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Exploring Sequence Space to Identify Binding Sites for Regulatory RNA-Binding Proteins
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Genetics
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JoVE Journal Genetics
Exploring Sequence Space to Identify Binding Sites for Regulatory RNA-Binding Proteins

Exploring Sequence Space to Identify Binding Sites for Regulatory RNA-Binding Proteins

DOI:
10.3791/59635-v

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11:34 min

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August 09, 2019

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1Department of Molecular, Cellular and Developmental Biology,University of Colorado at Boulder

Chapters

  • 00:04Title
  • 01:11Generation of DNA Random Library Pool
  • 02:14Synthesis of Pool 0 RNA
  • 04:58RNA-Protein Binding Reaction and Separation of Bound RNA
  • 08:35Reverse Transcription, PCR Amplification, and Protein Binding
  • 09:47Results: Enrichment of PTB-binding RNAs and Splice Site Switching Assay
  • 10:50Conclusion

Summary

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Sequence specificity is critical for gene regulation. Regulatory proteins that recognize specific sequences are important for gene regulation. Defining functional binding sites for such proteins is a challenging biological problem. An iterative approach for identification of a binding site for an RNA-binding protein is described here and is applicable to all RNA-binding proteins.

Tags

Keywords: Sequence SpaceBinding SitesRegulatory RNA-binding ProteinsSaturation MutagenesisProtein-binding SiteDisease DiagnosisTherapyRandom LibraryRNA PoolBinding ReactionHigh AffinitySpecific BindersPCRT7 RNA PolymeraseTranscriptionGel PurificationAutoradiography

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