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

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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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Christian K Roy

RNA Therapeutics Institute, Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, United States

Competing interests
No competing interests declared.
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.
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.
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.
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.

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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.