Annals of Botany 34: 439-446  (1920)
The Role of the Seed-coat in Relation to the Germination of Immature Seed.
BY
FRANKLIN KIDD
AND
CYRIL WEST
With six Tables and one Chart in the Text.

IN previous papers (9 and 10) it has been shown that the germination of seeds of Brassica alba, sown in the presence of certain percentages of carbon dioxide, can be completely inhibited, and that this inhibition of germination is often maintained indefinitely after the removal of the seeds to air. This is a remarkable phenomenon, and it is the more striking in that seeds rendered dormant by carbon dioxide show no signs of injury when finally brought to germination even after the lapse of twelve months. So far the authors have found only two certain methods of destroying the dormant condition into which the seeds are thrown by the carbon dioxide treatment. One method is to redry the seed, the other method is to remove the testa without drying.

In the present paper results are recorded which show that a condition of dormancy, similar in many ways to that produced when dry mature seeds of Brassica alba are sown in the presence of carbon dioxide, may be observed when immature seeds of this plant are sown immediately after removal from the parent and before the natural drying process has begun.

EXPERIMENTS WITH BRASSICA ALBA

The seeds in the following experiments were obtained from plants grown in the Botanic Gardens, Cambridge. The different degrees of ripeness of the seeds were as follows:

A. Green-ripe: Seeds fully swollen; still quite green; 50 per cent. to 80 per cent. of the dry weight of ripe seeds. Siliquas bright green.

B. Yellow-ripe: Seeds fully swollen; yellow in colour; of practically the same dry weight as fully ripe seeds. Siliquas still moist; beginning to turn yellow.

The germination results are set forth in the following tables (Tables I-IV) and chart. Yellow-ripe seeds with the testa intact, when sown immediately after removal from the parent, remained in a dormant condition or showed a long delay in germination. On the other hand, yellow-ripe seeds with the testa removed, or dried in the laboratory for twenty-four hours with the testa intact, germinated immediately (100 per cent.), even more rapidly than a sample of the previous year's seed used as a control.

Green-ripe seeds sown after the removal of the testa germinated completely (100 per cent.), but after a delay of a few days. When sown with the testa on they mostly died, and in the case of the few seeds which did germinate, it was noticed that germination was always preceded by a change in the colour of the radicle from green to yellow. If dried for forty-eight hours before sowing practically all the seeds were killed.

Table I.
Germination of White Mustard Seeds at different stages of maturity.

Lot A. Twenty yellow-ripe seeds gathered and removed from the siliquas on Sept. 8, 1918. Dried in the laboratory for one day.
Lot B. Twenty yellow-ripe seeds gathered on Sept. 8 and removed from the siliquas on Sept. 9. Sown immediately (i. e. not dried).
Lot C. Twenty green seeds gathered and removed from the siliquas on Sept. 8. Sown immediately (i.e. not dried).
Lot D. Twenty green seeds gathered and removed from the siliquas on Sept. 8. Testas removed before sowing.
All the seeds were sown on moist silica sand in a glass thermostat at 20° C.

 

  Germinations after:
16
hours
40
hours
45
hours
65
hours
4
days
5
days
7
days
9
days
12
days
21
days
Lot A 1 16 18 20 - - - - - -
Lot B 0 0 4 9 10 13 13 14 15 20
Lot C 0 0 0 0 0 1 5 8 10 14*
Lot D 0 0 0 0 2 3 20 - - -
  *The 6 ungerminated seeds were dead.

Table II.
Germination of White Mustard Seeds at different stages of maturity.

All the seeds were sown on moist silica sand in a thermostat at 20° C.

