Welcome to the new version of CaltechAUTHORS. Login is currently restricted to library staff. If you notice any issues, please email coda@library.caltech.edu
Published November 1, 2011 | Published + Supplemental Material
Journal Article Open

A versatile gene trap to visualize and interrogate the function of the vertebrate proteome

Abstract

We report a multifunctional gene-trapping approach, which generates full-length Citrine fusions with endogenous proteins and conditional mutants from a single integration event of the FlipTrap vector. We identified 170 FlipTrap zebrafish lines with diverse tissue-specific expression patterns and distinct subcellular localizations of fusion proteins generated by the integration of an internal citrine exon. Cre-mediated conditional mutagenesis is enabled by heterotypic lox sites that delete Citrine and "flip" in its place mCherry with a polyadenylation signal, resulting in a truncated fusion protein. Inducing recombination with Cerulean-Cre results in fusion proteins that often mislocalize, exhibit mutant phenotypes, and dramatically knock down wild-type transcript levels. FRT sites in the vector enable targeted genetic manipulation of the trapped loci in the presence of Flp recombinase. Thus, the FlipTrap captures the functional proteome, enabling the visualization of full-length fluorescent fusion proteins and interrogation of function by conditional mutagenesis and targeted genetic manipulation.

Additional Information

© 2011 by Cold Spring Harbor Laboratory Press. The Authors acknowledge that six months after the full-issue publication date, the Article will be distributed under a Creative Commons CC-BY-NC License (Attribution-NonCommercial 4.0 International License, http://creativecommons.org/licenses/by-nc/4.0/). Received July 7, 2011; revised version accepted September 16, 2011. We thank P. Pantazis for comments on the manuscript, and members of the Fraser laboratory for helpful discussions. We are grateful to LeighAnn Fletcher for fish care. The T2KXIGd-in and pMDS-eGFP plasmids were provided by K. Kawakami and S. Parinov, respectively. This work was supported by NHGRI Center of Excellence in Genomic Science grant P50HG004071.

Attached Files

Published - Genes_Dev.-2011-Trinh-2306-20.pdf

Published - Trinh2011p16329Genes_Dev.pdf

Supplemental Material - Binder2.pdf

Supplemental Material - ResourcePaperSI_text.pdf

Files

Binder2.pdf
Files (39.8 MB)
Name Size Download all
md5:152778f22367096247dbbfc6a5545fb5
36.9 MB Preview Download
md5:158c600a2fda3ca3a03c465f45224efe
1.4 MB Preview Download
md5:5a28957dc1ca254a34e2ba80726b69c2
1.4 MB Preview Download
md5:d5bab80a4ebc2b807b595d4ac388f27e
24.0 kB Preview Download

Additional details

Created:
August 22, 2023
Modified:
October 24, 2023