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Genetic control of germination and the effects of accelerated aging in mountain hemlock seeds and its relevance to gene conservation
Forest Ecology and Management 112 (1998) 203±211
Genetic control of germination and the effects of accelerated aging in mountain hemlock seeds and its relevance to gene conservation Y.A. El-Kassabya,*, D.G.W. Edwardsb a
Department of Forest Sciences, Faculty of Forestry, The University of British Columbia, Vancouver, BC, Canada V6T 1Z4 b Canadian Forest Service, Paci®c Forestry Centre, 506 West Burnside Road, Victoria, BC, Canada V8Z 1M5 Accepted 25 May 1998
Abstract Genetic control of germination parameters and the effects of accelerated aging in mountain hemlock (Tsuga mertensiana [Bong.] Carr.) seeds were investigated using standard germination tests and simulated aging, respectively. Germination parameters were studied on seeds collected from individual trees from two natural populations (Sooke and San Juan) located on southern Vancouver Island, BC. Strong genetic control was con®rmed by the high heritability estimates that ranged from 0.35 to 0.82 (strati®ed) and from 0.58 to 0.73 (unstrati®ed) for Sooke and from 0.30 to 0.85 (strati®ed) and from 0.45 to 0.84 (unstrati®ed) for San Juan. Simulated aging was conducted on eight seedlots collected from natural stands representing the species distribution in British Columbia. Strati®ed and unstrati®ed seeds were aged at 100% RH and 37.58C for 0 to 21 days at 3-day intervals. Seed aging accounted for the majority of variation which ranged from 97.61% to 99.86% of the total variation observed. Gradual loss of seed viability was observed over the aging treatments with a total loss of viability after aging for 12 days. Differential losses in viability among seedlots were maximum after aging for 6 days. Results are discussed in terms of the impact of long-term storage and the reliance on seed banks for ex-situ conservation of this species. # 1998 Elsevier Science B.V. All rights reserved. Keywords: Tsuga mertensiana; Seed germination; Heritability; Gene conservation
1. Introduction Mountain hemlock (Tsuga mertensiana) is a longlived, subalpine conifer that is most abundant in coastal regions of Alaska and British Columbia. Its range extends to central California (Parsons, 1972), with isolated populations in the interior of British Columbia, Idaho and Montana. In general, its populations are disjunct due the physical separation of the *Corresponding author. Tel.: +1-(250)-381-1404; fax: +1-(250)381-0252; e-mail: [email protected]
high elevation sites where they occur. It is a valuable species for watershed protection and wildlife habitat (Harestad, 1980). Timber harvesting in British Columbia is reaching higher into subalpine regions, but very little is known about the genetic diversity and regeneration potential/success of the species. Due to the disjunct nature of its distribution, mountain hemlock was included in a world list of threatened species (Farjon et al., 1993). This study is part of a larger project designed to investigate the genetic structure of mountain hemlock using gene markers, adaptive and quantitative
0378-1127/98/$ ± see front matter # 1998 Elsevier Science B.V. All rights reserved. PII: S0378-1127(98)00343-0
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Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
meters in mountain hemlock, and (2) how the genetic representation of the species will be affected by simulated long term storage and its relation to ex-situ gene conservation.
attributes and regeneration (germination) ecology for the purpose of developing a suitable conservation strategy. This can be accomplished by a combination of in-situ and ex-situ approaches. Although seed banks have been proposed as a viable means of ex-situ conservation, a thorough understanding of seed biology (germination and storage) is imperative in developing effective conservation tools. Standard germination tests provide estimates of the maximal potential of a seedlot for seedling production under favorable environmental conditions. These tests do not evaluate seedlot vigour, that is, the ability to produce seedlings under stressful conditions (Association of Of®cial Seed Analysts, 1983). Seed viability is highest at physiological maturity and declines with age (Edwards, 1980), the deterioration being gradual and ending with total loss of germinability and vigour (Delouche and Baskin, 1973). Seed vigour is known to be heritable (McDaniel, 1973; Kneebone, 1976; Dickson, 1980) and may be inherited maternally in some plants (Kueneman, 1983). Degree of seed deterioration/vigour can be revealed through a stress test, such as the accelerated aging test that simulates long-term storage (Delouche and Baskin, 1973; Baskin, 1977; Association of Of®cial Seed Analysts, 1983), which has been used successfully in several tree studies to evaluate the ef®cacy of ex-situ conservation (Pitel, 1980; Blanche et al., 1988, 1990; Marquez-Millano et al., 1991; Chaisurisri et al., 1993). It is important to determine whether this deterioration is independent among seedlots (i.e., genetic) and/or whether seedlots exhibit the same rate of deterioration over time. The objectives of this study were to determine: (1) the degree of genetic control of germination para-
2. Materials and methods 2.1. Seed source Seeds from individual trees in two natural stands of mountain hemlock on Vancouver Island were used to investigate the genetic control of germination parameters. The stands are located on TimberWest Ltd. (formerly Paci®c Forest Products Ltd.) land on the southern tip of Vancouver Island at Sooke (13 trees) and San Juan (20 trees). Eight natural stand bulk seedlots (Table 1) were obtained from the British Columbia Ministry of Forests' Tree Seed Centre for the simulated aging study. Seven of these seedlots were from coastal locations, while the Sale Mt. seedlot was included as a representative of an interior (more continental) seed source. 2.2. Germination Germination tests for individual trees within each population (Sooke and San Juan) were conducted on four replications (100 seeds each) of strati®ed and unstrati®ed seeds. Seeds were spread in 10104 cm, tightly-lidded clear-plastic boxes lined with one layer of Kimpak (cellulose wadding) overlain with three layers of Whatman no. 1 ®lter paper, moistened with 50 ml of distilled water. Boxes were placed in
Table 1 Sources of the eight mountain hemlock natural stand seedlots used in the germination study Seedlot no.a
Local source name
Year collected
Elevation (m)
Lat. (N)
Long. (W)
1000 PSWb (g)
03854 07727 07934 09783 09960 10904 13877 35050
Hoodoo Creek Garbage Creek Hkusam Mt. Kearsley Creek Port Alice Sale Mt. Lyon Lake Hanna Ridge
1979 1982 1982 1982 1982 1988 1982 1990
1250 850 950 1280 750 1700 1005 700
51820' 488 33' 50820' 49819' 50824' 51810' 49839' 56818'
125832' 124806 125850' 122822' 127827' 118810' 123854' 129820'
2493 2193 2283 2059 2387 2293 2323 2003
a b
British Columbia Ministry of Forests seedlot registration numbers. Estimated weight of 1000 pure seeds based on eight replications of 100 filled seeds (Edwards and El-Kassaby, 1996).
