Measurement of growth and physiological indices
(1) Measurement of seedling growth indices:
Before planting the seedlings, the height of each seedling were measured with a measuring tape (accuracy: 1 mm), respectively; the seedings were tagged accordingly. The duration of the experiment was one year; measurements of the plant height each seedling were repeated two weeks before the end of the experiment.
(2) Measurement of seedings biomass accumulation indices:
The duration of the experiment was one year; At the end of the experiment, each seedling was collected and finished in an oven at 110℃, then dried to constant weight after being adjusted to 80℃. An electronic balance with a precision of 0.01g was used to weigh the dry matter.
(3) Determination of antioxidant enzyme activities:
Antioxidant enzyme activities can reflect to a certain extent the differences in the adaptability of plant seedlings to the environment (Krauss, et al., 2014). Therefore, in this study, the amount of three stress-related substances, including superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA), were selected to determine the seedling adaptability to environmental changes.
① SOD activity was determined by the nitrogen blue tetrazolium (NBT) method. The total SOD activity (u·g-1) is determined by the unit number of enzyme per unit of fresh weight, where one unit of enzyme activity is defined as the amount of enzyme required to inhibit NBT reduction to half of the control.
② CAT activity was measured by the ultraviolet absorption method. The amount of enzyme that A240 reduces by 0.1 within 1 min is defined as one enzyme activity unit (u·g-1·min-1).
③ MDA content was determined by the thiobarbituric acid (TBA) method. The MDA content is expressed as the number of moles of MDA per unit of fresh weight. The unit is µmol·g-1.
The absorbance values of the above enzymes were all measured using the Lambda 35 UV/VIS spectrometer (Perkin Elmer, USA).