title: GSM3764492: iCLIP_SRSF4_1; Mus musculus; RIP-Seq
library_construction_protocol: Mice were sacrificed with carbon dioxide, hearts were extracted, perfused with PBS and tissue was inmediatly freezed in liquid nitrogen. Total RNA was purified using RNeasy Mini Kit (Qiagen). Briefly, crosslinked RNA was digested to lengths of 80-200 nucleotides using RNaseI (Ambion) and RNA-protein complexes were immunopurified using Protein G Dynabeads® (ThermoFisher Scientific) coupled with a goat anti-GFP antibody (D. Drechsel, MPI-CBG, Dresden). Isolated and purified RNA fragments were ligated to pre-adenylated DNA 3' adapters (IDT) and reverse-transcribed using barcoded RT-primers and SuperscriptIV (ThermoFisher Scientific). cDNA fragments were size-selected, circularized using CircLigaseTM (Epicentre) and re-linearized by BamHI (NEB). The final cDNA libraries now containing 5' and 3' adapters were amplified using AccuPrime (ThermoFisher Scientific)