Photosynthesis of plantlets in tissue culture containers is not considered important, compared to photosynthesis of ex vitro plants, due to the exogenous source of carbohydrates present
in tissue culture media. However, CO2 starvation can generate a burst of reactive oxygen species
(ROS). We examined this phenomenon in tissue culture, since CO2
levels may become very low
during the light period. The research was carried out with lily scales, regenerating adventitious
bulblets, and with Arabidopsis seedlings. CO2 starvation was achieved by placing a small vial of
concentrated KOH solution in the culture container. CO2 removal reduced the growth of regenerated
lily bulblets by 33% or 23%, with or without sucrose in the medium, respectively. In Arabidopsis
seedlings, CO2 removal decreased growth by 50% or 78% in the presence or absence of sucrose in
the medium, respectively. Therefore, the addition of sucrose as a replacement for photosynthesis
resulted in only partial recovery of growth. Staining with nitroblue tetrazolium (NBT) showed little
to no ROS in ex vitro growing seedlings, while abundant ROS were detected in seedlings grown
under in vitro CO2 starvation. Seedlings grown under normal tissue culture conditions (no CO2
withdrawal) showed low levels of ROS. In lily tissue culture, CO2 starvation decreased the maximum
quantum efficiency of photosystem II (Fv/Fm) from 0.69 to 0.60, and in Arabidopsis from 0.76 to 0.62.
Fv/Fm of ex vitro lily and Arabidopsis seedlings was 0.77 and 0.79, respectively. This is indicative of a
disturbance in photosynthesis functionality and the occurrence of in vitro stress under reduced CO2
concentrations. We conclude that poor growth, in the absence of CO2
, was partly due to strongly
reduced photosynthesis, while the detrimental effects were most likely due to a burst of ROS.