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
randomly-assigned positions in an incubator set for a constant 208C. Light (cool-white ¯uorescent) of approximately 100 lux at the ®lter paper surface was provided as a photoperiod of 8 h `day' and 16 h (no light) `night'. Seeds were counted as germinated when the radicle and hypocotyl equalled or exceeded four times the length of the individual seed. Germinants were assessed at 2-day intervals for the ®rst half of each test, then at 3±4 day intervals up to 28 days, and were classed as normal or abnormal according to the International Seed Testing Association, 1993. 2.3. Accelerated aging Based on a pilot study, it was determined that 37.58C was the optimal temperature for accelerated aging of mountain hemlock seeds. At higher temperature (408C and above), the decline in germination occurred too rapidly to clearly delineate differences among seedlots, while lower temperatures required very prolonged aging treatment to obtain complete loss of germinability. This temperature, 37.58C, is lower than that prescribed for agricultural seeds (Association of Of®cial Seed Analysts, 1983), but agrees with that recommended for tree seeds in other studies (Blanche et al., 1988; Chaisurisri et al., 1993). Accelerated aging was applied for eight periods from 0 to 21 days at 3-day intervals (i.e., 0, 3, , 21 days). For each aging period, eight replications of 100 seeds were placed on wire-mesh platforms in tightly-closed, clear-plastic germination boxes. Each box contained 50 ml of distilled water; the distance between the platform and the water surface was 1.5 cm. Thus, for every seedlot a total of 64 germination boxes containing 100 seeds each were used. For every seedlot and every aging period, the accelerated aging tests were conducted on four replications of strati®ed and unstrati®ed seeds. Prior to aging, four of the eight replications for each seedlot were imbibed for 24 h, drained, placed in plastic bags and strati®ed at 28C for 28 days. The remaining four replications were refrigerated (18C, dry) until required for the aging treatments. Aging and strati®cation treatments were scheduled so that all subsequent germination tests began at the same time. Thus, strati®ed and unstrati®ed samples that received the 21-day aging treatment were placed in the incu-
205
bator ®rst, and those to receive the 0-day treatment were last. Following aging, all samples were rinsed under running cold water for 1 to 2 min to reduce fungal mycelial growth. Germination tests for the aging trial were the same as described above. 2.4. Data analysis Results were expressed as: (i) germination capacity (GC), the percentage of seeds that had germinated normally at the end of the test; (ii) germination rate (R50), the number of days required for 50% of the seeds in the replication to germinate; (iii) germination speed (R50 '), the number of days for 50% of the germinating seeds to germinate (Thomson and ElKassaby, 1993); (iv) peak value (PV), an index of germination speed that expresses germination rate as the maximal quotient derived by dividing daily the accumulated number of normal germinants by the corresponding number of days (Czabator, 1962); this is the mean daily germination of the most-vigorous components of a seedlot, and a mathematical expression of the tangent drawn through the origin of a sigmoid curve representing the typical course of germination. And (v) germination value (GV), which combines germination capacity and speed into a single index (Czabator, 1962). Data were transformed to normalize the calculated response variables, and to achieve homogeneity of variances, as follows: GC by arcsin, R50 and R50' by (1-[1/x1]), and GV by (H[x0.5]), where x is the variable in question; PV was not transformed. The transformed data were analyzed using ANOVA based on the following two linear models: Yijk Ti Sj TSij " ijk
(1)
where overall mean, Tithe effect of the ith tree, i1 to 13 (Sooke) or 1 to 20 (San Jaun)(random), Sjthe effect of the jth seed pretreatment (strati®cation), j1 to 2 (®xed effect), TSijthe effect of the interaction between tree and seed pretreatment, "(ij)kthe residual term, k1 to 4 for germination tests. Yijkl Pi Sj Ak PSij SAjk PSAijk " ijkl
(2)
where overall mean, Pithe effect of the ith pretreatment (strati®cation), i1 to 2 (®xed effect),
206
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
Sjthe effect of the jth seed source, j1 to 8 (random effect), Akthe effect of the kth aging treatment, k1 to 8 (®xed effect), PSikthe effect of the interaction between pretreatment and seed source, PAikthe effect of the interaction between pretreatment and aging, SAjkthe effect of the interaction between seed source and aging treatment, PSAijkthe effect of the interaction between pretreatment (strati®cation), seed source and aging treatment, "(ijk)lthe residual term, l1 to 4 for accelerated aging tests. Expected mean squares (EMS) were calculated and % variance components estimated for each source of variation. Where signi®cant effects were observed, means were compared using the Student±Newman± Keuls range test (Steel and Torrie, 1980). Sources of variation and EMS are shown in Tables 2 and 3. 3. Results and discussion 3.1. Genetic control of germination parameters The germination data were analysed on an individual-population basis due to the lack of data con®rming that the allelic frequencies of genes controlling such attributes are uniform over these two populations. Therefore, heritability estimates were calculated for Sooke and San Juan separately. For both populations, the effects of seed pretreatment (P), expressed as a percentage of the total variation, on germination
parameters were highly signi®cant (P0.01), and accounted for between 20 (GC) to 79 (R50), and between 18 (GC) to 86 (R50') at Sooke and San Juan, respectively (Table 2(a)). Germination capacity ranged from 91% (unstrati®ed) to 97% (strati®ed) at Sooke (Fig. 1), and from 95% (unstrati®ed) to 97% (strati®ed) at San Juan (Fig. 2), while germination rate varied between 14 (unstrati®ed) and 17 (strati®ed) days to reach 50% germination in both populations. At Sooke, when seeds were strati®ed (Fig. 1), variation in germination was reduced (range: 94.6±99.6% vs. 76.9±100%), whereas at San Juan the variation was greatly reduced in 19 out of the 20 trees (range: 95.6± 99.6% vs. 88.7±98.6%) (Fig. 2). Tree number 7 at San Juan showed little response to the seed pretreatment indicating greater variability in seed dormancy among trees in this population. The observed differences between strati®cation treatments created strong polarization in the data, making it dif®cult to discern the true differences among trees. Therefore, complementary analyses were conducted for each seed pretreatment separately (Table 2(b and c)). When seed pretreatment was excluded from the analysis, a shift in the amount of variation was observed and differences among trees within populations became highly signi®cant for all germination parameters. At Sooke, the effects of trees (T), expressed as a percentage of the total variation, on germination parameters ranged from 35 (GC) to 82 (R50'), and from 58 (PV) to 73 (R50'), for strati®ed and
Fig. 1. Germination course of stratified (S) and unstratified (US) seeds from 13 trees and Sooke.
p-1 t-1 (p-1)(t-1) pt(r-1)
d.f.
