Ribosome profiling of WT E14, Dgcr8_KO, Dorsha_KO, Dicer_KO and Ago1&2_Double_KO mouse embryonic stem cells

miRNAs are short regulatory single stranded RNA sequences that upon complementary binding to mRNAs lead to the inhibition or degradation of their targets. This regulatory mechanisms has been shown to play crucial roles throughout the whole life cycle of animals and plants as well as in disease. While a plethora of methods exist to predict targets of miRNA, which suggest that up to 80% of the genome is miRNA regulated, it has recently been reported that many of these predictions are false positives, cell type specific or represent non-functional binding. In order to identify the subset of real functional miRNAs and their targets, we established miRNA pathway mutants in mouse embryonic stem cells (mESC), allowing the dissection of canonical and non-canonical functions of pathway members. Additional data integration of downstream regulatory layers (CLIP-seq, ribosome profiling and MS) enabled us to follow and track down real functional miRNA-gene interactions, which reduced the miRNA genome regulation to approximately 1%. Overall design: Purpose: The goals of this study was to obtain the ribosome binding profile of WT and RNAi mutant mESCs. Ribosome profiling of different RNAi mutant mouse embryonic stem cells (E14) were generated by deep sequencing, in duplicate, using Illumina HiSeq2500.