Cutting Temperature (OCT) medium (Sakura FineTek USA Inc., Torrance, CA), and tumor was scraped from multiple serial tumor sections using a hematoxylin and eosin (H&E) stained slide as a guide to identify dense tumor. At Columbia University, TGEN and Mayo Clinic, total RNA (50 ng) of each sample was used to generate whole transcriptome libraries for RNA sequencing using Illuminas TruSeq RNA Sample Prep Kit V2 following the manufacturers protocol. Total RNA was used to generate double stranded cDNA and subsequently amplified using Nugens SPIA linear amplification process. Amplified products were cleaned using Qiagens AIA-quick PCR Purification Kit and quantitated using Invitrogens Quant-iT Picogreen. Amplified cDNA (1 g) was fragmented to a target size of 300 bp. Illunimas TruSeq DNA Sample Preparation Kit was used to prepare libraries from 1 g amplified cDNA.