CTRL15
Total RNA (500 ng) was converted to cDNA libraries using TruSeq Stranded Total RNA with Ribo-Zero kit (Illumina) according to manufacturer's protocol. Libraries were evaluated qualitatively on Bioanalyzer 2100 using High Sensitivity DNA Kit and quantitatively on CFX96 Real-Time PCR system (Bio-Rad) using KAPA Library Quantification Kit (Kapa Biosystems), and then sequenced 2x76bp on HiSeq2500 Illumina platform. At least 25 M reads per sample were obtained with mean quality score (>=Q30)>94%.   Bioinformatics:  Raw reads in FASTQ files were subjected to trimming of low quality 5’ and 3’ ends, adapters and indexes of Illumina by Trimmomatic software (Bolger et al., 2014). Next, rRNA was removed from the samples using Bowtie2 software (Langmead et al., 2009). Alignment of remaining the RNA-seq reads to human genome GRCh38 release 76 (NCBI) was performed by a STAR splice junction mapper (Dobin et al., 2013). Then, the samples were filtered to remove the reads of low mapping quality (>=50) using Bamtools software (Barnett et al., 2011). PCR duplicates were removed with http://broadinstitute.github.io/picard/ and the samples were sorted using Bamtools (Barnett et al., 2011).