The Rutaceae comprise about 1500 species of trees and shrubs within about 130 genera which provide edible fruits (e.g. Casimiroa edulis Llave & Lex., white sapote), flavourings (e.g. Murraya koenigii (L.) Spreng., curry-leaf tree) and medicinal products (e.g. Ruta graveolens L., rue). The fruits may be dehiscent capsules, samara-like, or fleshy and berry-like. Seed storage behaviour is now thought to be orthodox. For example, Dictamnus albus L. is maintained in the long-term seed store at the Wakehurst Place Gene Bank, and Murraya exotica L. is known to show orthodox seed storage behaviour. Information on Citrus seed storage behaviour is provided in a subsequent section of this chapter.
SEED DORMANCY AND GERMINATION
Germination of the seeds can be slow and a part of the cause may be dormancy. Treatments to the seed covering structures and gibberellins can promote germination. Detailed information on seed dormancy and germination is provided in this chapter for the genus Citrus (including synonyms within Limonia and Poncirus). Information on other species is summarised in Table 64.1 and in addition the algorithm below may be helpful in developing suitable germination test procedures.
RBG Kew Wakehurst Place algorithm
The first step of the algorithm is to test seeds in an alternating temperature regime of 33°/19°C (12h/12h) with light applied for 12h/d during the period spent at the upper temperature in each cycle.
If this does not result in full germination then the second step in the algorithm is to co-apply 7 x 10-4 M GA3 to the germination test substrate and test a fresh sample of seeds in the above alternating temperature regime.
If this does not result in full germination then the third step in the algorithm is to chip the seed covering structures of a fresh sample of seeds and then test in the alternating temperature regime described in step one, with 7 x 10-4 M GA3 co-applied if the proportion of seeds which germinated in step two was significantly greater than the proportion which germinated in step one.
TABLE 64.1 Summary of germination test recommendations for species within the Rutaceae
|
Species and Authority |
Substrate |
Temperature |
Duration |
Additional directions |
Source |
|
Aegle marmelos (L.) Correa |
|
|
21d |
pre-soak, 24h, then warm stratification |
Riley |
|
Casimiroa spp.
