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Related Concept Videos

RNA-seq03:21

RNA-seq

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RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
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Related Experiment Video

Updated: Mar 12, 2026

PAR-CliP - A Method to Identify Transcriptome-wide the Binding Sites of RNA Binding Proteins
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PAR-CliP - A Method to Identify Transcriptome-wide the Binding Sites of RNA Binding Proteins

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DO-RIP-seq to quantify RNA binding sites transcriptome-wide.

Cindo O Nicholson1, Matthew B Friedersdorf1, Laura S Bisogno2

  • 1Department of Molecular Genetics & Microbiology, USA; Duke University Medical Center, Durham, NC 27710, USA.

Methods (San Diego, Calif.)
|November 15, 2016
PubMed
Summary
This summary is machine-generated.

Digestion optimized RNA immunoprecipitation with deep sequencing (DO-RIP-seq) quantifies RNA binding protein interactions at both whole transcript and binding site levels. This method provides continuous metrics for dynamic biological processes.

Keywords:
DO-RIP-seqQuantitationRNA-binding proteinsRNA-binding sitesWhole-transcript targets

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Post-transcriptional regulation relies on dynamic protein-RNA interactions.
  • Existing methods quantify either whole transcript binding or specific binding sites, but not both.
  • A need exists for a method quantifying RNA binding protein (RBP) interactions at multiple levels.

Purpose of the Study:

  • To introduce digestion optimized RNA immunoprecipitation with deep sequencing (DO-RIP-seq).
  • To demonstrate DO-RIP-seq's capability to quantify RBP binding at whole transcript (RIP-Seq-Like or RSL) and binding site (BSL) levels.
  • To validate the method using the RBP HuR/ELAVL1.

Main Methods:

  • DO-RIP-seq involves partial RNA digestion of cell lysates using nucleases under optimized conditions.
  • Immunoprecipitation captures digested RNA-protein complexes, assessing background interactions.
  • Deep sequencing of bound RNA fragments enables quantitative analysis.

Main Results:

  • DO-RIP-seq provides continuous metrics for both RSL and BSL binding events.
  • The method yields two types of enrichment scores: one for RSL and one for BSL.
  • Extensive read coverage allows seamless integration of binding site and whole transcript information.

Conclusions:

  • DO-RIP-seq is a novel method for quantifying RBP binding events.
  • It enables simultaneous quantification at whole transcript and binding site levels.
  • DO-RIP-seq is valuable for studying RBP binding during dynamic biological processes.