posted 10 July 2014 by Fillip
In figure S5 of our paper we report the generation of a GFP knock-in allele of wingless (wg) using our CRISPR/Cas system. In panel C we show a low-magnification image of imaginal discs from a wg:GFP animal. The image is a single confocal section and as a result not the entire expression domain of wg:GFP is visible. For those of you interested in this allele we provide here a movie of a three-dimensional Z-stack through a wg:GFP wing imaginal disc that shows the entire expression pattern.
The disc was dissected in PBS, fixed in 4% paraformaldehyde for 25 min, washed in PBT and mounted in Vectarshield mounting medium. Samples were imaged on a Zeiss 780 confocal micoscope with a 20x/0.5 NA air objective.
The wg:GFP expression pattern seems to recapitulate all aspects of the pattern observed when untagged endogenous Wg is visualised using a specific antibody (see for example: Figure 1G in Zeng and Verheyen, 2004 DOI: 10.1242/dev.01177 ). A more important question is of course whether wg:GFP can fully substitute for untagged wg. We are currently performing crosses to find out and will update this post once we know the answer.
Update: 15 September 2014
The wg:GFP allele is indeed homozygous viable without any obvious phenotypes. We currently backcross this allele into a wildtype background and will then deposit it at the Bloomington stock center.