In vivo study of gene expression with an enhanced dual-color fluorescent transcriptional timer
- Harvard Medical School, United States
- Chongqing University of Arts and Sciences, China
- Tufts University, United States
Abstract
Fluorescent transcriptional reporters are widely used as signaling reporters and biomarkers to monitor pathway activities and determine cell type identities. However, a large amount of dynamic information is lost due to the long half-life of the fluorescent proteins. To better detect dynamics, fluorescent transcriptional reporters can be destabilized to shorten their half-lives. However, applications of this approach in vivo are limited due to significant reduction of signal intensities. To overcome this limitation, we enhanced translation of a destabilized fluorescent protein and demonstrate the advantages of this approach by characterizing spatio-temporal changes of transcriptional activities in Drosophila. In addition, by combining a fast-folding destabilized fluorescent protein and a slow-folding long-lived fluorescent protein, we generated a dual-color transcriptional timer that provides spatio-temporal information about signaling pathway activities. Finally, we demonstrate the use of this transcriptional timer to identify new genes with dynamic expression patterns.
Data availability
All essential data are provided in the supplementary materials. All the reagents created by this study (plasmids and transgenic flies) will be donated to public domains including Addgene and Bloomington Stock Center.
Article and author information
Author details
Funding
National Institute of General Medical Sciences
- Norbert Perrimon
Damon Runyon Cancer Research Foundation
- Li He
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, He et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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In vivo study of gene expression with an enhanced dual-color fluorescent transcriptional timer
eLife 8:e46181.
https://doi.org/10.7554/eLife.46181
