Efficient activation of gene expression using a heat-shock inducible Gal4/Vp16-UAS system in medaka.
BACKGROUND
Genetic interference by DNA, mRNA or morpholino injection is a widely used approach to study gene function in developmental biology. However, the lack of temporal control over the activity of interfering molecules often hampers investigation of gene function required during later stages of embryogenesis. To elucidate the roles of genes during embryogenesis a precise temporal control of transgene expression levels in the developing organism is on demand.


RESULTS
We have generated a transgenic Gal4/Vp16 activator line that is heat-shock inducible, thereby providing a tool to drive the expression of specific effector genes via Gal4/Vp16. Merging the Gal4/Vp16-UAS system with the I-SceI meganuclease and the Sleeping Beauty transposon system allows inducible gene expression in an entirely uniform manner without the need to generate transgenic effector lines. Combination of this system with fluorescent protein reporters furthermore facilitates the direct visualization of transgene expressing cells in live embryos.


CONCLUSIONS
The combinatorial properties of this expression system provide a powerful tool for the analysis of gene function during embryonic and larval development in fish by ectopic expression of gene products.
Grabher
Clemens
C
Developmental Biology Program, European Molecular Biology Laboratory, 69117-Heidelberg, Germany. grabher@embl.de <grabher@embl.de>
Wittbrodt
Joachim
J
D000818:Animals; D004625:Embryo, Nonmammalian / Q000737:chemistry / Q000378:metabolism; D029941:Fish Proteins / Q000235:genetics; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D009990:Oryzias / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000502:physiology; D012313:RNA / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015534:Trans-Activators / Q000235:genetics*; D014162:Transfection / Q000379:methods; D015027:Zebrafish / Q000235:genetics; D029961:Zebrafish Proteins / Q000235:genetics
D029941:Fish Proteins; C070633:Gal-VP16; D018840:HSP70 Heat-Shock Proteins; D015534:Trans-Activators; D029961:Zebrafish Proteins; D012313:RNA