Beadstation 500 beadarray reader

Mouse microarrays were performed using Illumina^® MouseWG-6 v2.0 Expression BeadChips (Illumina). Samples were labeled and hybridized using Illumina^® TotalPrep^™ kit (Ambion) and then arrays were scanned using Illumina^® Beadstation 500 BeadArray reader and data acquisitioned with BeadStudio (Illumina^®). R 2.15.1 (http://www.R-project.org/) and tools in Bioconductor (http://www.bioconductor.org/) were used for all analyses unless otherwise stated. More detailed methodology, including flowchart and Sweave report, for microarray and survival analyses in Supplementary Methods.

Gene expression differences were determined between 40 NSCLCs (the cells that had IC₅₀≤1.55 μM and those cells that had IC₅₀ ≥10 μM) by microarray analysis. Briefly, approximately 5μg of total cellular RNA was isolated from each cell line and reverse transcribed into cDNA using standard techniques. The cDNA was indirectly labeled with a fluorescent probe using a two-step hybridization and labeling protocol where the gene chip (Illumina Human WG-6 V3, Cat No: BD-101–0203, BD-101–0603) was hybridized to cDNA overnight, washed stringently, and then post-stained with fluorescent dendrimers. After hybridization and washes, the gene chip was scanned using Illumina TotalPrep Kit (Ambion, Waltham, MA) and then arrays were scanned using Illumina Beadstation 500 BeadArray reader and data acquisitioned with Illumina BeadStudio for visualization and data mining.
Raw and processed data are available on GEO (accession GSE32036), http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?token=pfiphqkackiyubo&acc=GSE32036). Raw data was processed using default parameters of the MBCB package in R/Bioconductor. Statistically significant genes were determined using unpaired t-tests with multiple testing correction via the Bonferroni method (p<0.01).