Figure 7. Schematic view of the leaf types measured both in the common garden (CG) and chamber experiments. Plants are not to scale and it was not uncommon for more unmarked leaves to be located between leaf triplets than shown. LS = long shoot, SS = short shoot, YFD = youngest fully developed leaf. Figure credit: Veera Varpa. Modified by the author.
a practical consideration, keeping the airflow rate constant and relatively high (while letting the RH fluctuate a little) notably accelerates the
response times and thus hastens measuring. The leaf was kept at constant temperature (except for temperature response curves) by keeping the cooling block temperature either at 23°C (in the field) or at 20 – 22°C (in the chambers). PPFD given to the leaf was 1800 μmol photons m-2 s-1 (except for the variable PPFD supplied to the leaf in the light response curves in II, or the measurements in III). This PPFD is saturating but not photoinhibitory for chamber-grown silver birch (II). Before taking a measurement, the leaf was always allowed to acclimate to reach a steady-state. Measurements were always made during 8:00 – 16:00h (mostly during 9:00 – 15:00 and the majority of the measurements well before afternoon) to ensure maximal gas exchange (Sellin et al. 2010). Maximal gas exchange rates were verified before the experiment with diurnal measurements of gs (Fig. 8).
Figure 8. Diurnal patterns of stomatal conductance. Measured on 5.5.2017 from non-experimental, greenhouse-grown plants with the SC-1
porometer (Decagon Devices, Pullman, WA, USA).
In (I), light-saturated snapshot GE was measured in 2013, the third growing season at the CG, from early leaves on short shoots (SS) and late leaves on long shoots (LS). GE was recorded during two measuring
campaigns, during June 5th – 13th (day of year or DOY 156 – 164, SS leaves only) and during June 24th – July 7th (DOY 175 – 188, both SS and LS leaves) from 3 – 5 plants per genotype, using the same plants on both rounds. This data was utilized by constructing two models. In the first model, pooled data of the two measuring rounds was used to better detect the overall provenance-effect, and in the other model a subset of data was used to temporally restrict the measurements to the second measuring round to detect differences between leaf-type. In addition, a time-series of stomatal conductance was examined (Supplementary Fig. S7 of I).
In (II), snapshot GE was monitored from all plants through the lifespan of a single leaf (YFD1) from the moment it was fully expanded and
measurable to the time when it was already showing slight symptoms of senescence (four measuring rounds during 50 – 90 DAP). Additionally, GE response curves were made for temperature and light. Response curves were made by varying either the temperature or light given to a single leaf, but chamber temperature and illumination were kept stable. The
temperature response of Anet – T and gs – T was measured from YFD2 leaves from 20 plants from one chamber (during 73 – 98 DAP). First the leaf was cooled down to 10 – 12°C (until no decrease in temperature was
attained in 10min), a measurement was taken and then the temperature raised in 3°C steps, taking a measurement at each, through 13, 16, 19, 22, 25, 28 and 31°C. To assist the cooling provided by the instrument,
additional cooling was attained by cold-water tubing around the Peltier elements and ice gel packs around and under the leaf chamber. With this makeshift-solution it was possible to lower the measuring head
temperature down to 10°C. Because of the extremes of temperature, RH was kept lower than normal at 40% and RH and VPD closely monitored.
Rarely tested in research articles, VPD vs. gs and VPD vs. internal CO2
partial pressure (Ci) were checked in (II) to examine the variation in these properties due to varying VPD. Light response was measured from YFD1 leaves of 12 plants (during 92 – 94 DAP). First the leaf was given a very high PPFD of 2500 μmol photons m-2 s-1 for 15min, after which light levels were rapidly decreased in sequence, taking a measurement at each, through 2000, 1800, 1500, 1250, 1000, 750, 500, 300, 150, 100, 50, 25 and 0 μmol
photons m-2 s-1. The high initial PPFD fully opens the stomata and closes the PSII centers. While the PSII centers quickly react to the decrease in PPFD, the rapid technique does not allow the stomata to fully respond in time. Also, while this technique is fast, easy and generally keeps values stable, at lower light levels the sample cell RH can decrease below
desirable. During the measurements RH was therefore slightly below 50%
at times.
In (III), snapshot GE measurements were first measured from all available plants from leaf 3-1 (during 85 – 88 DAP) with PPFD of 200 μmol photons m-2 s-1 to correspond to chamber ambient PPFD. The plants were measured once during 8:00 – 12:00h and once during 12:00 – 16:00h.
These measurements had similar distributions, and were thus pooled and used together to represent a daily average photosynthesis. Later, GE was measured otherwise similarly, but with a saturating PPFD of 1000 μmol photons m-2 s-1 mainly from leaf 4-1 (during 111 – 112 DAP) to see if the measurements at ambient PPFD over a four day campaign were mostly similar to those made at saturation during a shorter time period. The distributions of values of both styles of measuring were not identical, but similar enough to base conclusions on either data. As there were more measurements in the first dataset, it was used in the main text of (III).
Further, in (III), mass-based net photosynthesis, Amass (μmol CO2 g-1 s-1), was calculated as Anet divided by leaf mass per area (LMA).
2.3 Growth, leaf longevity, structural leaf traits and biomass