12 36
19 57
a-Combined analyses Seed pretreatment (P) Tree (T) PT Residual
S.O.V.
b- Sooke. Among trees (B) Within (W) h2
c-San Juan. Among trees (B) Within (W) h2
PV
GV
2W 42B 2W
2W 42B 2W
84.76 15.24 0.85
77.78 22.12 0.78 81.82 18.18 0.82
81.61 18.39 0.82
R'50
29.95 70.05 0.30
34.83 65.17 0.35
GC
83.72 16.28 0.84
64.05 35.95 0.64
PV
80.14 19.86 0.80
60.01 39.99 0.60
GV
81.48 18.52 0.81
65.71 34.29 0.66
84.00 16.00 0.84
73.33 26.67 0.73
R'50
R50
85.9 00.0 06.8 07.4
R50
64.5 25.1 04.2 06.2 Unstratified
64.5 15.2 06.9 13.4
Stratified
61.6 14.0 10.7 13.8
E.M.S3
19.9 14.9 37.3 27.8
R'50
78.8 14.2 01.6 05.4
73.7 18.9 01.5 05.9
R50
GC
R50 R'50
San Juan
Sooke
2e k1 2PT k2 2P 2e k1 2PT k3 2T 2e k1 2PT 2e
E.M.S.2
44.79 55.21 0.45
72.98 27.02 0.73
GC
18.0 20.6 10.3 51.0
GC
79.44 20.56 0.79
58.41 41.59 0.58
PV
25.4 19.2 00.0 55.4
PV
77.11 22.89 0.77
67.60 32.40 0.68
GV
52.4 35.4 03.7 08.4
GV
Germination parameters are (i) germination rate (R50), (ii) germination speed (R50'), p (iii) germination capacity (GC), (iv)germination value (GV), and (v) peak value (PV). Data transformations are as follows: R50 and R50' by (1ÿ[1/x1]), GC by arcsin, GV by x ÿ 0:5, and PV was not transformed. 2 Coefficients of variance components are: Sooke (k14, k28, k352) and San Jaun (k14, k28, k380), 2p variance due to pretreatment, 2T variance due to tree, 2PT variance due to pretreatment tree interaction. 3 2 B variance among trees and 2W variance among replications within tree.
1
d.f.
S.O.V
Table 2 Estimation of variance components (as percentages of the total variation) and broad-sense heritabilities (h2) for germination parameters1 of 2 mountain hemlock populations (Sooke and San Juan).
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Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
Table 3 ANOVA and percent contribution of the variance components (Var. Comp.) for the germination parametersa of eight mountain hemlock seedlots following seed pretreatment and accelerated aging S.O.V.
d.f.
E.M.S.b
Var. Comp.
R'50
GC
GV
PV
Seed pretreatment (P) Seedlot (S) Aging (A) PS P A SA PSA Residual
1 7 7 7 7 49 49 384
2e 3:992PSA 31:972PS p 2e 3:992PSA 7:992AS 31:972PS 63:932S 2e 3:992PSA 7:992SA A 2e 3:992PSA 31:972PS 2e 3:992PSA PA 2e 3:992PSA 7:992SA 2e 3:992PSA 2e
P 2S A 2PS PA 2SA 2PSA 2e
1.17 0.00 97.61 0.01 0.68 0.17 0.00 0.48
0.11 0.03 99.58 0.00 0.06 0.08 0.00 0.16
0.00 0.04 99.86 0.00 0.02 0.16 0.00 0.18
0.00 0.03 99.81 0.00 0.13 0.09 0.00 0.16
a
Germination parameters are (i) germination speed (R50'), (ii) germination capacity (GC), (iii) germination value (GV), and (iv) peak value (PV). Data transformation are as follows: R50' by (1-[1/x1]), GC by arcsin, GV by (Hx0.5), and PV is not transformed. b Pvariance due to seed pretreatment (fixed), 2S variance due to seedlot effect (random), Avariance due to aging treatment (fixed), 2PS variance due to seed pretreatment seedlot interaction, PAvariance due to seed pretreatment aging treatment interaction, 2SA variance due to seedlot aging treatment interaction, 2PSA variance due to seed pretreatment seedlot aging treatment interaction, 2e residual effect.
unstrati®ed seeds, respectively (Table 2(b)). A similar trend was observed at San Juan, and the effect of trees (T) as a percent of total variation ranged from 30 (GC) to 85 (R50), and from 45 (GC) to 84 (R50'), for strati®ed and unstrati®ed seeds, respectively (Table 2(c)). These differences were further emphasized by the Student±Newman±Keuls range tests which grouped the trees in several statistically-different sets (data not shown). The magnitude of genetic control over germination parameters, estimated by broad sense heritability h2, ranged from 0.35 to
0.82 (strati®ed) and from 0.58 to 0.73 (unstrati®ed) for Sooke and, similarly from 0.30 to 0.85 (strati®ed) and from 0.45 to 0.84 (unstrati®ed) for San Juan (Table 2(b) and (c)), indicating that genetic differences exerted a major in¯uence on germination parameters. These results are supported by other investigations that have shown that germination parameters of other coniferous seeds are under strong genetic control also (Bramlett et al., 1983; Hoff, 1987; Chaisurisri et al., 1993; El-Kassaby et al., 1992, 1993; Davidson et al., 1995).
Fig. 2. Germination course of stratified (S) and unstratified (US) seeds from 20 trees at San Juan.
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
3.2. Simulated aging Accelerated aging produced a highly signi®cant (P<0.01) effect on all germination parameters and accounted for between 97.61% to 99.86% of the total variation observed (Table 3). The effect of aging on germination rate (R50) resulted in less than 50% germination in many cases, thus the R50 parameter was inestimable (Thomson and El-Kassaby, 1993). Seed pretreatment (strati®cation) produced a small, but detectable (P<0.05) effect on germination speed (R50'), accounting for 1.17% of the total variation in this parameter and indicating that pretreatment had enhanced germination speed by overcoming seed dormancy. The effect of seed pretreatment on the remaining germination parameters was almost nondetectable (Table 3). The other sources of variation, namely, seed source and all the interaction terms, accounted for almost no detectable effect on variation in germination (Table 3), indicating uniformity of response among the seedlots to aging treatments. The effect of accelerated aging on seed viability (expressed as mean GC) of all eight seedlots was similar for strati®ed and unstrati®ed seeds (Fig. 3). For strati®ed seeds, average viability decreased from 88% before aging, to 74%, 41%, and 2% after 3,6, and 9 days of aging, respectively (Fig. 3, solid line). In unstrati®ed seeds, over the same aging periods, average viability decreased from 91% to 77%, 40%, and
209
2% (Fig. 3, solid line). On average, strati®ed seeds lost all viability when aged for 12 days, while unstrati®ed seeds retained some viability through 18 days (Fig. 3). Initial viability among seedlots before aging ranged from 76% to 96% and 78% to 95% for strati®ed and unstrati®ed seeds, respectively. Aging treatment caused a broadening in this range after 3 and 6 days indicating differences among seedlots in their propensities to loss of viability. For example, in strati®ed seeds the range of viability increased to 50% to 94% and to 14% to 67% after 3 and 6 days of aging, respectively (Fig. 3, dotted lines). Similarly, in unstrati®ed seeds these ranges increased to 56% to 96% and 25% to 66% after the same aging treatments (Fig. 3, dotted lines). Thus, based on GC, these eight seedlots showed the most variation when aged for 6 days. Aging longer than 6 days substantially reduced this variation indicating that seed viability was in rapid decline. These results are similar to those reported for other conifers (Pitel, 1980; Blanche et al., 1988, 1990; Marquez-Millano et al., 1991; Chaisurisri et al., 1993). This study was not designed to correlate the duration of simulated aging treatment with the period of traditional long-term seed storage and their affects on viability retention. For this reason it cannot be determined how 6 days (or any other duration) of aging relates to a given period (years) of traditional storage. However, the study did demonstrate that mountain hemlock seedlots differ in their ability to resist stress,
Fig. 3. Average germination capacity (GC) (solid line) of stratified seeds (S) and unstratified seeds (US) of eight mountain hemlock populations subjected to accelerated aging treatments for 0; 3; ; 21 days (maximum and minimum are presented with dotted lines).
