Multi-omics study of lung cancer in smokers from EAGLE
Frozen tumor tissue with matched blood or normal tissue samples were immediately put into 1ml of 0.2 mg/ml Proteinase K in DNA lysis buffer (10 mMTris-Cl (pH 8.0), 0.1 M EDTA (pH 8.0), and 0.5% (w/v) SDS) for 24 hrs at 56°C with shaking at 850 rpm in Thermomixer R (Eppendorf) until the tissue was completely lysed. Genomic DNA was extracted from fresh frozen tissue using the QIAmp DNA Mini Kit. Each sample was eluted in 200 μl AE buffer and DNA concentration was determined by Nanodrop spectrophotometer. All DNA samples were aliquoted and stored at −80°C until use. DNA samples are required to meet minimum mass and concentration thresholds for each assay, as well as show no evidence of contamination or profile discordance in the Identifiler assay. Samples meeting these requirements were aliquoted at the appropriate mass needed for downstream assay processing. The resulting post-capture enriched multiplexed sequencing libraries were used in cluster formation on an Illumina cBOT. Paired-end sequencing was performed by the Broad Institute using the Illumina HiSeq X system following Illumina-provided protocols for 2x150bp paired-end sequencing.