The study of RNA chemical modifications is currently one of the most rapid-growing fields. Many types of RNA modifications in diverse RNA species have been shown to play versatile roles in a wide array of cellular processes. These modifications are installed and erased by writer and eraser enzymes, respectively. Additionally, RNA chemical modifications have downstream biological effects through either influencing changes in the chemistry or structure of RNA molecules or through recognition of the modification; these functions are primarily executed by the modification reader proteins. Reader proteins may bind to the modification site and cause a downstream signal cascade. One of the essential tools for studying erasers, writers, and readers is cross-linking immunoprecipitation followed by high-throughput sequencing (CLIP-seq). This method can detect the sites on endogenous RNAs bound by RNA-binding proteins or RNA modifying enzymes. Essentially, this strategy allows for snapshots of the epitranscriptome and molecular events occurring within the cell. In this article, we go through in detail the various steps involved in CLIP-seq.
Keywords: CLIP-seq; RNA binding protein; RNA modification; RNA modifying enzymes; Sequencing method.
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