Journal
GENOMICS PROTEOMICS & BIOINFORMATICS
Volume 16, Issue 2, Pages 136-143Publisher
ELSEVIER
DOI: 10.1016/j.gpb.2018.04.003
Keywords
RNA binding protein; UV crosslinking; CLIP; HaloTag; PTB
Categories
Funding
- Ministry of Science and Technology of China [2017YFA0504200, 2012CB910502, 2011CB966304]
- National Natural Science Foundation of China [91640105, 31770875, 31230041, 91640115, 31670827]
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Protein-RNA interaction networks are essential to understand gene regulation control. Identifying binding sites of RNA-binding proteins (RBPs) by the UV-crosslinking and immunoprecipitation (CLIP) represents one of the most powerful methods to map protein-RNA interactions in vivo. However, the traditional CLIP protocol is technically challenging, which requires radioactive labeling and suffers from material loss during PAGE-membrane transfer procedures. Here we introduce a super-efficient CLIP method (GoldCLIP) that omits all gel purification steps. This non-isotopic method allows us to perform highly reproducible CLIP experiments with polypyrimidine tract-binding protein (PTB), a classical RBP in human cell lines. In principle, our method guarantees sequencing library constructions, providing the protein of interest can be successfully crosslinked to RNAs in living cells. GoldCLIP is readily applicable to diverse proteins to uncover their endogenous RNA targets.
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