PUBLICATION

Two types of tet-on transgenic lines for doxycycline-inducible gene expression in zebrafish rod photoreceptors and a gateway-based tet-on toolkit

Authors
Campbell, L.J., Willoughby, J.J., and Jensen, A.M.
ID
ZDB-PUB-121221-14
Date
2012
Source
PLoS One   7(12): e51270 (Journal)
Registered Authors
Jensen, Abigail, Willoughby, John
Keywords
none
MeSH Terms
  • Animals
  • Animals, Genetically Modified
  • Base Sequence
  • DNA Primers
  • Doxycycline/pharmacology*
  • Gene Expression/drug effects*
  • Immunohistochemistry
  • Plasmids
  • Polymerase Chain Reaction
  • Retinal Rod Photoreceptor Cells/metabolism*
  • Zebrafish
PubMed
23251476 Full text @ PLoS One
Abstract

The ability to control transgene expression within specific tissues is an important tool for studying the molecular and cellular mechanisms of development, physiology, and disease. We developed a Tet-On system for spatial and temporal control of transgene expression in zebrafish rod photoreceptors. We generated two transgenic lines using the Xenopus rhodopsin promoter to drive the reverse tetracycline-controlled transcriptional transactivator (rtTA), one with self-reporting GFP activity and one with an epitope tagged rtTA. The self-reporting line includes a tetracycline response element (TRE)-driven GFP and, in the presence of doxycycline, expresses GFP in larval and adult rods. A time-course of doxycycline treatment demonstrates that maximal induction of GFP expression, as determined by the number of GFP-positive rods, is reached within approximately 24 hours of drug treatment. The epitope-tagged transgenic line eliminates the need for the self-reporting GFP activity by expressing a FLAG-tagged rtTA protein. Both lines demonstrate strong induction of TRE-driven transgenes from plasmids microinjected into one-cell embryos. These results show that spatial and temporal control of transgene expression can be achieved in rod photoreceptors. Additionally, system components are constructed in Gateway compatible vectors for the rapid cloning of doxycycline-inducible transgenes and use in other areas of zebrafish research.

Genes / Markers
Figures
Expression
Phenotype
Mutation and Transgenics
Human Disease / Model Data
Sequence Targeting Reagents
Fish
Antibodies
Orthology
Engineered Foreign Genes
Mapping
Errata and Notes