Stomatal traits
At the final harvest, one leaf from each of three different seedlings
was randomly selected for epidermal prints; the representative leaf
section at about one third from the apex and one third from the midvein
was examined. We first brushed acetone onto surfaces of these leaves,
and then pressed an acetate layer onto them firmly for 30 s and waited
for them to dry. Subsequently, we peeled off the acetate layer and
mounted it onto a slide for stomatal analysis. Stomatal number of each
of ten randomly selected views (0.12 mm2 at 100 ×
magnification) was counted and averaged. When prints did not have
sufficient large undamaged and clear areas, smaller areas (0.01- 0.05
mm2 at 400 × magnification) were examined. Stomatal
density was determined as the summed number of stomata on both upper and
lower surface per one-sided leaf area. The stomatal area was defined as
the area of a guard cell pair, i.e. double ellipse 2*a*b*π, where
a and b are the maximum length and maximum width of the guard cells of
ten randomly selected closed stomata, respectively (Cornelissen et
al. , 2003a). Leaf total stomatal area was defined as stomatal density
multiplied by average leaf area and average stomatal area.
Correspondingly, plant total stomatal area was defined as stomatal
density multiplied by plant total leaf area times average stomatal area.
Leaf total stomatal number was defined as stomatal density multiplied by
average leaf area. Similarly, plant total stomatal number was defined as
stomatal density multiplied by plant total leaf area.