|
|
|
21d |
complete removal of seed covering structures, |
Riley |
|
|
|
|
then pre-soak, 24h |
|
|
|
Clausena spp. |
|
|
21d |
pre-soak, 24h |
Riley |
|
Dictamnus albus L. |
TP |
20°/30°C |
21d |
light, pre-chill, 3°-5°C, 45d |
AOSA |
|
Feronia limonia |
|
|
21d |
pre-soak, 24h |
Riley |
|
Fortunella spp. |
|
|
21d |
pre-soak, 24h |
Riley |
|
Murraya exotica L. |
S |
25°-30°C |
18d |
light, continuous |
CHML |
|
Ruta graveolens L. |
TP; BP |
20°/30°C; 15°C; 20°C |
28d |
pre-chill |
ISTA |
CITRUS
|
C. aurantifolia Swingle [C. lima Lunan; C.
limetta Auth.; Limonia aurantifolia Christm.] |
lime |
|
C. aurantium L. [C. Bigardia Risso] |
sour orange, seville orange |
|
C. grandis Osbeck [C. aurantium var
grandis L.; C. decumana L.; C. maxima Merr.] |
akune burntan, shaddock, pummel, pompelmous |
|
C. jambhiri Lush |
rough lemon, jamberi |
|
C. karna Raf. |
karna khatta |
|
C. limettoides Tan. |
sweet lime |
|
C. limon Burm. f. [C. medica var Limon
L.; C. Limonum Risso] |
lemon |
|
C. limonia Osbeck [C. reticulata Blanco x C.
sinensis Osbeck] |
Rangpur lime |
|
C. natsudaidai Hayata |
natsudaidai |
|
C. paradisi Macf. [C. maxima var uvacarpa
Merr. & Lee] |
grapefruit, pomelo |
|
C. reticulata Blanco [C. nobilis Andr.; C.
deliciosa Ten.; C. nobilis var deliciosa Swingle] |
mandarin, tangerine, satsuma |
|
C. sinensis Osbeck |
sweet orange |
|
C. trifoliata L. [Poncirus trifoliata
Raf.] |
trifoliate-orange |
|
C. trifoliata L. x C. sinensis Osbeck |
troyer citrange |
I. Evidence of dormancy
In all Citrus spp. seed germination is generally slow and often erratic. For example, moist seeds of C. sinensis tested for germination at 23°C showed no germination in the first 15 days in test, and only thereafter was germination observed, with individual seeds continuing to germinate up to at least 31 days in test (17).
Moreover, drying Citrus seeds results in a considerable further delay to radicle emergence (18). There is a linear relationship between the mean rate of seed germination and initial seed moisture content, such that drying moist seeds extracted from fruits at 56% moisture content to 5% moisture content doubles the mean germination time of intact seeds at 20°C from 14 to 28 days, and 42 days are required for full germination (18). It has been suggested that this substantial delay to germination resulting from desiccation has been confused with loss in viability (germination test periods being too short to record the germination of dried seeds) and accounts for conflicts in the literature as to whether or not Citrus seeds can be dried safely (18). The delay to the germination of dried seeds results entirely from the considerable periods taken by the seeds to imbibe sufficiently to reach the fully imbibed state (33).
There is some doubt in the literature as to whether or not seeds of Citrus spp. exhibit dormancy. Despite emphatic reports that there is no dormancy in Citrus seeds (31), seed dormancy can be discerned in some lots (4,5). The major problem in testing seeds of Citrus spp. for germination, however, is the delay to germination. For Citrus seeds, treatments which do or do not reduce the delays to germination, or which do or do not increase the proportion of seeds germinating during tests of limited duration are included here as dormancy-breaking treatments according to the layout used for all other genera. This is partly for convenience and partly because there is a view that slow germination in seeds of Citrus spp. may result, in part, from dormancy (33). It should be noted that the majority of citations concern seeds that have not been dried.
II. Germination regimes for non-dormant seeds
C. aurantifolia
Constant temperatures: 25°C (28)
C. aurantium
Constant temperatures: 15°-38°C, 66d (7); 25°C (23,28); 26°C (24,25); 29°C (11)
Alternating temperatures: 20°/30°C (16h/8h) (3)
C. limon
