*5.5. Chlorophyll Fluorescence Measurements*

After measuring the rate of photosynthesis, a Chlorophyll a fluorescence measurement was performed with the third fully expanded leaf on the alfalfa seedlings. Before measurement, each seedling was held in a dark chamber for 30 min prior to being submitted to the chlorophyll fluorescence procedure using an Li-6800 portable photosynthesis system. Fluorescence parameters characterizing the state of the photosynthetic apparatus were calculated on the basis of induction fluorescence curves obtained using data from the JIP test, which is usually used to evaluate the state of PSII. On the basis of induction fluorescence curves (OJIP curves), the following parameters, which characterize the maximal PSII quantum yield (Fv/Fm), effective PSII quantum yield (ΦPSII, (F<sup>m</sup> <sup>0</sup> − Ft)/Fm), non-photochemical quenching (NPQ, Fm/F<sup>m</sup> <sup>0</sup> − 1) and electron transport rate (ETR), were determined. Here, F<sup>v</sup> is the value of variable fluorescence, equal to the difference between Fm and F0; F<sup>0</sup> is the minimum amplitude of fluorescence (F); and F<sup>m</sup> is the maximum amplitude of fluorescence. F<sup>m</sup> and Fm' are the maximum Chl fluorescence levels under dark- and light-adapted conditions, respectively. F<sup>v</sup> is the photoinduced change in fluorescence, and F<sup>t</sup> is the level of fluorescence before a saturation impulse is applied. F<sup>0</sup> is the initial Chl fluorescence level. All parameters were calculated according to the methods reported by Pashkovskiy et al. (2021) [62].