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Table 4 Germination capacity (GC) after 0 (initial) and 6 days accelerated aging for eight mountain hemlock seedlots Seedlot #
03854 07727 07934 09783 09960 10904 13877 35050
Stratified
Unstratified
Initial GC
6 days GC
% viability
Initial GC
6 days GC
% viability
77 85 88 97 95 82 93 91
15 54 18 29 52 34 64 68
19 64 20 30 55 42 69 75
79 96 93 94 94 88 95 93
26 46 28 30 34 31 64 67
33 48 30 32 36 35 67 72
`% viability' indicates GC after 6 days aging as percent of `initial' (minimum and maximum are bolded).
and that these differences can be readily detected. In the short term, for seeds aged for 6 days, the differential viability ranged between 19% and 75% and between 30% and 72% for strati®ed (seedlots 3854 and 35050) and unstrati®ed seeds (seedlots 7934 and 35050), respectively (Table 4). The differential losses in viability among various seedlots under short-term stressful environments suggest that similar losses can be expected over long-term storage. This means that the genetic contribution of seedlots is likely to change as seeds age during long-term storage, thus the reliance on seed banks for ex-situ conservation of the species genetic resources requires further evaluation. Acknowledgements This study was funded by Forest Renewal British Columbia Grant number HQ96059-RE. The technical assistance of L.M. El-Kassaby is highly appreciated. References Association of Official Seed Analysts. 1983. Seed vigor testing handbook. Assoc. Offic. Seed Anal., Contribution 32, p. 88. Baskin, C.C., 1977. Vigor test methods ± accelerated aging. Assoc. Offic. Seed Anal. Newslett. 51, 42±52. Blanche, C.A., Elam, W.W., Hodges, J.D., 1990. Accelerated aging of Quercus nigra seed: biochemical changes and applicability as a vigor test. Can J. For. Res. 20, 1611±1615. Blanche, C.A., Elam, W.A., Hodges, J.D., Bonner, F.T., Marquez, A.C., 1988. Accelerated aging of selected tree seeds. In: Worral, J., Loo-Dinkins, J., Lester, D.P. (Eds.), Proc. 10th North American Forest Biology Workshop `Physiology and genetics of reforestation'. University of British Columbia, Vancouver, BC, pp. 327±334.
Bramlett, D.L., Dell, T.R., Pepper, W.D., 1983. Genetic and maternal influences on Virgina pine seed germination. Silv. Genet. 32, 1±2. Chaisurisri, K., Edwards, D.G.W., El-Kassaby, Y.A., 1993. Accelerated aging of Sitka spruce seeds. Silv. Genet. 42, 303±308. Czabator, F.J., 1962. Germination value: an index combining speed and completeness of pine seed germination. For. Sci. 8, 386± 396. Davidson, R.H., Edwards, D.G.W., Sziklai, O., El-Kassaby, Y.A., 1995. Genetic variation in germination parameters among Pacific silver fir populations. Silv Genet. 45, 165±171. Delouche, J.C., Baskin, C.C., 1973. Accelerated aging techniques for predicting the relative storability of seed lots. Seed Sci Technol. 1, 427±452. Dickson, M.H., 1980. Genetic aspects of seed quality. Hortsci. 15, 771±774. Edwards, D.G.W., 1980. Maturity and quality of tree seeds ± a state-of-the-art review. Seed Sci. Technol. 8, 625±657. Edwards, D.G.W., El-Kassaby, Y.A., 1996. The effect of stratification and artificial light on the germination of mountain hemlock seeds. Seed Sci. Techol. 24, 225±235. El-Kassaby, Y.A., Edwards, D.G.W., Taylor, D.W., 1992. Genetic control of germination parameters in Douglas-fir and its importance for domestication. Silv. Genet. 41, 48± 54. El-Kassaby, Y.A., Chaisurisri, K., Edwards, D.G.W., Taylor, D.W., 1993. Genetic control of germination parameters of Douglasfir, Sitka spruce, western redcedar and yellow-cedar and its impact on container nursery production. In: Edwards, D.G.W. (Ed.), Dormancy and Barriers to Germination, Proc. Internat. Symposium on Seed Problems, IUFRO Project Group P2.0400. Victoria, B.C., April 1991, Forestry Canada, Pacific For. Cent.: pp. 37±42. Farjon, A., Page, C.N., Schellevis, N., 1993. A preliminary world list of threatened conifer taxa. Biodiversity and Conservation 2, 304±326. Harestad, A.S., 1980. Seasonal movement of black-tailed deer on northern Vancouver Island. Dissert Abstr. Internal. 40, 5088± 5089B.
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211 Hoff, R.J., 1987. Dormancy in Pinus monticola seed related to stratification time, seed coat, and genetics. Can. J. For. Res. 17, 94±298. International Seed Testing Association, 1993. International rules for seed testing. Seed Sci. Technol. 21 (supplement), 284. Kneebone, W.R., 1976. Some genetic aspects of seed vigor. J. Seed Technol. 1, 86±97. Kueneman, E.A., 1983. Genetic control of seed longevity in soy beans. Crop Sci. 23, 5±8. Marquez-Millano, A., Elam, W.A., Blanche, C.A., 1991. Influence of accelerated aging on fatty acid composition of slash pine (Pinus elliottii Engelm. var. elliottii) seeds. J. Seed Technol. 15, 29±41. McDaniel, R.G., 1973. Genetic factors influencing seed vigor; biochemistry of heterosis. Seed Sci. Technol. 1, 25±50.
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Parsons, D.J., 1972. The southern extensions of Tsuga mertensiana (mountain hemlock) in the Sierra nevada. MadronÄo 21, 536± 539. Pitel, J.A., 1980. Accelerated aging studies of seeds of jack pine (Pinus banksiana Lamb.) and red oak (Quercus rubra L.). In: Wang, B.S.P., Pitel, J.A. (Eds.), Proc. internat. Symp. `Fores tree seed storage', IUFRO Working party S2.01.06 `Seed Problems', Petawawa Nat. For. Inst., Chalk River, Ont., Canada. Environ. Can., Can. For. Serv., pp. 40±54. Steel, R.D.G., Torrie, J.H., 1980. Principles and Procedures of Statistics; a Biometrical Approach, 2nd. ed. McGraw-Hill, New York, p. 633. Thomson, A.J., El-Kassaby, Y.A., 1993. Interpretation of seed germination parameters. New Forests 7, 123±132.