Constant temperatures: 23°C (27)
Alternating temperatures: 20°/30°C (16h/8h) (3)
C. limonia
Constant temperatures: 21°-38°C, 61d (7)
C. paradisi
Constant temperatures: 15°-38°C, 45d (7)
Alternating temperatures: 20°/30°C (16h/8h) (3)
C. reticulata
Constant temperatures: 15°-35°C (22); 25°C (21,23)
C. sinensis
Constant temperatures: 18°-37°C, 66d (7); 23°-33°C (13); 23°C in light, 100 fc (17)
Alternating temperatures: 20°/30°C (16h/8h) (3)
C. trifoliata
Constant temperatures: 12°-35°C (22); 26°C (6)
C. trifoliata x C. sinensis
Constant temperatures: 26°C (6)
III. Unsuccessful dormancy-breaking treatments
C. aurantifolia
Pre-chill: 3°C, 14d, germinate at 25°C (33)
Pre-soak: 15 min, 50°C, germinate at 30°C, 20d (33)
Citric acid: pre-applied, 15 min, 50°C, 1%, with or without GA3, pre-applied, 24h, 500 ppm, germinate at 30°C, 20d (33)
Ethylenediaminetetraacetic acid: pre-applied, 15 min, 50°C, 0.001 M, with or without GA3, pre-applied, 24h, 500 ppm, germinate at 30°C, 20d (33)
C. aurantium
Pre-chill: 2-6w (12)
Pre-soak: 24h (10)
Pre-wash: 50°C (10)
GA3: pre-applied, 24h, 100, 1000 ppm (29)
Benzyladenine: pre-applied, 24h, 5-25 ppm (29)
Scarification: concentrated sulphuric acid, 20-60s (10)
Sodium hydroxide: pre-applied, 0.1 N, 100%, 20s-20 min (10)
2,4-Dichlorophenoxyacetic acid: pre-applied, 24h, 10-50 ppm (29)
C. limon
Constant temperatures: 20°C, 21d (18)
Pre-chill: 3°C, 7-21d, germinate at 20°C, 42d (33); 2-6w (12)
GA3: co-applied, 50, 150 ppm, germinate at 30°C (33)
C. reticulata
2,4-Dichlorophenoxyacetic acid: pre-applied, 24h, 10-50 ppm (29)
Benzyladenine: pre-applied, 24h, 5-25 ppm (29)
GA3: pre-applied, 24h, 100, 500 ppm (29)
C. sinensis
Constant temperatures: 21°C, 35d (4)
C. trifoliata, C. trifoliata x C. sinensis
GA3: pre-applied, 10 mg/l (6); co-applied, 10 mg/l (6)
IV. Partly-successful dormancy-breaking treatments
C. aurantifolia
Constant temperatures: 20°C, 70d (19); 20°C, 25°C, 42d (33)
GA3: pre-applied, 6,12h, 10-40 ppm (8); co-applied, 34.6, 3460 ppm, at 20°C, 42d (33)
1-Napthaleneacetic acid: pre-applied, 6, 12h, 10-40 ppm (8)
Thiourea: pre-applied, 6, 12h, 1-2% (8)
Potassium nitrate: pre-applied, 6, 12h, 1-2% (8)
C. aurantium
Constant temperatures: 20°C, 70d (18)
Pre-chill: 0°-5°C, 45d (30)
GA3: pre-applied, 24h, 500 ppm (29)
Removal of seed covering structures: testa, germinate at 25°C (10,23)
C. jambhiri
Pre-chill: 0°-5°C, 45d (30)
C. limettoides
Ethylenediaminetetraacetic acid: pre-applied, 15 min, 10-3 M, 50°C, then GA3, pre-applied, 24h, 200-500 ppm (1)
C. limon
Constant temperatures: 20°C, 42d (18); 20°C, 70d (19); 20°C, 25-56d (33)
Removal of seed covering structures: testa, germinate at 20°C, 42d (18)
Scarification: emery paper, germinate at 20°C, 42d (18)
C. limonia
Potassium nitrate: pre-applied 6, 12h, 1-2% (9)
Thiourea: pre-applied, 6, 12h, 1-2% (9)
1-Napthaleneacetic acid: pre-applied, 6, 12h, 10, 20 ppm (9)
C. paradisi
Removal of seed covering structures: slit or remove testa (34)
C. reticulata
Pre-chill: 0°-5°C, 45d (30)
GA3: pre-applied, 24h, 1000 ppm (29)
C. sinensis
Pre-chill: 3°-4°C, 7-21d, germinate at 21°C, 35d (4); 3°-4°C, 56d, germinate at 21°-32°C (4); 3°-4°C, 56d, then GA3, pre-applied, 24h, 1000 ppm, germinate at 21°C, 27°C, 32°C (4)
Pre-soak: 24h (5)
GA3: pre-applied, 24h, 1-10,000 ppm (5); pre-applied, 24h, 1000 ppm, germinate at 21°C (4)
Removal of seed covering structures: remove outer seed coat (34)
C. trifoliata
Pre-chill: 4°C, 28-84d, germinate at 26°C (6); 2-6w (12); 4.5°C, 2,4w, germinate at 25°C (21)
GA3: pre-applied, 24h, 1000 ppm, germinate at 18°C (26)
C. trifoliata x C. sinensis
Pre-chill: 4°C, 28-84d, germinate at 26°C (6)
V. Successful dormancy-breaking treatments
C. aurantifolia
Constant temperatures: 30°C, 35°C, 42d (33)
Alternating temperatures: 25°/35°C (12h/12h), 42d (33)
Pre-chill: 3°C, 14d, germinate at 25°C, 42d (33)
GA3: pre-applied, 12h, 40 ppm (8); pre-applied, 24h, 500 ppm, germinate at 30°C, 20d (33); co-applied, 346 ppm, germinate at 20°C, 42d (33)
1-Napthaleneacetic acid: pre-applied, 12h, 40 ppm (8)
Removal of seed covering structures: testa, germinate at 25°C (28)
C. aurantium
Removal of seed covering structures: seed coat, germinate at 25°C (28)
C. jambhiri
GA3: pre-applied, 24h, 500 ppm (32)
C. karna
Potassium nitrate: pre-applied, 24h, 750 ppm (32)
C. limon
Constant temperatures: 30°C, 28d (33)
Removal of seed covering structures: testa, germinate at 23°C (27); testa, germinate at 25°C (28)
Alcohol: pre-applied, 30,40 min, 80% (28)
Sodium hydroxide: pre-applied, 30 min, 5% (2)
C. limonia
GA3: pre-applied, 6, 12h, 10-40 ppm (9)
1-Napthaleneacetic acid: pre-applied, 6, 12h, 40 ppm (9)
C. sinensis
Ascorbic acid: pre-applied, 12h, 100 ppm (20)
C. trifoliata
Pre-chill: 4.5°C, 5-12w, germinate at 25°C (21); 5°C, 30-120d, germinate at 30°C (14)
VI. Comment
Although Citrus seeds will germinate over a wide range of constant temperatures, roughly 15°-38°C (7), seed germination is most rapid and complete within the range 30°-35°C (4,7,22,33). In dry Citrus seeds the time taken to germinate is affected by the rate of imbibition and the rate of the subsequent growth processes. Temperature affects both; because of the substantial delays to the germination of dried Citrus seeds it is important to optimise the germination test temperature with regard to these rates; at 30°C the rates are not at a maximum (22,33), but higher temperatures during the growth phase can cause seedling abnormalities and may reduce percentage germination in some seed lots (22,33). Consequently a germination test temperature of 30°C is recommended. Germination tests at this temperature should be continued until a full seven days have passed in which no seeds have begun to germinate. It is expected that this will require about 28-42 days.
Treatment with gibberellins can reduce the time taken to germinate (1,5,33), but the effect is marginal (33) and unreliable (5). Indeed in many cases control treatments where the seeds are pre-soaked in water with no gibberellins present account for most of the apparent effect of gibberellins compared to untreated seeds (5). Treatment with gibberellins after treatment with chelating agents (e.g. ethylenediaminetetraacetic acid) has been reported to reduce further the germination time due to increased permeability of the seed coat (1). Again, however, the effect appears unreliable since other workers report no effect of such combined treatments (33). If dormancy is present then pre-chill treatments are more effective than treatment with gibberellins and also reduce the time taken to germinate in the subsequent germination test regime (6) - although if the pre-chill period is included in the calculation of the time taken to germinate then there is no overall reduction (33).
It is not possible to give precise recommendations for pre-chill treatment periods where these are required to remove dormancy. Nevertheless, it is suggested that these be carried out at 3°-5°C, and that it is unlikely that treatment periods greater then 28 days are necessary.
Removing seedcoats can also reduce the time taken to germinate by both moist (11) and dry (18) seeds. In both cases the effect can be substantial (in contrast to the marginal effect of gibberellins). For example, removing seedcoats from moist seeds of C. aurantium reduced the mean germination time from about 35 to 5 days at 29°C (11). Removing seed coats as a regular procedure is not recommended, however, since it is time consuming and - provided sufficient time is allowed in the germination test - rarely results in greater cumulative percentage germination (18). Moreover, embryos can be damaged where the seed coats are removed from dried Citrus seeds (18).
