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Soaking, moist-chilling, and temperature effects on germination of Acer pensylvanicum seeds A. Bourgoin and J.D. Simpson
Abstract: The effect on germination of soaking duration, moist-chilling time, and temperature was evaluated using five seed lots of Acer pensylvanicum L. Seeds were soaked for 0, 48, 72, or 96 h, then moist chilled at 4 °C for 16, 24, or 32 weeks. Two temperature regimes were used for germination: (i) 16 h dark at 5 °C : 8 h light at 15 °C (5:15 °C) and (ii) 16 h dark at 20 °C : 8 h light at 30 °C (20:30 °C). Soaking and chilling seeds significantly increased germination. Germination was highest at 5:15 °C, but the germination speed was slow. Germination at 20:30 °C was lower, but 94%–98% of ungerminated seeds appeared to be viable, suggesting that they were dormant. Overall results showed that soaking seeds for 48 h, moist chilling for 16 weeks, and germinating at 5:15 °C produced an average germination of 92%. Résumé : L’effet du temps de trempage, de la durée d’exposition au froid humide et de la température sur la germination des graines a été évalué à l’aide de cinq lots de graines d’Acer pensylvanicum L. Les graines ont été mises à tremper pendant 0, 48, 72 ou 96 h et exposées ensuite au froid humide à 4 °C pendant 16, 24 ou 32 semaines. Deux régimes de température ont été utilisés pour la germination : (i) 16 h d’obscurité à 5 °C : 8 h de lumière à 15 °C (5:15 °C) et (ii) 16 h d’obscurité à 20 °C : 8 h de lumière à 30 °C (20:30 °C). Le trempage et l’exposition au froid ont significativement augmenté la germination des graines. Le taux de germination était maximal à 5:15 °C mais la vitesse de germination était lente. Le taux de germination était plus faible à 20:30 °C mais 94 % à 98 % des graines non germées semblaient viables; ce qui porte à croire qu’elles étaient dormantes. Dans l’ensemble, les résultats ont montré que le trempage des graines pendant 48 h, l’exposition au froid humide pendant 16 semaines et la germination à 5:15 °C produisent un taux de germination de 92 %. [Traduit par la Rédaction]
Bourgoin and Simpson
Introduction Striped maple (Acer pensylvanicum L.) ranges west from Cape Breton Island, Nova Scotia, to central Ontario, southward through New England and the Atlantic states, and along the Appalachian Mountains to northern Georgia (Sullivan 1983). This species is generally found on moist, cool sites, where it grows as an understorey shrub or tree (Sullivan 1983) in mixedwood and hardwood stands (Gabriel and Walters 1990). Acer pensylvanicum seeds germinate early, with radicle elongation presumed to start in April and May (Wilson et al. 1979). Seeds of A. pensylvanicum can lie dormant until the second year before they germinate (Gabriel and Walters 1990), which indicates that it has one of the lonReceived 4 December 2003. Accepted 11 May 2004. Published on the NRC Research Press Web site at http://cjfr.nrc.ca on 6 November 2004. A. Bourgoin. New Brunswick Department of Natural Resources, P.O. Box 6000, Fredericton, NB E3B 6H6, Canada. J.D. Simpson.1 Natural Resources Canada, Canadian Forest Service, National Tree Seed Centre, P.O. Box 4000, Fredericton, NB E3B 5P7, Canada. 1
Corresponding author (e-mail:
[email protected]).
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gest stratification requirements in the genus (Zasada and Strong 2003). Factors frequently contributing to the dormancy of A. pensylvanicum seeds are the presence of inhibitors, usually in the embryo, and the action of the seed coat in reducing the rate of imbibition and inhibitor leaching (Wilson et al. 1979) or restricting radicle elongation (Gabriel and Walters 1990). Peck and Lersten (1991) indicated that both the pericarp and seed coat have been identified as causes of dormancy in the Acer genus. Maguire (1975) defined seed dormancy as the inability of seeds to germinate under favorable conditions. This may be due to any or several of the following causes: (i) immature embryos, (ii) seed coats impermeable to water and (or) gases, (iii) inhibitors, (iv) light sensitivity, (v) mechanical restriction by seed coats, or (vi) adverse environmental conditions (Maguire 1975, 1976, 1984). Ellis et al. (1985) stated that there are three main types of dormancy: (i) Innate dormancy (primary dormancy), which is present immediately after the embryo ceases to grow. It can be divided into two categories: seed coat dormancy and embryo dormancy. (ii) Induced dormancy (secondary dormancy), when a seed has lost its innate dormancy but some other particular factor is unfavorable (e.g., high temperature or low oxygen). (iii) Enforced dormancy (environmental dormancy), which occurs
doi: 10.1139/X04-092
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when viable seeds do not germinate because of some limitation in the environment. There are four phases of seed dormancy: induction, maintenance, trigger, and germination (Amen 1968). Much research has focused on alleviating dormancy that is related to the trigger phase. Amen (1968) listed several triggers that effectively alleviate dormancy, including low temperature, light, and dry storage. Another important factor in alleviating dormancy is seed imbibition, because it is necessary to activate metabolism (Bonner et al. 1974; Khan 1982; Maguire 1984). Khan (1982) stated that stratification (moist chilling) is another effective method of alleviating dormancy. A project was undertaken to evaluate practical methods to alleviate dormancy and promote germination of A. pensylvanicum seed. Because of its noncommercial value, little research has been done on this species. Preliminary testing indicated that germination increased as the duration of moist chilling increased from 8, to 16, to 24 weeks (Daigle and Simpson 2001). Removal of the pericarp has been suggested as the only means of obtaining high germination levels (Wilson et al. 1979), but this is time consuming and not practical for large quantities of seed. Recommended germination methods for Acer spp. include prechilling seeds for 45–130 d followed by germination at 20:30 °C for 28 d (Association of Official Seed Analysts 2002). For this project, the effect on seed of three treatments was evaluated: duration of soaking, duration of moist chilling, and germination temperature.
