Hello,
I am working on transient expression of GFP (in leaves) in both Nicotiana benthamiana and a non-model plant.
Recombinant GFP is well detected in WB with the specific primary antibodies, for both N. benthamiana and the non-model plant.
My problem is that GFP specific fluorescence is visible in N. bentha, but not in the non-model plant.
Has anyone ever encountered this problem?
Thank you for sharing your experience
Maybe you should check the signal at different time intervals.
Hi, sometimes it is difficult to detect GFP with the microscopes, specially if the transduction efficiency is lower. Can you flow the cells from the plant in a flow cytometer? That would also give you a measure of the GFP protein in your plant. Hope it helps!
Full disclosure: no experience with plants, but plenty with GFP. By Western blot, are levels truly comparable (when normalized to total protein)? If yes, then it is indeed rather odd. How certain are you of your 'specific' Ab? Do you have an untouched control?
Less likely but still possible, could mean that GFP hasn`t yet folded/matured (pretty fast usually, at least in animal cells, 15 min or so, but could be somehow incomplete in your 'non-model' plant). Are you using the same construct for both? If not, it may have a freak mutation. Either way, if not done so yet, would perform the experiments side-by-side to limit possible interexperimental variations.
Bonne chance!
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