CLIP: construction of cDNA libraries for high-throughput sequencing from RNAs cross-linked to proteins in vivo.
UV cross-linking and immunoprecipitation assay (CLIP) can identify direct interaction sites between RNA-binding proteins and RNAs in vivo, and has been used to study several proteins in tissues and cell cultures. The main challenge of the method is to specifically amplify the low amount of isolated RNA. The current protocol is optimised for efficient RNA purification and ligation of barcoded RNA adapters. High-throughput sequencing of the multiplexed cDNA library allows for a comprehensive coverage of the target sequences.
Wang
Zhen
Z
MRC-Laboratory of Molecular Biology, Hills Road, Cambridge CB20QH, UK.
Tollervey
James
J
Briese
Michael
M
Turner
Daniel
D
Ule
Jernej
J
D000818:Animals; D002478:Cells, Cultured; D003432:Cross-Linking Reagents / Q000737:chemistry; D020869:Gene Expression Profiling / Q000379:methods*; D015723:Gene Library; D047468:Immunoprecipitation; D051379:Mice; D012313:RNA / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics*; D017421:Sequence Analysis / Q000379:methods*; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D016601:RNA-Binding Proteins; D012313:RNA