In eukaryotes, RNA Binding Protein (RBP) exists in a wide variety of categories, and plays an important role in multiple post-transcriptional regulation processes such as RNA cleavage, transport, sequence editing, intracellular localization and translational control, hence understanding the function of RBP is of great significance in addressing human diseases. CLIP-Seq (Cross-Linking and Immunoprecipitation High Throughput Sequencing) is a revolutionary technology that reveals the interaction of RNA and RNA-binding proteins at the genome-wide level.
In recent years, CLIP-Seq has been widely used in the field of miRNA target identification. In animals, mature single-stranded miRNAs form a miRNA-induced silencing complex (RISC) with a range of proteins, and the complexes bind to the 3′ UTR region of the target mRNAs, preventing the translation of the bound mRNAs or directly degrading the target mRNAs. Through the identification of the binding site of AGO2 protein to RNA in RISC by CLIP-Seq, the false positives predicted by miRNA binding sites can be significantly reduced and the range of miRNA binding site search space can also be reduced.
Advantages
● High accuracy: real in vivo interactions from the cross-linking of living cells
●Strong specificity: UV radiation does not cause cross-linking between proteins, specificity identification of the interaction between protein and RNA are available
● Wide application: especially applicable for splicing factor RNA-binding map, miRNA target, etc.
Bioinformatics Analysis
RiboBio can provide multiple bioinformatics analysis services including bascis and advanced analysis services, welcome to contact order@ribobio.com to know more about it.
FAQs
1. What is the difference between CLIP-Seq and RIP-Seq?
Both are techniques for detecting the interaction of RNA with RNA binding proteins, and their major difference lies in the gel purification process after UV crosslinking and precipitation of complex. CLIP-Seq can mark radioactive or non-radioactive labels on the RNA ends of RNA-protein complexes and separate these complexes in SDS-PAGE, thus increasing the specificity and improving the quality of cDNA libraries.
2. What sequencing methods are recommended for CLIP-Seq and RIP-Seq?
CLIP-Seq generally uses the SE50 sequencing mode, because the RNA fragments obtained by CLIP are generally small in length. The RNA fragment obtained by RIP is relatively longer, so the SE50 or PE150 sequencing modes are both optional according to the actual quality inspection requirement and research purpose. In short, the sequencing strategy depends mainly on the length of RNA after IP.
