Insect genetic modification technologies rely heavily on the use of fluorescent proteins as genetic markers, expression reporters etc. Colors vary, as do fluorescent outputs. Shearin et al. (2014) describe their efforts to increase signals from two popular fluorescent proteins, EGFP and mCherry, by creating multimers of their ORFs in a single transcription unit. They have linked six copies of EGFP and mCherry and report significant signal gains from each ‘super protein’.
There has been a lot of effort to engineer fluorescent proteins to have different colors and enhanced fluorescent output. tdTomato, for example, is a dimer of Tomato fluorescent protein’s ORF that has over 300% the fluorescent output of EGFP.
Another way to increase signals is to simply express more of each fluorescent protein. This was the approach of Shearin et al. (2014) but they did this by creating tandem fusions on EGFP and mCherry resulting in ~170 kDa proteins as predicted.
Cells expressing these large tandem fusions were brighter than the same cells expressing monomeric forms of the proteins. These bright forms are particularly useful when trying to observe either small numbers of cells or cells in which the promoter driving expression is relatively weak.
Expression of the hexameric mCherry reporter gene could be observed with the naked eye in cases where a promoter widely expressed in the animal was used to drive expression, resulting in red larvae or red ganglia.
Not surprisingly, the authors did detect some fitness costs in some cases where there was ubiquitous expression, but when expression was restricted to a limited number of cells, no lethality was observed.
The authors show how these super-reports pair well with binary expression systems such as Gal4/UAS but their utility could be exploited by direct expression studies as well.
These tools are worth keeping in mind by those who use fluorescent proteins routinely in transgenic insect studies.
Hexameric GFP and mCherry Reporters for the Drosophila GAL4, Q, and LexA Transcription Systems.
Harold K. Shearin, Ian S. Macdonald, Laura P. Spector and R. Steven Stowers
Genetics April 1, 2014 vol. 196 no. 4 951-960