RNA-seq analysis of BRAF-induced senescence

RNA-seq libraries on biological triplicate vector and BRAF^V600E-transduced Tig3 cells and senescent Tig3 cells were generated from 1 µg total RNA using the Illumina TruSeq RNA Sample Preparation kit, following the manufacturer's instruction. Libraries were single-end sequenced on the Illumina HTSeq 2000 platform. The reads were aligned to the human genome build GTC37.75 using TopHat 2.012 (Kim et al. 2013 (link)). HTSeq-count (Anders et al. 2015 (link)) was used to determine expression per gene using the default union mode. Differentially expressed genes were determined in R using edgeR (Robinson et al. 2010 (link)) and voom (Law et al. 2014 (link)) and were considered differentially expressed if they had a P-value <0.05 after Benjamini-Hochberg adjustment (Hochberg and Benjamini 1990 (link)). Relative expression of fragments per kilobase per million reads (FPKM) of each gene was calculated by normalizing the reads for each gene-by-gene length (kb) and library size (million reads).