Kind of Seed sown
(sown on Sept. 11)
Number
of Seeds
sown
18
hours
24
hours
41
hours
65
hours
5
days
7
days
10
days
21
days
Green-ripe, testas off 10 0 0 0 0 7 10 - -
"             on 10 0 0 0 0 0 0 1 1*
Yellow-ripe, testas off 10 9 10 - - - - - -
"             on 10 0 0 3 5 5 6 6 7†
Dry mature seed (1917) 10 0 6 10 - - - - -
Yellow-ripe, dried in the
laboratory air for 3 days
10 3 10 - - - - - -
* The nine ungerminated seeds were dead. † The three ungerminated seeds were not dead.

Table III.
Germination of White Mustard Seeds at different stages of maturity.

All the seeds were sown on moist sand in a thermostat at 20° C.

Kind of Seed sown
(sown on Sept. 11).
Number of
Seeds sown
Germination after:
1
day
3
days
5
days
8
days
17
days
Yellow-ripe, testas off, dried in the laboratory air for 2 days 10 8 10 - - -
Green-ripe, testas off, dried in the laboratory air for 2 days 10 0 1 2 2 2*
Green-ripe, testas on, dried in the laboratory air for 2 days 10 0 0 0 0 0*
* The ungerminated seeds were dead.

Table IV.
Germination of White Mustard Seeds at different stages of maturity.

Lot A consisted of ten green-ripe seeds.
Lot B consisted of ten bare embryos from similar seeds.
Sown on moist sand, July 14.
  Number of Germinations after:
2 days 4 days 6 days 7 days 12 days 18 days
Lot A 0 0 0 0 1 2*
Lot B 0 0 5 10† - -
*The eight ungerminated seeds finally died. † Healthy plants.

CHART. Germination of White Mustard seeds at different stages of maturity as affected by the seed-coat and by drying the seed (see Table II).

From the above experiments on the germination of immature White Mustard seeds it appears that the presence of the testa is fatal so long as the seeds are still green and have not reached their full dry weight. If they are sown without the testa they germinate in a healthy manner, but if sown with the testa on they perish. As the seed ripens, however, and turns yellow, the effect of the presence of the sced-coats is markedly reduced. The presence of the seed-coats in the yellow-ripe stage only causes dormancy or delayed germination. Finally, when the testa is completely yellow and the seed is dry, the testa no longer has any appreciable effect upon germination.

Since the embryos will germinate freely at all three stages if the seed-coats are removed, there does not seem to be any ground for seeking an explanation of the above differences in behaviour between green-ripe, yellow-ripe, and dry-ripe seeds in progressive changes occurring in the embryo itself.

1 For details as to the histological and microchemical characters of the testa in the case of the genus Brassica, the reader is referred to papers by Schroeder (21), Sempelowski (22), Holfert (8), Burchard (3), Gram (7), Kinzel (14), Pieters and Charles (20), Kondo (16), and Kidd and West (10).

The progressive changes which occur in the testa, to which we must therefore look for an explanation, may be described as follows. In the green-ripe seed the embryo is enclosed in a relatively thick green coat consisting of actively functioning tissue, while between this coat and the embryo a considerable quantity of liquid is usually found. In the yellow-ripe seed the testa is relatively thinner and presumably less active, but is still living. The liquid between the embryo and the seed-coats has by this time disappeared. In the dry-ripe seed the testa is a thin membrane and presumably dead.1

Considered as an obstacle to the gaseous exchange of the embryo the testa probably behaves in relation to its thickness in the same way as a film of water; in other words, the thicker the testa the more slowly do gases pass through.

2 In this paper the literature dealing with the effect of soaking seeds in water is critically reviewed.

It is a well-known fact that immersion in water does inhibit the germination of most seeds, and that prolonged immersion is fatal (see Kidd and West, 11 and 132).

Further, the living testa, i. e. the testa in the green-ripe and yellow-ripe stages as opposed to the dry-ripe stage, will not only consume oxygen and, as it were, steal it on its way to the embryo, but on account of its production of carbon dioxide will also tend to hinder the escape of this gas from the tissues of the embryo. The progressive changes in the testa described above will therefore for two reasons be in favour of a progressively greater concentration of oxygen available for the embryo, and a more rapid escape of carbon dioxide.

