Greenhouse germination experiment
Since our knowledge of the most appropriate pre-treatment for seed
germination of V. negundo var. heterophylla is unknown, we
carried out pre-experiments. The mature seeds of V. negundo var.heterophylla were collected from 10 naturally distributed plants
(ground diameter >10 cm, growing evenly) of the study site
in September 2017. After collection, the fruit stalks and persistent
calyxes were rubbed off, then these seeds were stored in a refrigerator
at 4 ℃. Greenhouse germination experiment was carried out from November
to December 2017. The seeds were soaked for 24 hours with gibberellin
acid (GA3) of different concentrations containing zero
(CK), 0.2 ‰, 0.6 ‰, 0.8 ‰, 1.0 ‰ and 0.2% peroxide hydrogen
(H2O2). The 9-cm Petri dishes with moist
double-decked filter papers were used as germination beds, 20 seeds were
placed uniformly in each dish, and 30 dishes (six pre-treatments × five
replicates) were kept in an incubator at 25℃, 60% relative humidity, 14
h light and 10 h darkness for 20 days. The filter papers on the dishes
were kept moist throughout the experiment. Seeds were considered
germinated at the emergence of the radicle (Bewley and Black 1994), and
without new seed germination for 7 consecutive days was marked as the
end of the test. Seed germination was recorded daily and expressed as a
% percentage of the total number of tested seeds (germination
percentage). The final germination rate (germination percentage at 20
days after the test started) and germination energy (germination
percentage at 8 days after the test started) were calculated. Trough
studying the effect of different concentration and type of solution on
the seed growth, the optimum pre-treatment solution was obtained in the
greenhouse germination process. The results of this part of the
experiment are shown in Fig 1.