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CARIP-Seq and ChIP-Seq: Methods to Identify Chromatin-Associated RNAs and Protein-DNA Interactions in Embryonic Stem Cells
Published on: May 25, 2018
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Mapping RNA-chromatin interactions by sequencing with iMARGI.
Weixin Wu1, Zhangming Yan1,2, Tri C Nguyen1
1Department of Bioengineering, University of California, San Diego, La Jolla, CA, USA.
Nature Protocols
|October 18, 2019
Summary
The iMARGI technique efficiently maps RNA-genome interactions and discovers chromatin-associated RNAs (caRNAs) with reduced cell input and experimental time. This method reveals genomic interaction loci for caRNAs, advancing transcriptional regulation studies.
Area of Science:
- Molecular Biology
- Genomics
- Epigenetics
Background:
- RNA-chromatin interactions are crucial for gene and transposable element transcriptional regulation.
- Current methods for analyzing chromatin-associated RNAs (caRNAs) are often limited to single RNA analysis.
Purpose of the Study:
- To introduce the iMARGI (in situ mapping of RNA-genome interactome) technique for discovering caRNAs and their genomic interaction sites.
- To provide a standardized bioinformatic tool, iMARGI-Docker, for processing sequencing data.
Main Methods:
- iMARGI involves in situ crosslinking and genome fragmentation.
- Proximal RNA-DNA pairs are converted into chimeric RNA-linker-DNA sequences.
- Chimeric sequences are prepared for paired-end sequencing, with data analyzed by iMARGI-Docker.
Main Results:
- iMARGI requires a 100-fold reduction in input cells (3-5 million) compared to MARGI.
- The experimental time is significantly reduced, with library construction taking 8 days and data processing 1 day.
- The technique successfully decodes caRNA-DNA interactions.
Conclusions:
- iMARGI offers an improved method for mapping RNA-genome interactions and identifying caRNAs.
- The technique streamlines the discovery of caRNA-DNA interactions, facilitating research in transcriptional regulation.
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