Dataset features


Application: RNA-seq analysis
Number of samples: 36
Release date: Apr 24 2018
Last update date: Nov 28 2018
Access: Public
Diseases: Head and Neck Neoplasms, Neoplasms
Dataset link Integrated time course omics analysis distinguishes immediate therapeutic response from acquired resistance (RNA-Seq)

Experimental Protocol

RNA isolation and sequencing were performed for 23 samples including 2 replicates of parental SCC25 cetuximab sensitive cell line, generations of SCC25 cell lines treated with either cetuximab or PBS over weeks (each week of treatment labeled C1 to C11), and a set of 11 stable cetuximab resistant clones derived from SCC25. Sequencing was performed at the Johns Hopkins Medical Institutions (JHMI) Deep Sequencing & Microarray Core Facility. Total RNA was isolated from a total of 1x106 cells using the AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany) following manufacturer’s instructions. The RNA concentration was determined by the spectrophotometer Nanodrop (Thermo Fisher Scientific, Waltham, MA) and quality was assessed using the 2100 Bioanalyzer (Agilent, Santa Clara, CA) system. An RNA Integrity Number (RIN) of 7.0 was considered as the minimum to be used in the subsequent steps for RNAseq. Library preparation was performed using the TrueSeq Stranded Total RNAseq Poly A1 Gold Kit (Illumina, San Diego, CA), according to manufacturer’s recommendations, followed by mRNA enrichment using poly(A) enrichment for ribosomal RNA (rRNA) removal. Sequencing was performed using the HiSeq platform (Illumina) for 2X100bp sequencing. Reads were aligned to hg19 with MapSplice16 and gene expression counts were quantified with RSEM17. Gene counts were upper-quartile normalized and log transformed for analysis following the RSEM v2 pipeline used to normalize TCGA RNA-seq data.










Elana Fertig