In vivo imaging of plants became a common research tool all over the world for gene expression, studies of circadian rhythm, etc.
Plants are transfected either with Luciferase or photoproteins like GFP (Fig. 1), YFP or dsRED.


Epi-illumination (from above) is easy to perform with plants not kept in Petri dishes (Fig.1).
However, epi-illumination of Petri dishes causes problems due to reflection of the illumination devices seen on the surface of Petri dishes.
Trans-illumination (from below) could also excitate transfected plants but in this case the light of the transilluminator shines directly into the camera. Therefore the blocking factor of the emission filter and the overlap of the emission spectra of transilluminators and the emission filters are the critical factors.


Different Petri dishes were used, 12 x 12 cm dishes from Greiner-BioOne and 9 x 9 cm dishes from Falcon. Agar (8 g/L) and growth media (8 g/L agar, 2,15 g/L MS-salt obtained from Duchefa, 0,5 g/L MES, adjusted to pH 5,7 with KOH) were filled into the dishes at four different heights.
The transilluminators were positioned in the optical axis of NightSHADE LB 985 IKlu, manufactured by Berthold Technologies. Holes in the base plate allow easy positioning. An interference emission filter with central peak at 520 nm and a half band width of 10 nm was attached in front of the lens. Intensities of light were measured in relative light units (RLU = cps in this case) with indiGO software.


The linear dependency of background intensities against exposure time and transilluminator intensity shows very clearly that the limiting factor of trans-illumination is the blocking factor of the used interference filter.
The 470 nm transilluminator shows much lower background than a 315 nm or 365 nm transilluminator.
With a 470 nm transilluminator model from Berthold set to 10% intensity, all GFP-transfected plants in Petri dishes filled with agar can be detected, which show higher emission intensities than 800-1000 RLU.
Compared with luminescence, Luciferase-transfected plants can be detected, if the signal intensities are higher than 30 RLU. But for screening and clone-picking of GFP-transfected seedlings in Petri dishes the trans-illumination method could be an alternative.