Assessing long-distance RNA sequence connectivity via RNA-templated DNA-DNA ligation

  1. Christian K Roy
  2. Sara Olson
  3. Brenton R Graveley
  4. Phillip D Zamore
  5. Melissa J Moore  Is a corresponding author
  1. Howard Hughes Medical Institute, University of Massachusetts Medical School, United States
  2. University of Connecticut Health Center, United States

Abstract

Many RNAs, including pre-mRNAs and long non-coding RNAs, can be thousands of nucleotides long and undergo complex post-transcriptional processing. Multiple sites of alternative splicing within a single gene exponentially increases the number of possible spliced isoforms, with most human genes currently estimated to express at least ten. To understand the mechanisms underlying these complex isoform expression patterns methods are needed that faithfully maintain long-range exon connectivity information in individual RNA molecules. Here, we describe SeqZip, a methodology that uses RNA-templated DNA-DNA ligation to retain and compress connectivity between distant sequences within single RNA molecules. Using this assay, we test proposed coordination between distant sites of alternative exon utilization in mouse Fn1 and we characterize the extraordinary exon diversity of Drosophila melanogaster Dscam1.

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Author details

  1. Christian K Roy

    RNA Therapeutics Institute, Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, United States
    Competing interests
    No competing interests declared.
  2. Sara Olson

    Institute for Systems Genomics, Department of Genetics and Developmental Biology, University of Connecticut Health Center, Farmington, United States
    Competing interests
    No competing interests declared.
  3. Brenton R Graveley

    Institute for Systems Genomics, Department of Genetics and Developmental Biology, University of Connecticut Health Center, Farmington, United States
    Competing interests
    No competing interests declared.
  4. Phillip D Zamore

    RNA Therapeutics Institute, Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, United States
    Competing interests
    Phillip D Zamore, Reviewing Editor, eLife.
  5. Melissa J Moore

    RNA Therapeutics Institute, Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, United States
    For correspondence
    melissa.moore@umassmed.edu
    Competing interests
    No competing interests declared.

Copyright

© 2015, Roy et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Christian K Roy
  2. Sara Olson
  3. Brenton R Graveley
  4. Phillip D Zamore
  5. Melissa J Moore
(2015)
Assessing long-distance RNA sequence connectivity via RNA-templated DNA-DNA ligation
eLife 4:e03700.
https://doi.org/10.7554/eLife.03700

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https://doi.org/10.7554/eLife.03700

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