High-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP, also known as CLIP-Seq) is a genome-wide means of mapping protein–RNA binding sites or RNA modification sites in vivo.[1][2][3] HITS-CLIP was originally used to generate genome-wide protein-RNA interaction maps for the neuron-specific RNA-binding protein and splicing factor NOVA1 and NOVA2;[2] since then a number of other splicing factor maps have been generated, including those for PTB,[4] RbFox2,[5] SFRS1,[6] hnRNP C,[7] and even N6-Methyladenosine (m6A) mRNA modifications.[3][8]
HITS-CLIP of the RNA-binding protein Argonaute has been performed for the identification of microRNA targets[9] by decoding microRNA-mRNA and protein-RNA interaction maps in mouse brain,[10][11] and subsequently in Caenorhabditis elegans,[12] embryonic stem cells[13] and tissue culture cells.[14]
As a novel modification of HITS-CLIP, m6A-CLIP was developed to precisely map N6-Methyladenosine(m6A) locations in mRNA by UV-crosslinking m6A antibody to the target RNA.[3][8] Recently, improved bioinformatics applied to Argonaute HITS-CLIP enables identification of binding sites with single nucleotide resolution.[15]
Similar methods
- PAR-CLIP, for identifying the binding sites of cellular RNA-binding proteins (RBPs) and microRNA-containing ribonucleoprotein complexes (miRNPs) in tissue culture cells.
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