Monitored eCLIP : high accuracy mapping of RNA-protein interactions.
Rémi Hocq, Janio Paternina, Quentin Alasseur, Auguste Genovesio, Hervé Le Hir.
Abstract
CLIP-seq methods provide transcriptome-wide snapshots of RNA-protein interactions in live cells. Reverse transcriptases stopping at cross-linked nucleotides sign for RNA-protein binding sites. Reading through cross-linked positions results in false binding site assignments. In the ’monitored enhanced CLIP’ (meCLIP) method, a barcoded biotinylated linker is ligated at the 5’ end of cross-linked RNA fragments to purify RNA prior to the reverse transcription. cDNAs keeping the barcode sequence correspond to reverse transcription read-throughs. Read through occurs in unpredictable proportions, representing up to one fourth of total reads. Filtering out those reads strongly improves reliability and precision in protein binding site assignment.
Nucleic Acids Research, 2018 Sep 25. doi : 10.1093/nar/gky858.