Genetic control of germination and the effects of accelerated aging in mountain hemlock seeds and its relevance to gene conservation Y.A. El-Kassabya,*, D.G.W. Edwardsb a
Department of Forest Sciences, Faculty of Forestry, The University of British Columbia, Vancouver, BC, Canada V6T 1Z4 b Canadian Forest Service, Paci®c Forestry Centre, 506 West Burnside Road, Victoria, BC, Canada V8Z 1M5 Accepted 25 May 1998
Abstract Genetic control of germination parameters and the effects of accelerated aging in mountain hemlock (Tsuga mertensiana [Bong.] Carr.) seeds were investigated using standard germination tests and simulated aging, respectively. Germination parameters were studied on seeds collected from individual trees from two natural populations (Sooke and San Juan) located on southern Vancouver Island, BC. Strong genetic control was con®rmed by the high heritability estimates that ranged from 0.35 to 0.82 (strati®ed) and from 0.58 to 0.73 (unstrati®ed) for Sooke and from 0.30 to 0.85 (strati®ed) and from 0.45 to 0.84 (unstrati®ed) for San Juan. Simulated aging was conducted on eight seedlots collected from natural stands representing the species distribution in British Columbia. Strati®ed and unstrati®ed seeds were aged at 100% RH and 37.58C for 0 to 21 days at 3-day intervals. Seed aging accounted for the majority of variation which ranged from 97.61% to 99.86% of the total variation observed. Gradual loss of seed viability was observed over the aging treatments with a total loss of viability after aging for 12 days. Differential losses in viability among seedlots were maximum after aging for 6 days. Results are discussed in terms of the impact of long-term storage and the reliance on seed banks for ex-situ conservation of this species. # 1998 Elsevier Science B.V. All rights reserved. Keywords: Tsuga mertensiana; Seed germination; Heritability; Gene conservation
1. Introduction Mountain hemlock (Tsuga mertensiana) is a longlived, subalpine conifer that is most abundant in coastal regions of Alaska and British Columbia. Its range extends to central California (Parsons, 1972), with isolated populations in the interior of British Columbia, Idaho and Montana. In general, its populations are disjunct due the physical separation of the *Corresponding author. Tel.: +1-(250)-381-1404; fax: +1-(250)381-0252; e-mail: [email protected]
high elevation sites where they occur. It is a valuable species for watershed protection and wildlife habitat (Harestad, 1980). Timber harvesting in British Columbia is reaching higher into subalpine regions, but very little is known about the genetic diversity and regeneration potential/success of the species. Due to the disjunct nature of its distribution, mountain hemlock was included in a world list of threatened species (Farjon et al., 1993). This study is part of a larger project designed to investigate the genetic structure of mountain hemlock using gene markers, adaptive and quantitative
0378-1127/98/$ ± see front matter # 1998 Elsevier Science B.V. All rights reserved. PII: S0378-1127(98)00343-0
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meters in mountain hemlock, and (2) how the genetic representation of the species will be affected by simulated long term storage and its relation to ex-situ gene conservation.
attributes and regeneration (germination) ecology for the purpose of developing a suitable conservation strategy. This can be accomplished by a combination of in-situ and ex-situ approaches. Although seed banks have been proposed as a viable means of ex-situ conservation, a thorough understanding of seed biology (germination and storage) is imperative in developing effective conservation tools. Standard germination tests provide estimates of the maximal potential of a seedlot for seedling production under favorable environmental conditions. These tests do not evaluate seedlot vigour, that is, the ability to produce seedlings under stressful conditions (Association of Of®cial Seed Analysts, 1983). Seed viability is highest at physiological maturity and declines with age (Edwards, 1980), the deterioration being gradual and ending with total loss of germinability and vigour (Delouche and Baskin, 1973). Seed vigour is known to be heritable (McDaniel, 1973; Kneebone, 1976; Dickson, 1980) and may be inherited maternally in some plants (Kueneman, 1983). Degree of seed deterioration/vigour can be revealed through a stress test, such as the accelerated aging test that simulates long-term storage (Delouche and Baskin, 1973; Baskin, 1977; Association of Of®cial Seed Analysts, 1983), which has been used successfully in several tree studies to evaluate the ef®cacy of ex-situ conservation (Pitel, 1980; Blanche et al., 1988, 1990; Marquez-Millano et al., 1991; Chaisurisri et al., 1993). It is important to determine whether this deterioration is independent among seedlots (i.e., genetic) and/or whether seedlots exhibit the same rate of deterioration over time. The objectives of this study were to determine: (1) the degree of genetic control of germination para-
2. Materials and methods 2.1. Seed source Seeds from individual trees in two natural stands of mountain hemlock on Vancouver Island were used to investigate the genetic control of germination parameters. The stands are located on TimberWest Ltd. (formerly Paci®c Forest Products Ltd.) land on the southern tip of Vancouver Island at Sooke (13 trees) and San Juan (20 trees). Eight natural stand bulk seedlots (Table 1) were obtained from the British Columbia Ministry of Forests' Tree Seed Centre for the simulated aging study. Seven of these seedlots were from coastal locations, while the Sale Mt. seedlot was included as a representative of an interior (more continental) seed source. 2.2. Germination Germination tests for individual trees within each population (Sooke and San Juan) were conducted on four replications (100 seeds each) of strati®ed and unstrati®ed seeds. Seeds were spread in 10104 cm, tightly-lidded clear-plastic boxes lined with one layer of Kimpak (cellulose wadding) overlain with three layers of Whatman no. 1 ®lter paper, moistened with 50 ml of distilled water. Boxes were placed in
Table 1 Sources of the eight mountain hemlock natural stand seedlots used in the germination study Seedlot no.a
Local source name
Year collected
Elevation (m)
Lat. (N)
Long. (W)
1000 PSWb (g)
03854 07727 07934 09783 09960 10904 13877 35050
Hoodoo Creek Garbage Creek Hkusam Mt. Kearsley Creek Port Alice Sale Mt. Lyon Lake Hanna Ridge
1979 1982 1982 1982 1982 1988 1982 1990
1250 850 950 1280 750 1700 1005 700
51820' 488 33' 50820' 49819' 50824' 51810' 49839' 56818'
125832' 124806 125850' 122822' 127827' 118810' 123854' 129820'
2493 2193 2283 2059 2387 2293 2323 2003
a b
British Columbia Ministry of Forests seedlot registration numbers. Estimated weight of 1000 pure seeds based on eight replications of 100 filled seeds (Edwards and El-Kassaby, 1996).
