Gal4 zebrafish transgenesis. How to identify germline carriers.
I suggest that you use another fish marker like you can make bicassette using two promoters. 1- the Gal4 under your specific promoter. 1-GFP under crystalline promoter that will drive the expression of the GFP to the eye only and by that way u can screen your fish by the eye color without distrusting of Gal4. Best of luck
Yes i would recommend the same. crystalline promoter for the eye is good because short. cmlc2 for the heart also works fine, although it is going to enlarge your construct significantly. We often use CFP as a label.
Alternatively just inject in a UAS:GFP line or similar. Then incross those adults to look for germline transmission. Keep in mind though that your UAS:GFP element might get mobilized by the coinjection of t TOL2 RNA. This has so far never been a problem for us, but in case you observe mosaik expression (and its a problem for you) backcross with the native UAS:GFP line.
You can use the 2A self cleaving peptide polycistronic strategy. Put a 2A peptide (e.g., T2A) between mCherry (without stop codon) and GAL4. It will make a 1:1 Cherry and GAL4 in each cell where the GAL4 is expressed.
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