It is worth noting that the extreme period taken by dry Citrus seeds to imbibe can also affect the results of rapid viability tests. Tetrazolium staining studies with seeds of C. trifoliata (21) and C. natsudaidai (16) show that such tests are unreliable for dried seeds. A further important point to note concerns the availability of moisture during the germination test. Slight water stress (-2.3 bars) substantially reduced the percentage of seeds - moist, C. sinensis - germinating - at least within 31 days - and dramatically increased the time taken to germinate (17), whereas in sunflower and lettuce, the same degree of moisture stress caused only a minor delay to germination and had no significant effect on the percentage of seeds which had germinated by the end of the test (17). It is thus vital, and much more important than in most other species, to ensure that sufficient moisture is available to Citrus seeds throughout germination tests. Testing between paper and regularly re-moistening the paper should be sufficient to prevent problems arising from this source.
VII. References
1. Achituv, M. and Mendel, K. (1973). Effect of certain treatments on the germination of sweet lime (Citrus limettoides Tan.) seed. The Plant Propagator, 19 (4), 15-20.
2. Anonymous, (1980). Methods of hastening the germination of citrus seed. Information Bulletin, Citrus and Sub-Tropical Fruit Research Institute, 91, 3-4. (From Seed Abstracts, 1981, 4, 2820.)
3. Barton, L.V. (1943). The storage of citrus seed. Contributions from the Boyce Thompson Institute, 13, 47-55.
4. Burger, D.W. and Hackett, W.P. (1982). Influence of low temperature and gibberellic acid treatment on the germination of 'Valencia' orange seed. HortScience, 17, 801-803.
5. Burns, R.M. and Coggins, C.W. (1969). Sweet orange germination and growth aided by water and gibberellin seed soak. California Agriculture, 23, 18-19.
6. Button, J., Bornman, C.H. and Hackland, B.A. (1971). Effect of some pre-sowing treatments on the germination of Poncirus trifoliata and troyer citrange seeds. The Citrus and Sub-Tropical Fruit Journal, 45, 9-11.
7. Camp, A.F., Mowry, H. and Loucks, K.W. (1933). The effect of soil temperature on the germination of citrus seeds. American Journal of Botany, 20, 348-357.
8. Choudhari, B.K. and Chakrawar, V.R. (1980). Effect of some chemicals on the germination of kagzi lime (Citrus aurantifolia Swingle) seeds. Journal of Maharashtra Agricultural Universities, 5, 173-174. (From Seed Abstracts, 1982, 5, 308.)
9. Choudhari, B.K. and Chakrawar, V.R. (1981). Note on the effect of some chemicals on the germination of Rangpur lime seeds. Indian Journal of Agricultural Sciences, 51, 201-203.
10. Cohen, A. (1956). Studies on the viability of citrus seeds and certain properties of their coats. Bulletin of the Research Council of Israel, Section D, 5, 200-209.
11. Demni, S. and Bouzid, S. (1979). Premières informations sur la germination des graines de bigaradier (Citrus aurantium L.). Fruits, 34, 283-287.
12. Elze, D.L. (1949). Germination of citrus seeds in relation to certain nursery practices. Palestine Journal of Botany, 7, 69-80.
13. Fawcett, H.S. (1929). Temperature experiments in germinating orange seeds. California Citrograph, 14, 515. (Cited by Edwards, T.I. (1932). Temperature relations of seed germination. Quarterly Review of Biology, 7, 428-444.)
