were assigned randomly to each of 10 benches that were raised 61 cm off the greenhouse floor. Shade treatments were assigned randomly to each of the trays under commercially prepared 30%, 55%, 80%, or 95% shade cloth (PAK 100% polypropylene shade fabric, Hummert, Intl., Earth City, Mo.). Control plants were grown on benches that were not shaded. For each shade treatment, a wooden shade frame (91 × 244 × 91 cm) was affixed to the bench corners. Shade cloth was draped over the frame, cut, and stapled to the frame so that it did not overlap on the top or sides of the frame. A vertical slit was made through the cloth down one long side of each frame to provide access to the seedlings for growth measurements. The slit remained closed otherwise. All seedlings were fertigated twice weekly with 250 mg·L-1 of a 26N–8.6P–16.6K water-soluble fertilizer also containing trace elements (Peters 20–20–20; Scotts Co., Marysville, Ohio).
On 3 Sept. 1996, height, leaf number, and total leaf area were determined for each plant. Leaf area was measured with a CI-203 portable laser leaf area meter and a CI-203-CA conveyor attachment (C.I.D., Vancouver, Wash.). Roots were washed of attached soil and patted dry with a paper towel. Then lateral roots were trimmed from the taproots. Stems, leaves, lateral roots, and taproots of each plant were dried for at least 48 h at 50 circC in a convection oven. Dry weight (DW) was determined gravimetrically using an electronic balance (model A-160; Fischer Scientific, Pittsburgh, Pa.).
OUTDOOR STUDY (EXPT. 2). A randomized compete block design experiment was conducted outdoors with two blocks. Treatments consisted of five levels of shade (0%, 28%, 51%, 81%, or 98%), as in the greenhouse study, with 20 seedlings included in each experimental treatment per block. Seeds were sown 29 Apr. 1998, and allowed to germinate in the greenhouse as described previously. Once seedlings in Rootrainers reached the two- to three-leaf stage (11 June 1998), they were placed outdoors, and the same benches and attached shade structures described in the greenhouse study were used to impose the shade treatments. Benches were placed directly on grass in a field far enough from building structures to avoid shading. Control seedlings were not shaded. Blocks were oriented in an east–west orientation on a gentle slope. Plants were watered as needed and fertilized weekly with 250 mg·L-1 of a 20N–8.6P–16.6K water-soluble fertilizer containing soluble trace elements (Peters 20–20–20). Plants were destructively harvested on 27 Aug. 1998, 11 weeks after the start of the experiment. Leaf number, seedling height, and leaf area were recorded. Weight of dried stems, leaves, and roots was also determined gravimetrically as described for Expt. 1.
PHOTOSYNTHETIC PHOTON FLUX, AIR TEMPERATURE, AND SPECTRAL ANALYSIS. At noon on a representative sunny day, four replicate measurements per treatment were made at midcanopy level and recorded for a) PPF (400 to 700 nm), using a quantum/radiometer/photometer with a quantum sensor (LI-185B and Q7072, respectively; LI-COR, Inc., Lincoln, Nebr.), b) red (660 nm) and far-red (730 nm) light with a portable spectroradiometer (LI-1800), and c) UV light (250 to 400 nm) using an ultraviolet meter (Apogee Instruments, Spectrum Technologies, Inc., Plainfield, Ill.). In the greenhouse experiment, air temperatures were measured inside and outside shade structures for each bench at seedling height, using a thermistor thermometer (Digisense; Cole-Parmer, Vernon Hills, Ill.). In the outdoor experiment, the temperature inside and outside shade structures was measured every 30 min for 24 h using sensors (TEMP HOBO; Onset Computer Corp., Pocasset, Mass.).
CHLOROPHYLL DETERMINATION. At the time of destructive harvest in the greenhouse and outdoor studies, leaf discs were collected from the apical, medial, and basal expanded leaves of each plant. Four discs (0.32 cm² each) were removed from the middle of the leaf using a paper hole punch. Discs were extracted in 8 mL of N,N-dimethylformamide (Sigma, St. Louis, Mo.) in darkness at 5 °C for at least 36 h. Absorbance of extracts was read at 625, 664, and 647 nm on a DU-70 UV/Vis spectrophotometer (Beckman Instruments, Fullerton, Calif.). Calculations for chlorophyll (chl) a, chl b, and protochlorophyll (p) chl were made as described by Moran (1982).
Four discs (0.32 cm² each) were removed from the middle of the leaf using a paper hole punch. Discs were extracted in 8 mL of N,N-dimethylformamide (Sigma, St. Louis, Mo.) in darkness at 5 °C for at least 36 h. Absorbance of extracts was read at 625, 664, and 647 nm on a DU-70 UV/Vis spectrophotometer (Beckman Instruments, Fullerton, Calif.). Calculations for chlorophyll (chl) a, chl b, and protochlorophyll (p) chl were made as described by Moran (1982).
STATISTICAL ANALYSIS. Data from both studies were subjected to GLM analysis of variance (ANOVA) or stepwise GLM regression analysis using the statistical program Costat (CoHort Software, Minneapolis, Minn.). Where appropriate, treatment means were separated using least significant difference (LSD) or an unpaired t test.
GREENHOUSE STUDY (EXPT. 1). Incident light levels were reduced by 33%, 56%, 81%, and 98% using the 30%, 55%, 80%, and 95% shade fabric, respectively (Table 1). Temperature and red to far-red light ratios were similar in all treatments. UV irradiance declined with increased shading (Table 1).
ANOVA indicated there was a significant interaction between shading and Cu(OH) 2(P < 0.001) for total leaf area, average leaf area, average plant SLW, lateral root DW, and total root, shoot, and plant DW. In plants without container treatment with Cu(OH) 2, leaf number showed a quadratic response with increasing shade level, with plants under 81% or 98% shade exhibiting a decline in leaf number compared
area, average plant SLW, lateral root DW, and total root, shoot, and plant DW. In plants without container treatment with Cu(OH) 2, leaf number showed a quadratic response with increasing shade level, with plants under 81% or 98% shade exhibiting a decline in leaf number compared
Fig. 1. Effect of shading and treatment of containers with (open symbols) and without (closed symbols) cupric hydroxide [Cu(OH)2] on growth of pawpaw seedlings in the greenhouse. (A) Leaf number of plants with (y = 13.632 + 0.135x - 0.002x2, R2=0.89, P=0.001) and without Cu(OH)2 (y = 14.938 + 0.1655x - 0.002x2, R2=0.92, P=0.001), (B) average leaf area in plants with (y = 34.22 + 1.08x - 0.01x2, R2=0.64, P = 0.001) and without Cu(OH)2 (y = 28.01 + 0.89x - 0.01x2, R2=0.68, P = 0.001), and (C) average plant specific leaf DW (SLW) in plants with (y = 4.160 - 0.017x - 0.001x2, R2=0.78, P=0.04) and without Cu(OH)2 (y = 4.216 - 0.035x + 0.001x2, R2=0.98, P = 0.001). Each symbol is the mean of two blocks ± se.*Indicates that means were significantly different between Cu(OH)2 treated and non-treated plants using an unpaired t test (P < 0.05, n = 2).
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