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4.2.2 Water content and imbibition rate
Germination physiology
Water content was determined by weighing seeds before and after placing them in a
drying oven set at 110°C. The weighing continued for six weeks until there was no
further loss in seed weight. Per species, four replicates of 5 seeds each were used.
The imbibition rate was determined by weighing the seeds after imbibing them for 3,
6, 24, 48, 150, 168, 216, 264 and 336 h. Seeds were placed in 6.5 cm disposable
plastic Petri dishes with filter paper (Whatman No.1) moistened with 3.5 ml of distilled
water and subsequently kept moist by adding 1 to 3 ml of distilled water when
needed. At each time interval, the seeds were removed from the Petri dishes, blotted
dry, weighed and replaced in the respective Petri dishes. This experiment was
conducted at room temperature (25 ± 0.5°C) using four replicates of 5 seeds for each
species.
4.2.3 Scanning electron microscopy
Seeds were rinsed in 70% ethanol for 30 seconds and then allowed to dry on a paper
towel. Seeds were mounted on 12 mm stubs and the micropylar regions and
surfaces viewed using a scanning electron microscope (Zeiss EVO LS15 variable
pressure).
4.2.4 Ex vitro germination experiments
Experiments were conducted to test the effect of temperature and light, cold and
warm stratification, acid scarification, mechanical scarification, plant growth
promoting substances and deficiency of nitrogen, phosphorous and potassium on
germination of different Romulea species.
If not stated otherwise, a growth chamber set at 20°C with a 16 h light: 8 h dark
photoperiod and a photosynthetic photon flux density (PPFD) of 30.1 ± 4.0 µmol m -2
s -1 provided by cool-white fluorescent lamps was used. Seeds were decontaminated
by placing them in 0.2% mercuric chloride for 5 min and rinsing them once with tap
water and twice with distilled water. Unless otherwise stated, seeds were placed in
6.5 cm disposable plastic Petri dishes with two circles of filter paper (Whatman No.1)
moistened with 3.5 ml of distilled water and test solutions. The filter paper was kept
moist with the respective solutions when needed. In all germination experiments 10
seeds were placed in each Petri dish with 4 replicates per treatment. All germination
experiments were terminated after 90 days. Germination was recorded every second
day and was considered complete when the radicle had emerged up to 2 mm. Seeds
100