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
randomly-assigned positions in an incubator set for a constant 208C. Light (cool-white ¯uorescent) of approximately 100 lux at the ®lter paper surface was provided as a photoperiod of 8 h `day' and 16 h (no light) `night'. Seeds were counted as germinated when the radicle and hypocotyl equalled or exceeded four times the length of the individual seed. Germinants were assessed at 2-day intervals for the ®rst half of each test, then at 3±4 day intervals up to 28 days, and were classed as normal or abnormal according to the International Seed Testing Association, 1993. 2.3. Accelerated aging Based on a pilot study, it was determined that 37.58C was the optimal temperature for accelerated aging of mountain hemlock seeds. At higher temperature (408C and above), the decline in germination occurred too rapidly to clearly delineate differences among seedlots, while lower temperatures required very prolonged aging treatment to obtain complete loss of germinability. This temperature, 37.58C, is lower than that prescribed for agricultural seeds (Association of Of®cial Seed Analysts, 1983), but agrees with that recommended for tree seeds in other studies (Blanche et al., 1988; Chaisurisri et al., 1993). Accelerated aging was applied for eight periods from 0 to 21 days at 3-day intervals (i.e., 0, 3, , 21 days). For each aging period, eight replications of 100 seeds were placed on wire-mesh platforms in tightly-closed, clear-plastic germination boxes. Each box contained 50 ml of distilled water; the distance between the platform and the water surface was 1.5 cm. Thus, for every seedlot a total of 64 germination boxes containing 100 seeds each were used. For every seedlot and every aging period, the accelerated aging tests were conducted on four replications of strati®ed and unstrati®ed seeds. Prior to aging, four of the eight replications for each seedlot were imbibed for 24 h, drained, placed in plastic bags and strati®ed at 28C for 28 days. The remaining four replications were refrigerated (18C, dry) until required for the aging treatments. Aging and strati®cation treatments were scheduled so that all subsequent germination tests began at the same time. Thus, strati®ed and unstrati®ed samples that received the 21-day aging treatment were placed in the incu-
205
bator ®rst, and those to receive the 0-day treatment were last. Following aging, all samples were rinsed under running cold water for 1 to 2 min to reduce fungal mycelial growth. Germination tests for the aging trial were the same as described above. 2.4. Data analysis Results were expressed as: (i) germination capacity (GC), the percentage of seeds that had germinated normally at the end of the test; (ii) germination rate (R50), the number of days required for 50% of the seeds in the replication to germinate; (iii) germination speed (R50 '), the number of days for 50% of the germinating seeds to germinate (Thomson and ElKassaby, 1993); (iv) peak value (PV), an index of germination speed that expresses germination rate as the maximal quotient derived by dividing daily the accumulated number of normal germinants by the corresponding number of days (Czabator, 1962); this is the mean daily germination of the most-vigorous components of a seedlot, and a mathematical expression of the tangent drawn through the origin of a sigmoid curve representing the typical course of germination. And (v) germination value (GV), which combines germination capacity and speed into a single index (Czabator, 1962). Data were transformed to normalize the calculated response variables, and to achieve homogeneity of variances, as follows: GC by arcsin, R50 and R50' by (1-[1/x1]), and GV by (H[x0.5]), where x is the variable in question; PV was not transformed. The transformed data were analyzed using ANOVA based on the following two linear models: Yijk Ti Sj TSij " ijk
(1)
where overall mean, Tithe effect of the ith tree, i1 to 13 (Sooke) or 1 to 20 (San Jaun)(random), Sjthe effect of the jth seed pretreatment (strati®cation), j1 to 2 (®xed effect), TSijthe effect of the interaction between tree and seed pretreatment, "(ij)kthe residual term, k1 to 4 for germination tests. Yijkl Pi Sj Ak PSij SAjk PSAijk " ijkl
(2)
where overall mean, Pithe effect of the ith pretreatment (strati®cation), i1 to 2 (®xed effect),
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Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
Sjthe effect of the jth seed source, j1 to 8 (random effect), Akthe effect of the kth aging treatment, k1 to 8 (®xed effect), PSikthe effect of the interaction between pretreatment and seed source, PAikthe effect of the interaction between pretreatment and aging, SAjkthe effect of the interaction between seed source and aging treatment, PSAijkthe effect of the interaction between pretreatment (strati®cation), seed source and aging treatment, "(ijk)lthe residual term, l1 to 4 for accelerated aging tests. Expected mean squares (EMS) were calculated and % variance components estimated for each source of variation. Where signi®cant effects were observed, means were compared using the Student±Newman± Keuls range test (Steel and Torrie, 1980). Sources of variation and EMS are shown in Tables 2 and 3. 3. Results and discussion 3.1. Genetic control of germination parameters The germination data were analysed on an individual-population basis due to the lack of data con®rming that the allelic frequencies of genes controlling such attributes are uniform over these two populations. Therefore, heritability estimates were calculated for Sooke and San Juan separately. For both populations, the effects of seed pretreatment (P), expressed as a percentage of the total variation, on germination
parameters were highly signi®cant (P0.01), and accounted for between 20 (GC) to 79 (R50), and between 18 (GC) to 86 (R50') at Sooke and San Juan, respectively (Table 2(a)). Germination capacity ranged from 91% (unstrati®ed) to 97% (strati®ed) at Sooke (Fig. 1), and from 95% (unstrati®ed) to 97% (strati®ed) at San Juan (Fig. 2), while germination rate varied between 14 (unstrati®ed) and 17 (strati®ed) days to reach 50% germination in both populations. At Sooke, when seeds were strati®ed (Fig. 1), variation in germination was reduced (range: 94.6±99.6% vs. 76.9±100%), whereas at San Juan the variation was greatly reduced in 19 out of the 20 trees (range: 95.6± 99.6% vs. 88.7±98.6%) (Fig. 2). Tree number 7 at San Juan showed little response to the seed pretreatment indicating greater variability in seed dormancy among trees in this population. The observed differences between strati®cation treatments created strong polarization in the data, making it dif®cult to discern the true differences among trees. Therefore, complementary analyses were conducted for each seed pretreatment separately (Table 2(b and c)). When seed pretreatment was excluded from the analysis, a shift in the amount of variation was observed and differences among trees within populations became highly signi®cant for all germination parameters. At Sooke, the effects of trees (T), expressed as a percentage of the total variation, on germination parameters ranged from 35 (GC) to 82 (R50'), and from 58 (PV) to 73 (R50'), for strati®ed and
Fig. 1. Germination course of stratified (S) and unstratified (US) seeds from 13 trees and Sooke.
p-1 t-1 (p-1)(t-1) pt(r-1)
d.f.