In this connexion it is interesting to note that Demoussy (5) found that hydrogen peroxide increased the percentage of germination of old Cress seeds. Hydrogen peroxide has a lethal action upon the saprophytic flora of the dead seed-coats. He suggested that non-germination in these old seeds was to some extent due to the respiratory action of moulds or bacteria present in the seed-coat, attributing to them a role in causing dormancy similar to that which we are here attributing to the respiratory activity of the living testa.

EXPERIMENTS WITH PISUM SATIVUM

The results of certain experiments with immature seeds of Pisum sativum are recorded in Table V, and appear to be essentially similar to those obtained with Brassica alba.

Table V.
Germination of seeds and bare embryos of Pisum sativum at different stages of maturity.

A { Lot I Twenty immature seeds. These had attained their full size (the funicle comes away with the pea when taken from the pod).
Lot II Twenty bare embryos from similar seeds.
B { Lot I Twenty less immature seeds (peas break from the funicle when taken from the pod).
Lot II Twenty bare embryos from similar seeds.
C { Lot I Twenty seeds picked ten days later.
Lot II Twenty bare embryos from similar seeds.
D { Twenty seeds picked at a still later stage.
E { Controls = Twenty mature seeds gathered during previous season.

Seeds sown in garden soil. Temperature 15°-20° C.

  Number of Germinations after:
5 days 10 days 14 days
A Lot I 0 1 (19 dead) -
Lot II 20 20 (18 healthy) -
B Lot I 0 0 (all dead) -
Lot II 20 20 (all healthy) -
C Lot I - 10 (10 ungerminated dead) -
Lot II - 20 (all healthy) -
D   - - 18
E [Controls] 20 - -

 

1 It is interesting to note here that this is the same relation between seeds of Brassica alba and those of Pisum sativum in regard to dormancy as that found when the germination of these seeds was inhibited by atmospheres containing certain percentages of carbon dioxide (cf. Kidd, 9). Seeds of White Mustard exhibit secondary dormancy when removed to air. No such phenomenon can be obtained with peas.

If the testa is not removed a large proportion of the seeds perish when sown. The most immature seeds tested showed a high mortality, but as maturity was approached so the injurious effect of the testa was decreased, nevertheless the seeds that survived showed an appreciable delay in germination. No real dormancy similar to that described above in the case of Brassica alba was observed.1

In order to test our hypothesis that the effect of the testa, as shown in the experiments above, is to be attributed to its property in limiting the gaseous exchange of the embryo, further experiments were carried out in the following way. It was argued that, assuming the hypothesis to be correct, these immature seeds, when sown under germinating conditions, must either be in a condition in which the limitation of the gaseous exchange of the embryo is so great as to become actually harmful (e.g. in the cases where sowing on damp sand is followed by death), or at any rate must be near the point at which any further limitation will cause injury. It follows that if we impose even for a short period a condition which further limits the gaseous exchange of the embryo we should, if our hypothesis is correct, obtain a pronounced result. In order to further limit the gaseous exchange, the seeds were immersed in water for short periods before sowing. The result, as Table VI shows, is striking and bears out our hypothesis. Whereas fully ripe dry pea seeds will endure immersion in water for several hours without showing any obvious decrease in the percentage of germination (Kidd and West, 11), the unripe pea seeds suffer heavily even after a few hours' immersion. The amount of injury shown is more or less proportional to the period of soaking. On the other hand, soaking per se for the periods used in this experiment is in no way harmful to the embryo, as is shown by the results of a parallel series of experiments with the bare embryos from unripe pea seeds.

Table VI.

The seeds used in this experiment were similar to those of category C in Table V. Twelve seeds were used in each experiment. They were sown in garden soil. Temperature 15°-20° C.

Condition of the Seed Treatment Results observed 11 days after sowing:
Percentage of
seeds dead
Percentage of
vigorous plants
Average length
of the shoots
Immature
With testas
Dried in air for 15 hours before sowing 20 58 2 cm.
Sown immediately after removal from parent plant 33 50 2 "
Soaked in tap-water for 1 1/2 hours before sowing 50 42 3 "
Soaked in tap-water for 5 hours before sowing 58 33 2 "
Immature
Without testas
Dried in air for 15 hours before sowing 0 50 2.5 "
Sown immediately after removal from parent plant 0 92 3 "
Soaked in tap-water for 1 1/2 hours before sowing 0 100 4.5 "
Soaked in tap-water for 5 hours before sowing 0 100 4.5 "
Mature intact seeds Soaked for 24 hours 0 100 8.5 "

The above table also shows results which were obtained with bare embryos and with intact seeds which were dried in the air for fifteen hours before sowing. The germination of the bare embryos is reduced from 100 per cent. to 50 per cent. by the drying process, which is thus shown to be injurious to unripe pea seeds. A similar reduction in the percentage of germination occurs when the testa is allowed to remain in situ whether the seeds are dried previous to sowing or not. When the seeds are not subjected to the drying process before sowing, the living testa is responsible for the 50 per cent. mortality observed. When the seeds are sown after having been dried the testa is presumably a dead membrane, and it is now the drying of the seed which causes the 50 per cent. mortality observed, just as in the case of the bare embryos which had been allowed to dry.

Experiments conducted by Dr. F. F. Blackman and Miss N. Darwin (4), of which an abbreviated account was read at the British Association Meeting held at Sheffield in 1910, but of which no published record is at present available, are significant in relation to the hypothesis put forward above to the effect that the living testa of unripe seeds limits the gaseous exchange of the embryo in the same way as a continuous film of water. They worked with barley grains, the vitality of which had been reduced (by age or by immersion in the swollen condition in hot water at 50° C. (circa) for twenty minutes), but which still showed a full percentage of germination when sown under ideal conditions. It was found that slight films of water greatly delayed the germination, reduced the germination percentage, and resulted in the death of a large proportion of such seeds. The results obtained were more or less proportional to the thickness of the water films.

CONCLUSIONS.

Many authors (see especially Nobbe (19), Maze (18), Windisch (23), Eberhart (6), Atterberg (1), Babcock (2), Kinzel (15), and Kondo (17)) have described experiments dealing with the dormancy or delayed germination observed when certain seeds, which, although immature and with a relatively high moisture content, have nevertheless attained their full size, are sown immediately after removal from the parent plant. The process of drying has generally been found to terminate the dormant condition of such seeds. Different theories have been put forward to account for the dormancy of unripe seeds sown in the moist condition immediately after removal from the parent plant (see Kidd and West, 12). In the present paper it has been shown that in the case of Brassica alba and Pisum sativum the removal of the testa not only accelerated the germination and terminated the dormant condition of unripe seeds, but also increased the germination percentage. It is clear that the rest period observed when attempts are made to germinate unripe seeds fresh from the parent plant may be largely attributed to the presence of the testa, and there are strong indications that under these conditions the living testa limits the gaseous exchange of the embryo. A fact which should always be borne in mind in this connexion is that the testa, considered as a membrane through which the gaseous exchange of the embryo must occur, undergoes great modifications during the ripening and drying off of the seed.

Botany School,
Cambridge, 1919.

