Cells were cultured in RPMI-1640, supplemented with 10% FBS and 1X Penicillin/Streptomycin. Xenografts were grown by subcutaneous injection of two million trypsin-dissociated tumor cells into non-obese diabetic/severe combined immunodeficient mice.
Extracted molecule
total RNA
Extraction protocol
Total RNA was isolated from confluent cell lines or fresh-frozen xenograft tissues using Trizol reagent (Life Technologies, Carlsbad, CA, USA) according to the manufacturer's instructions, followed by Dnase I treatment (Life Technologies) and purification using the Qiagen RNeasey kit (Qiagen, Venlo, Netherlands). RNA quantity and quality was assessed using the Qubit Fluorometer (Life Technologies), NanoDrop 1000 (Nanodrop Technologies, Wilmington, DE, USA), and Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA).
Label
biotin
Label protocol
Microarray labeling, hybridization and scanning was performed according to the manufacturer's recommendations by the University Health Networks (UHN) Microarray Centre (Toronto, Ontario, Canada).
Hybridization protocol
Microarray labeling, hybridization and scanning was performed according to the manufacturer's recommendations by the University Health Networks (UHN) Microarray Centre (Toronto, Ontario, Canada).
Scan protocol
Microarray labeling, hybridization and scanning was performed according to the manufacturer's recommendations by the University Health Networks (UHN) Microarray Centre (Toronto, Ontario, Canada).
Data processing
Image files were quantified in BeadStudio Version 3.3.8 (Illumina) and raw data output were loaded into the R statistical environment (v2.14.0) using the lumi package (v2.6.0) implemented in the Bioconductor libraries. The probes were re-annotated against miRBase v16, log2 transformed and normalized using the Robust Spline Normalization (RSN) algorithm. No background correction was performed.
