Transcriptome analysis upon REXO2 silencing

Investigation of the role of human REXO2 in mitochondrial RNA metabolism. Overall design: The dataset corresponds to RNAseq studies and comprises experiment performed in triplicate, with the exception of REXO2 silencing sample for which duplicate was analyzed. The aim of this study was to examine the influence of REXO2 silencing on mitochondrial transcriptome. To this end we engineered two stable cell lines with the use of human HeLa Flp-In T-Rex cells as parental. First cell line inducible expressed miRNAs silencing endogenous copy of REXO2 gene while second one expressed additionally transgenic version of REXO2 which contained silent mutations causing insensitivity to miRNA. We also analyzed RNA isolated from parental HeLa Flp-In T-Rex cells. RNAseq libraries were prepared with the use of strand-specific library preparation procedures. RNAs were random fragmented and reverse transcribed using random oligomers as primers (dUTP-based protocol, see PMID: 29590189, PMID: 22609201; this pipeline enables analysis of RNAs (> ~100 nucleotides)). RNA was isolated from unfractionated cells using TRI-Reagent. Before preparation of the libraries total RNA was subjected to depletion of nuclear-encoded rRNAs (Ribo-Zero rRNA Removal Kit (Human/Mouse/Rat), Epicenter). Libraries were sequenced with the help of Illumina sequencing platform.