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Transcriptome-Wide Profiling of Protein-RNA Interactions by Cross-Linking and Immunoprecipitation Mediated by FLAG-Biotin Tandem Purification
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JoVE Journal Genetik
Transcriptome-Wide Profiling of Protein-RNA Interactions by Cross-Linking and Immunoprecipitation Mediated by FLAG-Biotin Tandem Purification

Transcriptome-Wide Profiling of Protein-RNA Interactions by Cross-Linking and Immunoprecipitation Mediated by FLAG-Biotin Tandem Purification

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08:46 min

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May 18, 2020

DOI:

10.3791/60730-v

DOI:
10.3791/60730-v

•

08:46 min
May 18, 2020

•

3 Views
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1Tsinghua-Peking Center for Life Sciences, School of Medicine and School of Life Sciences,Tsinghua University, 2Department of Molecular Biology,University of Texas Southwestern Medical Center

Summary

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Here we present a modified CLIP-seq protocol called FbioCLIP-seq with FLAG-biotin tandem purification to determine the RNA targets of RNA-binding proteins (RBPs) in mammalian cells.

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Protein-RNA InteractionsCLIP-seqFLAG-BiotinTranscriptome-wide ProfilingRBPsMammalian CellsDirect Protein-bound RNAsIsotope-freeSDS-PAGEMembrane TransferMouse Embryonic Stem CellsUV Cross-linkingCell LysisDNase I TreatmentFLAG BeadsFLAG Peptide Elution

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Bi, X., Zhang, X., Shen, X. Transcriptome-Wide Profiling of Protein-RNA Interactions by Cross-Linking and Immunoprecipitation Mediated by FLAG-Biotin Tandem Purification. J. Vis. Exp. (159), e60730, doi:10.3791/60730 (2020).

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