Protein Interaction Profile Sequencing PIP-Seq sequences RBP interaction sites on both unprocessed and mature RNA species in crosslinked or noncrosslinked cells (Silverman et al., 2014) (Silverman et al., 2014). In PIP-seq, both RNase-sensitive and RNase-insensitive fragments are isolated and processed separately to differentiate which sequences […]

Photoactivatable Ribonucleoside_Enhanced Crosslinking and Immunoprecipitation PAR-CLIP maps RBP sites on the target RNAs (Hafner et al., 2008). This approach is similar to HITS-CLIP and CLIP-Seq, but it uses much more efficient crosslinking to stabilize the protein-RNA complexes. The requirement to introduce a photoactivatable ribonucleoside limits […]

miRNA Trapping by RNA in Vitro Affinity Purification miTRAP identifies the target RNA sequences of miRNA species in vitro utilizing MS2 stem loops attached to the target RNA sequence as bait for the miRNA (Braun et al., 2014). MS2 is a 19 nt bacteriophage RNA […]

MicroRNA Crosslinking and Immunoprecipitation miR-CLIP was developed to identify mRNA targets of miRNA, using synthetic miRNAs (Imig et al., 2014). The capture probes in miR-CLIP are pre-miRNAs conjugated to psoralen and biotin that can be crosslinked with mRNA to isolate their targets from total RNA.First, […]

Individual Nucleotide Resolution CLIP iCLIP maps protein-RNA interactions, in a process similar to HITS-CLIP and PAR-CLIP (Konig et al., 2010). This approach includes additional steps to digest the proteins after crosslinking and to map the crosslink sites with reverse transcriptase. In iCLIP, specific crosslinked RNA-protein […]

RNA Hybrid and Individual-Nucleotide Resolution Ultraviolet Crosslinking and Immunoprecipitation hiCLIP sequences RNA duplexes bound to RBPs in vivo (Sugimoto et al., 2015). The unique cloning and linker-adapter system in hiCLIP identifies whether the RBP-bound duplex originates from the same RNA or different RNAs.Similar to CLIP […]

Enhanced Cross-linking Immunoprecipitation eCLIP maps the binding sites of RBPs on their target RNAs using a modified individual nucleotide resolution CLIP (iCLIP) protocol, improving efficiency and decreasing execution complexity (Van Nostrand et al., 2016). The hallmark of this method is the ligation of barcoded single-stranded […]

Directly Ligate Sequencing Adapters to the First-Strand cDNA DLAF is a strand-specific RNA sequencing method that avoids second-strand cDNA synthesis by directly attaching unique double-stranded adapters to the first-strand cDNA (Agarwal et al., 2015). First, mRNA is isolated and depleted of rRNA. Next, RNA fragments […]

High-Throughput Sequencing of CLIP cDNA Library Crosslinking and immunoprecipitation (CLIP), with the use of RNase T1 trimming, was first described by Ule et al (Ule et al., 2005). and later applied to high-throughput sequencing to map protein-RNA binding sites in vivo (Licatalosi et al., 2008) […]

Crosslinking, Ligation, and Sequencing of Hybrids CLASH maps RNA-RNA interactions (Kudla et al., 2011) (Helwak et al., 2013). In this method, RNA-protein complexes are UV-crosslinked and affinity-purified. RNA-RNA hybrids are ligated, isolated, and reverse-transcribed into cDNA. Deep sequencing of the cDNA provides high-resolution chimeric reads […]