LITERATURE CITED

  1. ATTERBERG, A.: Die Nachreife des Getreides. Landw. Versuchs-Stat., lxvii, 1907, p. 129.
  2. BABCOCK, S. M.: Metabolic Water: Its Production and Role in Vital Phenomena. Univ. of Wisconsin Agric. Expt. Sta., Research Bull., xxii, 1912.
  3. BURCHARD, O.: Ueber den Bau der Samenschale einiger Brassica- und Sinapis-Arten. Journ. f. Landwirtschaft, xlii, 1894, p. 125.
  4. DARWIN, N., and BLACKMAN, F. F.: Germination Conditions and the Vitality of Seeds. Rep. Brit. Assoc. Advanc. Sci., Sheffield, 1910, p. 786.
  5. DEMOUSSY, E.: Influence de 1'Eau oxygenee sur la Germination. C. R. de l'Acad. des Sci. (Paris), clxii, 1916, p. 435.
  6. EBERHART, C.: Versnche iiber die Keimungsverhaltnisse frischgeemteter Samen. Fiihling's Landw. Ztg., lv, 1906, p. 583.
  7. GRAM, B.: Untersuchungen iiber die Futtermittel des Handels, etc. XIX. Ueber Rapskuchen und deren Verunreinigung. Landw. Versuchs-Stat., 1, 1898, p. 449.
  8. HOLFERT, J.: Die Nahrschicht der Samenschalen. Flora, 73, 1890, p. 279.
  9. KIDD, F.: The Controlling Influence of Carbon Dioxide in the Maturation, Dormancy, and Germination of Seeds. I. Proc. Roy. Soc, Lond., B., lxxxvii, 1914, p. 408.
  10. KIDD, F., and WEST, C.: The Controlling Influence of Carbon Dioxide. IV. On the Production of Secondary Dormancy in Seeds of Brassica alba following Treatment with Carbon Dioxide, and the Relation of this Phenomenon to the Question of Stimuli in Growth Processes. Ann. Bot., xxxi, 1917, p. 457.
  11. ————————: Physiological Predetermination: The Influence of the Physiological Condition of the Seed upon the Course of Subsequent Growth and upon the Yield. I. The Effects of Soaking Seeds in Water. Ann. Appl. Biol., v, 1918, p. 1.
  12. ————————: Physiological Predetermination, &c III. Review of Literature. Chapter II. Ibid., 1919, p. 157.
  13. ————————: Physiological Predetermination, &c. IV. Review of Literature. Chapter III. Ibid., p. 220.
  14. KINZEL, W.: Ueber die Samen einiger Brassica- und Sinapis-Arten, mit besonderer Beriick sichtigung der ostindischen. Landw. Versuchs-Stat., Hi, 1899, p. 169.
  15. ———:Frost und Licht als beeintlussende Krafte bei der Samenkeimung. Stuttgart, 1913.
  16. KONDO, M.: Untersuchung der Samen der in Japan vertretenen Brassica-Arten. Ein Beitrag zur genauen Feststellung der Sortenunterschiede. Ber. d. Ohara-Inst. f. landw. Forsch., i, 1917, p. 123.
  17. ————: Ueber Nachreife und Keimung verschieden reifer Reiskorner (Oryza sativa, L.). Ibid., 1918, p. 361.
  18. MAZÊ, P.: La Maturation des Graines et l'Apparition de la Faculte germinative. C. R. de l'Acad. des Sci. (Paris), cxxxv, 1902, p. 1130.
  19. NOBBE, F.: Ueber kiinstliche Getreidetrocknung mit Bezug auf die Keimfahigkeit. Mitt. Deutsch. Landw. Gesellsch., xii, 1897, p. 185.
  20. PIETERS, A. J., and CHARLES, V. K.: The Seed-coats of certain Species of the Genus Brassica. U.S. Dept. Agric, Div. of Bot., Bull. 29, 1901.
  21. SCHROEDER, J.: Untersuchung der Samen der Brassica-Arten und Varietäten. Landw. Versuchs-Stat., xiv, 1871, p. 179.
  22. SEMPELOWSKI, : Ueber den Bau der Schale landwirthschaftlich wichtiger Samen. Landw. Jahrb., iii, 1874, p. 823.
  23. WINDISCH, W.: Warum keimt die getrocknete, bezw. abgelagerte Gerste besser als die frisch geerntete? Biedermann's Centralbl. f. Agrikulturchemie, xxxiv, 1905, p. 623.