Green fluorescent protein tagging Drosophila proteins at their native genomic loci with small P elements

Genetics. 2003 Nov;165(3):1433-41.

Abstract

We describe a technique to tag Drosophila proteins with GFP at their native genomic loci. This technique uses a new, small P transposable element (the Wee-P) that is composed primarily of the green fluorescent protein (GFP) sequence flanked by consensus splice acceptor and splice donor sequences. We demonstrate that insertion of the Wee-P can generate GFP fusions with native proteins. We further demonstrate that GFP-tagged proteins have correct subcellular localization and can be expressed at near-normal levels. We have used the Wee-P to tag genes with a wide variety of functions, including transmembrane proteins. A genetic analysis of 12 representative fusion lines demonstrates that loss-of-function phenotypes are not caused by the Wee-P insertion. This technology allows the generation of GFP-tagged reagents on a genome-wide scale with diverse potential applications.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Artificial Gene Fusion
  • Base Sequence
  • DNA Primers
  • DNA Transposable Elements*
  • Drosophila / genetics*
  • Drosophila Proteins / genetics*
  • Genomics*
  • Green Fluorescent Proteins
  • Luminescent Proteins / genetics
  • Recombinant Fusion Proteins / genetics
  • Subcellular Fractions / metabolism

Substances

  • DNA Primers
  • DNA Transposable Elements
  • Drosophila Proteins
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins