Ann. Bot. 47, 467-471, 1981
Germination Rate of Douglas-fir [Pseudotsuga
menziesii (Mirb.) Franco] Seeds Affected by Their
Orientation
F. C. SORENSEN and R. K. CAMPBELL
Accepted: 11 September 1980
ABSTRACT
Douglas-fir [Pseudotsuga menziesii (Mirb.) Franco] seeds develop with the adaxial surface continuous with the
cone scale. On separation, the scale side of the seed is lighter coloured and flatter than the side not previously
in contact with the scale. Seeds oriented scale-side-up on a moist surface germinate more rapidly than seeds
placed scale-side-down. When seeds were germinated under laboratory conditions, the response to orientation
varied among seeds of different geographic origin. Most of the geographic variation in germination rate
occurred when seeds were oriented scale-side-up.
Key words: Pseudotsuga menziesii (Mirb.) Franco, Douglas-fir germination, orientation of seeds.
INTRODUCTION
Two types of seed-form variants (seed polymorphisms) have been described: (1) two or
more forms of seeds for a species exist on separate plants growing in the same habitat,
or (2) two or more forms of seeds are produced on different parts of the same plant or
flower (Harper, Lovell and Moore, 1970). The differences in form are adaptive and are
often associated with differences in germination behaviour.
We report here a variation similar to polymorphism in that it is based on differences
in form, but in this case the differences are between the two sides of a single seed. Because
of the differences between the sides, seed orientation affects germination rate. The
differences in germination rate may be adaptive, because response to orientation differs
among seeds of different geographic origins.
The physical basis for the polymorphism resides in the shape and development of a
Douglas-fir [Pseudotsuga menziesii (Mirb.) Franco] seed. A wingless, mature seed is
flattened and ovate, approximately 8 mm x 3 mm x 2 mm. The seed coat is differentiated
into three layers, of which the outer is continuous with the adaxial surface of the
ovuliferous scale. Separation of the seed wing from the scale results from dissolution of
the middle lamella beneath the ovule and seed wing (Allen and Owens, 1972). When the
seed is separated from the scale, the side previously adhering to the scale is lighter in
colour and somewhat flatter than the upper side. Because the seed is somewhat flattened,
it tends to rest on a more or less even surface with the scale side up or down.
Polymorphisms are generally assumed to be adaptive (Harper et al., 1970; Stebbins,
1971; Nobs and Hagar, 1974; McDonough, 1977; Sorensen, 1978), and we hypothesized
that the seed orientation effect on germination might have an adaptive function in
Douglas-fir. This paper reports a preliminary testing of that hypothesis based on seed
collections from several widely separated locations in Oregon.
This article was written and prepared by US Government employees on official time, and it is therefore in
the public domain.
Downloaded from http://aob.oxfordjournals.org/ at DigiTop USDA's Digital Desktop Library on October 11, 2012
U.S. Department of Agriculture, Forest Service, Pacific Northwest Forest and Range Experiment Station,
Portland, Oregon, U.S.A.
m http://aob.oxfordjournals.org/ at DigiTop USDA's Digital Desktop Library on October 11, 2012
1. Analysis of variance of days to 50 per cent germination for Douglas-fir seed samples from two trees at each of seven locations.
Treatments included two light intensities and two seed orientations with three replications. Trees-in-populations are random, other effects
are fixed
TABLE
Sources of variation
Light (L)
Replications-in-L (R)
Orientation (O)
OxL
Provenance (P)
Trees-in-P (T/P)
PxL
d.f.
/-I
T/PxO
PxLxO
T/P x L x 0
Residual
Total
<rj+ oprto\
/(r-1)
0-1
(T^ + r/(TQ^>(pj -f- '*O*7"LT(P) "t"'*'^o*LT(P) ~^~ rp'°LO"t~ ^ipt<T{)
(/-lXo-1)
0-^ + rlfrjy^i pj + roo^np\ -f- rlofT^ip) 4- rpto^Q
p-\
p(t— 1)
(p— 1X/— l)
a-l + r/oo-^p, + rloto\,
T/PxL
PxO
Estimated mean squares
o^ + r/oo-^p)
o\ + rtxr^-piP) ~t" rot(j\jp
&b ~^~ FOG LT( P)
(p-lXo-l)
Pit-m>-\)
(p— 1X/— 1X°— •)
p(,_l)/_l)( 0 _l)
Kppt-\\r-X)
loprt - 1
(T^ + r/o^yi-^pj
° i + ro1x>T( p> + ' • ' " l o p
^E + ^loTIP)
Calculated
mean squares
(xlO«)
Significance
45113
<001
3
0023
149-823
0066
50-699
14-246
2-299
2-417
11-624
0-776
1-269
0-517
0-242
<0.05
n.s.
n.s.
< 001
n.s.
<001
<001
<001
n.s.
n.s.
§
I3
Sorensen and Campbell—Germination of Douglas-fir Seeds
469
MATERIAL AND METHODS
RESULTS AND DISCUSSION
Seeds placed scale-side-up reached 50 per cent germination in 4-6 days (average for all
treatments), or 1-7 days earlier than those placed scale-side-down. It should be noted
that this difference occurred at temperatures promoting rapid germination. In addition,
the effect of orientation depended on population and trees-in-population, both interactions being highly significant (Table 2).
TABLE 2. Components of variance (cr x 103) and standard errors (in parentheses) estimated
from analyses of germination rates (days'1). Estimates are for seed samples germinated
scale-side-up vs. scale-side-down. Percentages refer to total variability among means
exclusive of light and replication effects
Components of variance
Sources of variation
Populations (P)
Trees-in-populations (T/P)
Error
Total
Estimated mean squares
<J\ + (XT\IV+\2<T\.
<TI + (XJ\IT
a\
Scale-side-up
(<j' x 101) (%)
Scale-side-down
3-92(2-3) 72
1-30(0-6) 24
0-21
4
5-43
100
0-05(0-4) 3
1-07(0-5) 76
0-30
21
1-42
100
Downloaded from http://aob.oxfordjournals.org/ at DigiTop USDA's Digital Desktop Library on October 11, 2012
Seeds were collected from two trees at each of seven locations along a west-to-east
transect at approximately 45° N latitude. The sites included three locations in the Coast
Ranges, two in the Cascade Range, and two in central Oregon east of the Cascades.
Elevations ranged from 45 to 1750 mm, and distances from the Pacific Ocean ranged from
20 to 420 km.
Seeds were soaked in distilled water for 24 h at room temperature (c. 22 °C) and were
moist chilled for 60 days at 3-4 °C. Germination was on moist filter paper in covered
Petri dishes at 18 °C in a controlled environment chamber. Moisture in the Petri dishes
was kept below the level at which a visible film could be seen on the seed coats. Light
was supplied by four cool white fluorescent plus two incandescent lamps (c. 400 fc =
4300 lx).
Because the two sides of the seed differ in colour and do not transmit light equally
(Richardson, 1959), we anticipated that light adsorption might be critical to the
orientation effect and included a light vs. dark contrast in the test. Seeds in the dark
treatment were kept in a box lined with black polyethlene, and their germination was
counted under a single cool white fluorescent lamp filtered through two sheets of 7-mil
(018 mm) medium green roscolene (plastic sheeting).
The design model included a pair of experiments (Snedecor and Cochran, 1967, p. 375),
one in light and one in dark, in a single environment chamber. Seed orientations and
seed samples, replicated three times, formed a 14 x 2 factorial within each experiment.
Seed samples (trees) were nested within populations. The combined analysis of variance ,
was based on a mixed model with random trees-in-populations and other variables fixed
(Table 1).
The unit of observation was the average germination rate of 35 seeds in a single Petri
dish. Mean germination rates, defined at days to 50 per cent total germination (Bliss,
1967, p. 116), were determined for each dish by plotting cumulative percentage
germination against days"1 on probit paper (Campbell and Sorensen, in preparation) and
visually fitting a straight line to the plot. Counts were made at 0025-day"1 intervals
starting at the time of first germination.
470
Sorensen and Campbell—Germination of Douglas-fir Seeds
The evidence for an adaptive basis for the variation pattern comes from the observation
that both mean germination rate (day"1) for a provenance and the difference between
germination rate scale-side-up and scale-side-down increased with distances from the
ocean (Table 3, lines 1 and 2, respectively). In addition, the differences between
germination rates scale-side-up and -down were very closely related to mean germination
rate for the provenance (Table 3, line 3). In other words, provenances whose seeds had
the most rapid germination rates also showed the greatest response to orientation, and
both germination rate and response to orientation increased with increasing distance from
the ocean. Length of frost-free season and mean annual precipitation decrease regularly
with increasing distance from the ocean in the area sampled (Johnsgard, 1963).
Comparison
1
2
3
0-00026
0-00023
0-94
0000085
0000091
008
0-81
0-75
0-98
< 005
=005
< 001
• Probability that the coefficient of correlation differs significantly from zero.
As a preliminary hypothesis, we propose that the orientation response has evolved as
a strategy for regulating variation in timing of germination. In our experiment, the
orientation effect was greater in faster-germinating populations (Table 3, line 3). Seeds
of populations of Douglas fir from habitats with short growing seasons characteristically
germinate rapidly. In regions where rapid or early germination is advantageous, a factor
which delays or spreads out germination also may be advantageous. In such a case, not
all seeds would germinate early if the environmental cues influencing germination were,
by chance, mistimed with respect to frost or other damaging event. Furthermore,
selection against seeds germinating at a particular time would be based partly on a
non-genetic factor (the orientation of the seed). Therefore, genetic variance of germination
rate would not be so greatly altered by the selection associated with a mistimed climatic
event.
We do not know the mechanism by which orientation affects germination rate. Light,
seed shape, moisture uptake and gas exchange and chemistry of the seed coat influence
activation of enzymes and other processes. We included a light contrast. Although seed
germinated faster in the light (by 0-8 days), the effect of light did not vary between
orientations or among populations (Table 1).
Because the size and shape of seeds influence germination (Dr A. G. Gordon, 1979,
pers. comm.), we correlated germination rate with a number of measures of seed size and
shape and found a significant correlation^ = 0-55;d.f. = 12;P < 005) between the ratio
seed thickness:seed weight and germination rate scale-side-up. The families whose seeds
were thickest for their weight germinated the fastest, suggesting that the distance between
the embryo and the seed coat could be important. This effect may hinge on the sensitivity
of the seeds to moisture content (Belcher, 1975; Hegarty, 1978). A small supplementary
test indicated that Douglas-fir seeds placed on a moist surface equilibrate at different
moisture contents depending on orientation.
Downloaded from http://aob.oxfordjournals.org/ at DigiTop USDA's Digital Desktop Library on October 11, 2012
T A B L E 3. Estimated coefficients (b) of regression, sample standard deviations 0>b) of the
regression coefficients, and coefficients (r) of correlation associated with (1) germination rates
(days'1) vs. provenance distance from the ocean, (2) difference between germination rates
scale-side-up and scale-side-down vs. distance from the ocean and (3) difference between
germination rates scale-side-up and -down vs. mean germination rate for provenance
LITERATURE CITED
ALLEN, G. S. and OWENS, J. N., 1972. The life history of Douglas-fir. Canadian Forestry Service. Environment
Canada. Ottawa, Canada.
BARNETT, J. P., 1976. Delayed germination of southern pine seeds related to seed coat constraint. Can. J. For.
Res. 6, 504-10.
BELCHER, E. W., 1975. Influence of substrate moisture level on the germination of seed and selected Pinus
species. Seed Set. Tech. 3, 597-604.
BLISS, C. I., 1967. Statistics In Biology, vol. 1. McGraw-Hill, New York.