Materials and methods Five seed lots were selected, collected in the fall of 2000, from south-central New Brunswick and eastern Nova Scotia, Canada. Three were single-tree collections and two were bulk collections (seed lot 119 comprised 7 trees and seed lot 120 consisted of 10 trees). Seeds were stored at –20 °C for 2 months before initiating the experiment. Seed moisture content at the time of storage averaged 7.9% and ranged from 7.5% to 8.0%. Seeds were submerged and soaked in aerated, distilled water at room temperature (22 °C) for 48, 72, or 96 h. The control sample was not soaked. Water was changed every 24 h. After soaking was complete, the seeds in each seed lot were surface dried with paper towel to remove excess water. Using a single replicate, a 2-g sample (10–12 seeds) was removed from each seed lot and soak time to determine moisture content, following procedures outlined by the International Seed Testing Association (2003). Three chilling times of 16, 24, or 32 weeks were used. Seeds were placed on Kimpak®, moistened with 125 mL of deionized water, in Petawawa germination boxes (Wang and Ackerman 1983). An additional 125 mL of water was added to the reservoir of each box. One hundred seeds were placed in each box. The boxes were placed in a walk-in cooler maintained at 4 °C. Seeds were checked every 3 weeks to ensure the Kimpak remained moist. Kimpak in any boxes that required more water was remoistened with 50 mL of deionized water. When the moist-chilling period was complete, germination boxes were placed in Conviron™ G30 germinators. One germinator was set at 5 °C for 16 h without light and 15 °C
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for 8 h with light (5:15 °C); the second was set at 20 °C for 16 h without light and 30 °C for 8 h with light (20:30 °C). Relative humidity was maintained at 85%. The 20:30 °C germination temperature regime is the standard used for most germination tests, and the cooler 5:15 °C temperature regime was used to approximate natural conditions during April and May in forest stands in New Brunswick. Seeds were assessed every 7 d and were left in the germinators for 8 weeks. Seeds were deemed to have germinated when the radicle, hypocotyl, and cotyledons had emerged from the seed. Germinants, at this stage, were counted and removed from the boxes. At the end of 8 weeks, ungerminated seeds were cut to determine whether they were viable. Seeds were considered viable if the cotyledons and radicle were firm and green. The experiment was set up using a nonreplicated factorial design. Total germination at week 8 was used for data analysis. Before analysis, data were transformed to arcsin px/100, where x is the percent germination. Data were analyzed using the ANOVA procedure of SAS, and means were subjected to a Duncan’s multiple range test to evaluate significant differences within main effects and reported as untransformed values (SAS Institute Inc. 1990).
Results For all seed lots, soaking seeds rapidly increased moisture content during the first 48 h (7.9%–58.7%, fresh mass basis), but its effect diminished thereafter (63.5% and 65.8% after 72 and 96 h, respectively) (results not shown). Mean germination of all seed lots for each chilling and soaking time was much higher in the 5:15 °C germination condition than in the 20:30 °C condition (Figs. 1A and 1B). Chilling beyond 16 weeks improved germination at 20:30 °C only. The decline in germination at 5:15 °C for seeds soaked for 96 h and chilled for 24 weeks was due to the consistently poorer germination of seed lot 123 and the comparatively lower germination of seed lots 118 and 120 for that treatment combination. Each of the four main effects exhibited significant differences, but first-order interactions involving soaking time were not significant at P < 0.01 (Table 1). Chilling time and germination temperature significantly interacted with seed lot as well as with each other. Germination was significantly different among seed lots, with seed lot 123 clearly being inferior (Table 2). Seeds that were soaked had a higher germination, but increased soaking time did not significantly improve germination (Table 2). Germination increased with chilling time, with the largest increase occurring from 16 to 24 weeks, due primarily to increased germination at 20:30 °C. The 5:15 °C germination temperature regime resulted in higher germination, however 7–8 weeks were required for complete germination to occur. Germination at 20:30 °C was completed in 3–4 weeks, because the seeds had started to germinate during chilling.
Discussion Soaking seeds for 48 h increased germination but longer soak times did not substantially improve it (Table 2). In similar studies, Webb (1974), Webb and Dumbroff (1969), and © 2004 NRC Canada
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Fig. 1. Mean of five seed lots of Acer pensylvanicum seeds germinated at (A) 5:15 °C and (B) 20:30 °C after four soaking times and three periods of moist chilling.
Table 1. Analysis of variance summary for germination of Acer pensylvanicum seeds. Source Seed lot Soak time Chill time Germination temperature Seed lot × soak time Seed lot × chill time Seed lot × germination temperature Soak time × chill time Soak time × germination temperature Chill time × germination temperature Error
df 4 3 2 1 12 8 4 6 3 2 74
MS 0.5884 0.4992 0.0601 21.0851 0.0142 0.0421 0.1186 0.014 0.0319 0.6015 0.013
P value
