GoldCLIP： Gel-omitted Ligation-dependent CLIP  

作　　者：Jiaqi Gu Ming Wang Yang Yang Ding Qiu Yiqun Zhang Jinbiao Ma Yu Zhou Gregory J. Hannon Yang Yu 

机构地区：[1]State Key Laboratory of Genetic Engineering, Department of Biochemistry, School of Life Sciences, Fudan University Shanghai 200438, China [2]CAS Key Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China [3]Hubei Key Laboratory of Cell Homeostasis, College of Life Sciences, Wuhan University, Wuhan 430072, China [4]Cancer Research UK, Li Ka Shing Centre, University of Cambridge, Cambridge CB2 ORE, United Kingdom

出　　处：《Genomics, Proteomics & Bioinformatics》2018年第2期136-143,共8页基因组蛋白质组与生物信息学报（英文版）

基　　金：supported in part by grants from the Ministry of Science and Technology of China(Grant No.2017YFA0504200 to YY,Grant Nos.2012CB910502 and 2011CB966304 to JM);the National Natural Science Foundation of China(Grant Nos.91640105 and 31770875 to YY,Grant No.31230041 to JM,and Grant Nos.91640115 and 31670827 to YZ)

摘　　要：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 nonisotopic 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.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 nonisotopic 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.

关 键 词：RNA binding protein UV crosslinking CLIP HALOTAG PTB 

分 类 号：Q75[生物学—分子生物学]