12 36
19 57
a-Combined analyses Seed pretreatment (P) Tree (T) PT Residual
S.O.V.
b- Sooke. Among trees (B) Within (W) h2
c-San Juan. Among trees (B) Within (W) h2
PV
GV
2W 42B 2W
2W 42B 2W
84.76 15.24 0.85
77.78 22.12 0.78 81.82 18.18 0.82
81.61 18.39 0.82
R'50
29.95 70.05 0.30
34.83 65.17 0.35
GC
83.72 16.28 0.84
64.05 35.95 0.64
PV
80.14 19.86 0.80
60.01 39.99 0.60
GV
81.48 18.52 0.81
65.71 34.29 0.66
84.00 16.00 0.84
73.33 26.67 0.73
R'50
R50
85.9 00.0 06.8 07.4
R50
64.5 25.1 04.2 06.2 Unstratified
64.5 15.2 06.9 13.4
Stratified
61.6 14.0 10.7 13.8
E.M.S3
19.9 14.9 37.3 27.8
R'50
78.8 14.2 01.6 05.4
73.7 18.9 01.5 05.9
R50
GC
R50 R'50
San Juan
Sooke
2e k1 2PT k2 2P 2e k1 2PT k3 2T 2e k1 2PT 2e
E.M.S.2
44.79 55.21 0.45
72.98 27.02 0.73
GC
18.0 20.6 10.3 51.0
GC
79.44 20.56 0.79
58.41 41.59 0.58
PV
25.4 19.2 00.0 55.4
PV
77.11 22.89 0.77
67.60 32.40 0.68
GV
52.4 35.4 03.7 08.4
GV
Germination parameters are (i) germination rate (R50), (ii) germination speed (R50'), p (iii) germination capacity (GC), (iv)germination value (GV), and (v) peak value (PV). Data transformations are as follows: R50 and R50' by (1ÿ[1/x1]), GC by arcsin, GV by x ÿ 0:5, and PV was not transformed. 2 Coefficients of variance components are: Sooke (k14, k28, k352) and San Jaun (k14, k28, k380), 2p variance due to pretreatment, 2T variance due to tree, 2PT variance due to pretreatment tree interaction. 3 2 B variance among trees and 2W variance among replications within tree.
1
d.f.
S.O.V
Table 2 Estimation of variance components (as percentages of the total variation) and broad-sense heritabilities (h2) for germination parameters1 of 2 mountain hemlock populations (Sooke and San Juan).
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Table 3 ANOVA and percent contribution of the variance components (Var. Comp.) for the germination parametersa of eight mountain hemlock seedlots following seed pretreatment and accelerated aging S.O.V.
d.f.
E.M.S.b
Var. Comp.
R'50
GC
GV
PV
Seed pretreatment (P) Seedlot (S) Aging (A) PS P A SA PSA Residual
1 7 7 7 7 49 49 384
2e 3:992PSA 31:972PS p 2e 3:992PSA 7:992AS 31:972PS 63:932S 2e 3:992PSA 7:992SA A 2e 3:992PSA 31:972PS 2e 3:992PSA PA 2e 3:992PSA 7:992SA 2e 3:992PSA 2e
P 2S A 2PS PA 2SA 2PSA 2e
1.17 0.00 97.61 0.01 0.68 0.17 0.00 0.48
0.11 0.03 99.58 0.00 0.06 0.08 0.00 0.16
0.00 0.04 99.86 0.00 0.02 0.16 0.00 0.18
0.00 0.03 99.81 0.00 0.13 0.09 0.00 0.16
a
Germination parameters are (i) germination speed (R50'), (ii) germination capacity (GC), (iii) germination value (GV), and (iv) peak value (PV). Data transformation are as follows: R50' by (1-[1/x1]), GC by arcsin, GV by (Hx0.5), and PV is not transformed. b Pvariance due to seed pretreatment (fixed), 2S variance due to seedlot effect (random), Avariance due to aging treatment (fixed), 2PS variance due to seed pretreatment seedlot interaction, PAvariance due to seed pretreatment aging treatment interaction, 2SA variance due to seedlot aging treatment interaction, 2PSA variance due to seed pretreatment seedlot aging treatment interaction, 2e residual effect.
unstrati®ed seeds, respectively (Table 2(b)). A similar trend was observed at San Juan, and the effect of trees (T) as a percent of total variation ranged from 30 (GC) to 85 (R50), and from 45 (GC) to 84 (R50'), for strati®ed and unstrati®ed seeds, respectively (Table 2(c)). These differences were further emphasized by the Student±Newman±Keuls range tests which grouped the trees in several statistically-different sets (data not shown). The magnitude of genetic control over germination parameters, estimated by broad sense heritability h2, ranged from 0.35 to
0.82 (strati®ed) and from 0.58 to 0.73 (unstrati®ed) for Sooke and, similarly from 0.30 to 0.85 (strati®ed) and from 0.45 to 0.84 (unstrati®ed) for San Juan (Table 2(b) and (c)), indicating that genetic differences exerted a major in¯uence on germination parameters. These results are supported by other investigations that have shown that germination parameters of other coniferous seeds are under strong genetic control also (Bramlett et al., 1983; Hoff, 1987; Chaisurisri et al., 1993; El-Kassaby et al., 1992, 1993; Davidson et al., 1995).
Fig. 2. Germination course of stratified (S) and unstratified (US) seeds from 20 trees at San Juan.
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211
3.2. Simulated aging Accelerated aging produced a highly signi®cant (P<0.01) effect on all germination parameters and accounted for between 97.61% to 99.86% of the total variation observed (Table 3). The effect of aging on germination rate (R50) resulted in less than 50% germination in many cases, thus the R50 parameter was inestimable (Thomson and El-Kassaby, 1993). Seed pretreatment (strati®cation) produced a small, but detectable (P<0.05) effect on germination speed (R50'), accounting for 1.17% of the total variation in this parameter and indicating that pretreatment had enhanced germination speed by overcoming seed dormancy. The effect of seed pretreatment on the remaining germination parameters was almost nondetectable (Table 3). The other sources of variation, namely, seed source and all the interaction terms, accounted for almost no detectable effect on variation in germination (Table 3), indicating uniformity of response among the seedlots to aging treatments. The effect of accelerated aging on seed viability (expressed as mean GC) of all eight seedlots was similar for strati®ed and unstrati®ed seeds (Fig. 3). For strati®ed seeds, average viability decreased from 88% before aging, to 74%, 41%, and 2% after 3,6, and 9 days of aging, respectively (Fig. 3, solid line). In unstrati®ed seeds, over the same aging periods, average viability decreased from 91% to 77%, 40%, and
209
2% (Fig. 3, solid line). On average, strati®ed seeds lost all viability when aged for 12 days, while unstrati®ed seeds retained some viability through 18 days (Fig. 3). Initial viability among seedlots before aging ranged from 76% to 96% and 78% to 95% for strati®ed and unstrati®ed seeds, respectively. Aging treatment caused a broadening in this range after 3 and 6 days indicating differences among seedlots in their propensities to loss of viability. For example, in strati®ed seeds the range of viability increased to 50% to 94% and to 14% to 67% after 3 and 6 days of aging, respectively (Fig. 3, dotted lines). Similarly, in unstrati®ed seeds these ranges increased to 56% to 96% and 25% to 66% after the same aging treatments (Fig. 3, dotted lines). Thus, based on GC, these eight seedlots showed the most variation when aged for 6 days. Aging longer than 6 days substantially reduced this variation indicating that seed viability was in rapid decline. These results are similar to those reported for other conifers (Pitel, 1980; Blanche et al., 1988, 1990; Marquez-Millano et al., 1991; Chaisurisri et al., 1993). This study was not designed to correlate the duration of simulated aging treatment with the period of traditional long-term seed storage and their affects on viability retention. For this reason it cannot be determined how 6 days (or any other duration) of aging relates to a given period (years) of traditional storage. However, the study did demonstrate that mountain hemlock seedlots differ in their ability to resist stress,
Fig. 3. Average germination capacity (GC) (solid line) of stratified seeds (S) and unstratified seeds (US) of eight mountain hemlock populations subjected to accelerated aging treatments for 0; 3; ; 21 days (maximum and minimum are presented with dotted lines).
