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Reactivation Assay to Identify Direct Targets of the Nonsense-Mediated mRNA Decay Pathway in Drosophila

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mRNA Decay

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1720))

Abstract

Transcriptome analysis provides a snapshot of cellular gene expression and is used to determine how cells and organisms respond to genetic or environmental changes. Identifying the transcripts whose expression levels are regulated directly by the manipulation being examined from those whose expression changes as a secondary cause from the primary changes requires additional analyses. Here we present a technique used to distinguish direct targets of the nonsense-mediated mRNA decay (NMD) pathway in Drosophila from secondary gene expression effects caused by loss of this pathway. This technique uses pulsed reexpression of an essential NMD gene in Drosophila lacking this NMD factor, followed by analysis of the transcriptome over time. In this way, RNAs with a rapid reduction in expression upon reactivation of NMD activity, corresponding to primary NMD targets, can be identified. This technique could potentially be modified to identify direct targets of other mRNA decay mechanisms in Drosophila or other organisms.

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Author information

Authors and Affiliations

  1. Department of Human Genetics, University of Utah, Salt Lake City, UT, USA

    Jonathan O. Nelson & Mark M. Metzstein

  2. Life Sciences Institute, University of Michigan, Ann Arbor, MI, USA

    Jonathan O. Nelson

Authors
  1. Jonathan O. Nelson
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  2. Mark M. Metzstein
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Corresponding author

Correspondence to Mark M. Metzstein .

Editor information

Editors and Affiliations

  1. Murdoch Children’s Research Institute, The Royal Children’s Hospital, Parkville, Victoria, Australia

    Shireen R. Lamandé

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Nelson, J.O., Metzstein, M.M. (2018). Reactivation Assay to Identify Direct Targets of the Nonsense-Mediated mRNA Decay Pathway in Drosophila. In: Lamandé, S. (eds) mRNA Decay. Methods in Molecular Biology, vol 1720. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7540-2_15

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  • DOI: https://doi.org/10.1007/978-1-4939-7540-2_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7539-6

  • Online ISBN: 978-1-4939-7540-2

  • eBook Packages: Springer Protocols

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