Authors:
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
Summary:
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.
Source:
Nature Methods; 8, 506-512 (05/08/11)