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Table 4 Germination capacity (GC) after 0 (initial) and 6 days accelerated aging for eight mountain hemlock seedlots Seedlot #
03854 07727 07934 09783 09960 10904 13877 35050
Stratified
Unstratified
Initial GC
6 days GC
% viability
Initial GC
6 days GC
% viability
77 85 88 97 95 82 93 91
15 54 18 29 52 34 64 68
19 64 20 30 55 42 69 75
79 96 93 94 94 88 95 93
26 46 28 30 34 31 64 67
33 48 30 32 36 35 67 72
`% viability' indicates GC after 6 days aging as percent of `initial' (minimum and maximum are bolded).
and that these differences can be readily detected. In the short term, for seeds aged for 6 days, the differential viability ranged between 19% and 75% and between 30% and 72% for strati®ed (seedlots 3854 and 35050) and unstrati®ed seeds (seedlots 7934 and 35050), respectively (Table 4). The differential losses in viability among various seedlots under short-term stressful environments suggest that similar losses can be expected over long-term storage. This means that the genetic contribution of seedlots is likely to change as seeds age during long-term storage, thus the reliance on seed banks for ex-situ conservation of the species genetic resources requires further evaluation. Acknowledgements This study was funded by Forest Renewal British Columbia Grant number HQ96059-RE. The technical assistance of L.M. El-Kassaby is highly appreciated. References Association of Official Seed Analysts. 1983. Seed vigor testing handbook. Assoc. Offic. Seed Anal., Contribution 32, p. 88. Baskin, C.C., 1977. Vigor test methods ± accelerated aging. Assoc. Offic. Seed Anal. Newslett. 51, 42±52. Blanche, C.A., Elam, W.W., Hodges, J.D., 1990. Accelerated aging of Quercus nigra seed: biochemical changes and applicability as a vigor test. Can J. For. Res. 20, 1611±1615. Blanche, C.A., Elam, W.A., Hodges, J.D., Bonner, F.T., Marquez, A.C., 1988. Accelerated aging of selected tree seeds. In: Worral, J., Loo-Dinkins, J., Lester, D.P. (Eds.), Proc. 10th North American Forest Biology Workshop `Physiology and genetics of reforestation'. University of British Columbia, Vancouver, BC, pp. 327±334.
Bramlett, D.L., Dell, T.R., Pepper, W.D., 1983. Genetic and maternal influences on Virgina pine seed germination. Silv. Genet. 32, 1±2. Chaisurisri, K., Edwards, D.G.W., El-Kassaby, Y.A., 1993. Accelerated aging of Sitka spruce seeds. Silv. Genet. 42, 303±308. Czabator, F.J., 1962. Germination value: an index combining speed and completeness of pine seed germination. For. Sci. 8, 386± 396. Davidson, R.H., Edwards, D.G.W., Sziklai, O., El-Kassaby, Y.A., 1995. Genetic variation in germination parameters among Pacific silver fir populations. Silv Genet. 45, 165±171. Delouche, J.C., Baskin, C.C., 1973. Accelerated aging techniques for predicting the relative storability of seed lots. Seed Sci Technol. 1, 427±452. Dickson, M.H., 1980. Genetic aspects of seed quality. Hortsci. 15, 771±774. Edwards, D.G.W., 1980. Maturity and quality of tree seeds ± a state-of-the-art review. Seed Sci. Technol. 8, 625±657. Edwards, D.G.W., El-Kassaby, Y.A., 1996. The effect of stratification and artificial light on the germination of mountain hemlock seeds. Seed Sci. Techol. 24, 225±235. El-Kassaby, Y.A., Edwards, D.G.W., Taylor, D.W., 1992. Genetic control of germination parameters in Douglas-fir and its importance for domestication. Silv. Genet. 41, 48± 54. El-Kassaby, Y.A., Chaisurisri, K., Edwards, D.G.W., Taylor, D.W., 1993. Genetic control of germination parameters of Douglasfir, Sitka spruce, western redcedar and yellow-cedar and its impact on container nursery production. In: Edwards, D.G.W. (Ed.), Dormancy and Barriers to Germination, Proc. Internat. Symposium on Seed Problems, IUFRO Project Group P2.0400. Victoria, B.C., April 1991, Forestry Canada, Pacific For. Cent.: pp. 37±42. Farjon, A., Page, C.N., Schellevis, N., 1993. A preliminary world list of threatened conifer taxa. Biodiversity and Conservation 2, 304±326. Harestad, A.S., 1980. Seasonal movement of black-tailed deer on northern Vancouver Island. Dissert Abstr. Internal. 40, 5088± 5089B.
Y.A. El-Kassaby, D.G.W. Edwards / Forest Ecology and Management 112 (1998) 203±211 Hoff, R.J., 1987. Dormancy in Pinus monticola seed related to stratification time, seed coat, and genetics. Can. J. For. Res. 17, 94±298. International Seed Testing Association, 1993. International rules for seed testing. Seed Sci. Technol. 21 (supplement), 284. Kneebone, W.R., 1976. Some genetic aspects of seed vigor. J. Seed Technol. 1, 86±97. Kueneman, E.A., 1983. Genetic control of seed longevity in soy beans. Crop Sci. 23, 5±8. Marquez-Millano, A., Elam, W.A., Blanche, C.A., 1991. Influence of accelerated aging on fatty acid composition of slash pine (Pinus elliottii Engelm. var. elliottii) seeds. J. Seed Technol. 15, 29±41. McDaniel, R.G., 1973. Genetic factors influencing seed vigor; biochemistry of heterosis. Seed Sci. Technol. 1, 25±50.
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Parsons, D.J., 1972. The southern extensions of Tsuga mertensiana (mountain hemlock) in the Sierra nevada. MadronÄo 21, 536± 539. Pitel, J.A., 1980. Accelerated aging studies of seeds of jack pine (Pinus banksiana Lamb.) and red oak (Quercus rubra L.). In: Wang, B.S.P., Pitel, J.A. (Eds.), Proc. internat. Symp. `Fores tree seed storage', IUFRO Working party S2.01.06 `Seed Problems', Petawawa Nat. For. Inst., Chalk River, Ont., Canada. Environ. Can., Can. For. Serv., pp. 40±54. Steel, R.D.G., Torrie, J.H., 1980. Principles and Procedures of Statistics; a Biometrical Approach, 2nd. ed. McGraw-Hill, New York, p. 633. Thomson, A.J., El-Kassaby, Y.A., 1993. Interpretation of seed germination parameters. New Forests 7, 123±132.