Abstract
Techniques for conditional gene or protein expression are important tools in developmental biology and in the analysis of physiology and disease. On the protein level, the tunable and reversible expression of proteins can be achieved by the fusion of the protein of interest to a destabilizing domain (DD). In the absence of its specific ligand (Shield-1), the protein is degraded by the proteasome. The DD-Shield system has proven to be an excellent tool to regulate the expression of proteins of interests in mammalian systems but has not been applied in teleosts like the medaka. We present the application of the DD-Shield technique in transgenic medaka and show the ubiquitous conditional expression throughout life. Shield-1 administration to the water leads to concentration-dependent induction of a YFP reporter gene in various organs and in spermatogonia at the cellular level.
Highlights
The analysis of gene function in development and in adult organisms requires the controlled expression of proteins of interest
We present the application of the destabilizing domain (DD)-Shield technique in transgenic medaka and show the ubiquitous conditional expression throughout life
HEK293T cells were transfected with pDS-actin-DD-YFP, cultured for 24 hours and treated with Shield-1
Summary
The analysis of gene function in development and in adult organisms requires the controlled expression of proteins of interest. Many genes have pleiotropic functions during embryogenesis and adulthood, the expression of a transgene may cause embryonic phenotypes that impede the analysis of later developmental stages. Several conditional expression systems have been developed to avoid embryonic phenotypes and to restrict the expression temporally and spatially [1]. Two transgenes are used to control the expression of the gene of interest. The effector transgene defines the time and tissue of expression, activating or repressing the target transgene. In combination with transgenic methods to express fluorescent proteins in specific cell types, the observation of environmental or genetic effects is possible at the level of tissues or cells [2]
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