RNA-sequencing of LCL and B cells

Global gene expression profiling using RNAseq was carried out for n = 5 LCL and CD19+ B cells (matching donors). Total RNA was first isolated using the RNeasy Mini Kit (Qiagen) before RNAseq library preparation using the TruSeqV2 Library Preparation kit (Illumina). The indexed libraries were pooled 10 plex, and 50 bp single-end reads were sequenced on the HiSeq 2500 (Illumina). Reads were assessed for quality using FastQC, aligned to hg19 using TopHat2 [22 (link)], and summarised to RPKM gene level expression using SAMmate [23 (link)]. An average of 18 million mapped reads was obtained for each sample at an overall alignment rate of 80.5%. Differentially expressed genes were calculated using EdgeR [24 (link)]. A cut-off of 1% false discovery rate was selected to select differentially expressed genes, see Additional file 1: Table S1. The raw and processed sequencing data generated have been submitted to the NCBI Gene Expression Omnibus (GEO; http://www.ncbi.nlm.nih.gov/geo/) under accession number GSE126379 [25 ].