CROCKER, W., 1948. Growth of Plants. Reinhold, New York.
HARPER, J. L., LOVELL, P. H. and MOORE, K. G., 1970. The shapes and sizes of seeds. A. Rev. Ecol. System.
1, 327-56.
HEGARTY, T. W., 1978. The physiology and seed hydration and dehydration, and the relation between water
stress and the control of germination: a review. PI., Cell, Environ. I, 101-19.
IKUMA, H. and THIMANN, K. V., 1963. The role of the seed-coats in germination of photosensitive lettuce seeds.
PI. Cell Physiol. 4, 169-85.
JOHNSGARD, G. A., 1963. Temperature and the water balance for Oregon weather stations. Special Report ISO,
Agricultural Experiment Station, Oregon State University, Corvallis.
MCDONOUGH, W. T., 1977. Seed physiology. Chapter VI in Rangeland Plant Physiology, ed. R. E. Sosebee, pp.
155-184. Society of Range Management, Range Science Series 4.
NOBS, M. A. and HAOAR, W. G., 1974. Analysis of germination and flowering rates of dimorphic seeds from
Atriplex hortensis. Carnegie Inst. of Washington Yearbook 73, 859-864.
NYMAN, B., 1963. Studies on the germination in seeds of Scots pine (Pinus silvestris L.) with special reference
to the light factor. Studia Forestalia Suecica No. 2.
RICHARDSON, S. D., 1959. Germination of Douglas-fir seed as affected by light, temperature and gibberellk
acid. For. Sci. 5, 174-81.
SNEDECOR, G. W. and COCHRAN, W. G., 1967. Statistical Methods. The Iowa State University Press, Ames.
SORENSEN, A., 1978. Somatic polymorphism and seed dispersal. Nature, Lond. 276, 174-6.
STEBBINS, G. L., 1971. Adaptive radiation of reproductive characteristics in angiosperms. II. Seeds and
seedlings. A. Rev. Ecol. System. 2, 237-60.
STONE, E. C , 1957. Embryo dormancy of Pinus Jeffrey! Murr. seed as affected by temperature, water uptake,
stratification and seed coat. PI. Physiol. 32, 93-99.
WEBB, D. P. and WAREING, P. F., 1972. Seed dormancy in Acer pseudoplatanus L.: the role of covering structure.
J. exp. Bot. 23, 813-29.
Downloaded from http://aob.oxfordjournals.org/ at DigiTop USDA's Digital Desktop Library on October 11, 2012
Sorensen and Campbell—Germination of Douglas-fir Seeds
471
Also, the structure or chemistry of the seed coats could be factors. Seed coats are known
to influence germination rate in many ways (Crocker, 1948; Stone, 1957; Ikuma and
Thimann, 1963; Nyman, 1963; Webb and Wareing, 1972; Barnett, 1976), and the
differences in seed colour of the two sides of a Douglas-fir seed may be associated with
structural or chemical differences affecting germination.
The large effect of orientation under laboratory conditions and its correlation with seed
origin suggest that orientation is a potentially important part of the reproductive biology
of Douglas fir. The effect may be particularly significant on sites with relatively short
growing seasons; for example, a difference of 1-8 days occurred between orientations for
a central Oregon sample with an average germination rate of 3-7 days. The relationship
between orientations and germination rate, however, is undoubtedly more complex than
we have described it. Our experiment included a limited number of germination
conditions and seed samples. We arranged seeds with equal numbers scale-side-up or
-down, an unlikely distribution under natural seedfall conditions. In a small test of
distributions, attempted indoors in still air by tapping the base of freshly opened cones,
approximately 55 per cent of the winged seeds came to rest scale-side-down. On a natural,
rougher surface, the orientation would be less determinate. Seeds could rest on their sides,
partially tilted, or even apex up or down. The critical factor may not be the extreme
orientations that we have tested, but whether or not the scale side of the seed is in contact
with the moisture substrate. Considering the apparent adaptive significance of the
response, it may be worthwhile addressing other experiments to these questions.