GSM3318545: MK47_5; Homo sapiens; RNA-Seq
The total RNA was purified using RNeasy Mini Kit (QIAGEN, cat# 74106) following the manufacturer's instruction. Libraries were prepared using the Illumina protocol. The quality of the library was confirmed using Advanced Analytical Fragment Analyzer (Advanced Analytical Technologies, Heidelberg, Germany) or with an Agilent Bioanalyzer, and the concentrations of the libraries were quantified with Qubit® Fluorometric Quantitation (Life Technologies, ThermoFisher). The sequencing was performed using a HiSeq2500 Next-Generation Sequencing platform at the Finnish Functional Genomics Centre (FFGC). The quality control of raw sequencing reads was performed with FastQC, and adapters and low-quality bases were trimmed by Trimmomatic (Bolger et al., 2014). The trimmed reads were then aligned to the human reference genome GRCh37.75 (Ensembl release 75) using Tophat2 (Kim et al., 2013). Htseq-count (Anders et al., 2015) was used to calculate summarized read counts for each gene. The R/Bioconductor packages edgeR (Robinson et al., 2010) and DESeq2 (Love et al., 2014) were used to identify differentially expressed (DE) genes. Genes with a false discovery rate (FDR) <0.05 and detected using both methods were considered significantly DE.