14. Fu, W.H. (1951). Germination and storage of trifoliata orange seeds. California Citrograph, 37, 38-39.
15. Honjo, H. and Nakagawa, Y. (1978). Suitable temperature and seed moisture content for maintaining the germinability of citrus seed for long-term storage. In Long-term Preservation of Favourable Germplasm in Arboreal Crops (eds. T. Akihama and K. Nakajima), pp. 31-35, Fruit Tree Research Station, Fujimoto, Japan.
16. Honjo, H. and Nakagawa, Y. (1979). [Tetrazolium test for evaluating the germination capacity of citrus seed.] Bulletin of the Fruit Tree Research Station, Series A, 6, 37-42.
17. Kaufmann, M.R. (1969). Effect of water potential on germination of lettuce, sunflower and Citrus seeds. Canadian Journal of Botany, 47, 1761-1764.
18. King, M.W. and Roberts, E.H. (1980). The desiccation response of seeds of Citrus limon L. Annals of Botany, 45, 489-492.
19. King, M.W., Soetisna, U. and Roberts, E.H. (1981). The dry storage of Citrus seeds. Annals of Botany, 48, 865-872.
20. Misra, R.S. and Verma, V.K. (1980). Studies on the seed germination of Kinnow orange in the Central Himalayas. Progressive Horticulture, 12, 79-84. (From Seed Abstracts, 1981, 4, 3153.)
21. Mobayen, R.G. (1980). Germination of trifoliate orange seed in relation to fruit development, storage and drying. Journal of Horticultural Science, 55, 285-289.
22. Mobayen, R.G. (1980). Germination and emergence of citrus and tomato seeds in relation to temperature. Journal of Horticultural Science, 55, 291-297.
23. Monselise, S.P. (1953). Viability tests with citrus seeds. Palestine Journal of Botany, 8, 152-157.
24. Monselise, S.P. (1959). Citrus germination and emergence as influenced by temperature and seed treatments. Bulletin of the Research Council of Israel, Section D, 7, 29-34.
25. Monselise, S.P. (1962). Citrus seed biology. XVIth International Horticultural Congress, Brussels, 559-565.
26. Moss, G.I. (1980). Propagation of citrus for future planting. Proceedings of the International Society of Citriculture, 1978, Griffith, NSW, Australia, 132-135.
27. Mumford, P.M. and Grout, B.W.W. (1979). Desiccation and low temperature (-196°C) tolerance of Citrus limon seed. Seed Science and Technology, 7, 407-410.
28. Panggabean, G. (1981). Dry storage of citrus seeds and factors affecting their germination. M.Sc. Thesis, University of Birmingham.
29. Rawash, M.A., Montaser, A., Habib, S.S., Nabawy, S.E. and Mahmoud, N. (1980). Germination of some citrus seeds as affected by soaking in growth regulators, water washing and sowing date. Research Bulletin, Faculty of Agriculture, Ain Shams University, 1299, 1-10.
30. Rawash, M.A. and Mougheith, M.G. (1978). Effect of some storage treatments on seed germination of some citrus rootstocks. Research Bulletin, Faculty of Agriculture, Ain Shams University, 835, 1-11.
31. Schneider, H. (1968). The anatomy of citrus. In The Citrus Industry (eds. W.L. Rather, L. Batchelor and H. Webber), Volume II, pp. 1-85, University of California, Berkeley, USA.
32. Singh, H.K., Shankar, G. and Makhija, M. (1979). A study on citrus seed germination as affected by some chemicals. Haryana Journal of Horticultural Science, 8, 194-195. (From Seed Abstracts, 1981, 4, 143.)
33. Soetisna, U., King, M.W. and Roberts, E.H. (1985). Germination test recommendations for estimating the viability of moist or dry seeds of lemon (Citrus limon L. Burm.) and lime (C. aurantifolia (Christm.) Swing.). Seed Science and Technology, 13, 87-110.
34. Tager, J.M. and Cameron, S.H. (1957). The role of the seed coat in chlorophyll deficiency (albinism) of citrus seedlings. Physiologia Plantarum, 10, 302-305.
