In vivo protein trapping produces a functional expression codex of the vertebrate proteome

Karl J. Clark, Darius Balciunas, Hans Martin Pogoda, Yonghe Ding, Stephanie E. Westcot, Victoria M. Bedell, Tammy M. Greenwood, Mark D. Urban, Kimberly J. Skuster, Andrew M. Petzold, Jun Ni, Aubrey L. Nielsen, Ashok Patowary, Vinod Scaria, Sridhar Sivasubbu, Xiaolei Xu, Matthias Hammerschmidt, Stephen C. Ekker

Research output: Contribution to journalArticle

108 Scopus citations

Abstract

We describe a conditional in vivo protein-trap mutagenesis system that reveals spatiotemporal protein expression dynamics and can be used to assess gene function in the vertebrate Danio rerio. Integration of pGBT-RP2.1 (RP2), a gene-breaking transposon containing a protein trap, efficiently disrupts gene expression with >97% knockdown of normal transcript amounts and simultaneously reports protein expression for each locus. The mutant alleles are revertible in somatic tissues via Cre recombinase or splice-site-blocking morpholinos and are thus to our knowledge the first systematic conditional mutant alleles outside the mouse model. We report a collection of 350 zebrafish lines that include diverse molecular loci. RP2 integrations reveal the complexity of genomic architecture and gene function in a living organism and can provide information on protein subcellular localization. The RP2 mutagenesis system is a step toward a unified 'codex' of protein expression and direct functional annotation of the vertebrate genome.

Original languageEnglish (US)
Pages (from-to)506-512
Number of pages7
JournalNature Methods
Volume8
Issue number6
DOIs
StatePublished - Jun 2011

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Clark, K. J., Balciunas, D., Pogoda, H. M., Ding, Y., Westcot, S. E., Bedell, V. M., Greenwood, T. M., Urban, M. D., Skuster, K. J., Petzold, A. M., Ni, J., Nielsen, A. L., Patowary, A., Scaria, V., Sivasubbu, S., Xu, X., Hammerschmidt, M., & Ekker, S. C. (2011). In vivo protein trapping produces a functional expression codex of the vertebrate proteome. Nature Methods, 8(6), 506-512. https://doi.org/10.1038/nmeth.1606