Scientia Horticulturae 104 (2005) 91–99 www.elsevier.com/locate/scihorti
Pollinizer influence on almond seed dormancy Marta Garcı´a-Gusano, Pedro Martı´nez-Go´mez, Federico Dicenta* Departamento de Mejora y Patologı´a Vegetal, Centro de Edafologı´a y Biologı´a Aplicada del Segura (CSIC), P.O. Box 164, E-30100 Espinardo, Murcia, Spain Accepted 30 June 2004
Abstract Breaking seed dormancy is necessary for the completion of germination. Maternal tissues (integument and endocarp) appear to control seed germination, although the effect of the pollinizer has also been described. In this work, the influence of the pollinizer flowering time was studied on the stratification requirements of almond seed germination. Stratification requirements were determined in seeds (with and without endocarp) from crosses between a female progenitor and pollinizers of different flowering dates. The results showed an influence of the pollinizer in the stratification requirements of seeds with endocarp. However, no correlations were found between pollinizer flowering date and stratification requirements, which seem to be determined mainly by the female progenitor. In the case of seed stratification without endocarp, the results did not show any influence of pollinizer on stratification requirements. The observed variability within females (within or between crosses) could be due to differences in the shell characteristics, the different accumulation of inhibitors in the integument, or the genetic nature of the embryos. # 2004 Elsevier B.V. All rights reserved. Keywords: Prunus dulcis; Seed germination; Xenia; Flowering time
1. Introduction Seed dormancy is a mechanism to protect temperate fruits from frost damage during winter, and its breakage is necessary for the completion of germination. Stratification in cold chambers, application of hormones, and the removal of the integument are methods * Corresponding author. Tel.: +34 968 396 309; fax: +34 968 396 213. E-mail address:
[email protected] (F. Dicenta). 0304-4238/$ – see front matter # 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.scienta.2004.06.006
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traditionally used to break dormancy in fruit species (Chao and Walker, 1966; Du Toit et al., 1979; Mehanna et al., 1985; Frisby and Seeley, 1993). In almond (Prunus dulcis (Mill.) D.A. Webb), different investigators have indicated the hormonal nature of dormancy and the positive effect of stratification in its breakage (Grigorian, 1972; Gonza´ lez-Cepeda, 1975). In addition, Garcı´a-Gusano et al. (2004) studied the effect of temperature, genotype, and endocarp on dormancy breakage. Seeley et al. (1998) and Martı´nez-Go´ mez and Dicenta (2001) characterised seed dormancy in peach (P. persica L.) as two independent mechanisms: external seed dormancy controlled by the seed coat (maternal tissue) with a hormonal component and manifested by the inhibition of germination, and internal seed dormancy, controlled by the embryo (female and male contribution) and expressed in later plant growth. Maternal tissues (integument and endocarp) appear to control seed germination (Chao and Walker, 1966; Du Toit et al., 1979; Seeley et al., 1998; Martı´nez-Go´ mez and Dicenta, 2001). Mehlenbacher and Voordeckers (1991) observed that seeds from late-flowering apple cultivars needed longer stratification requirements than seeds from early-flowering cultivars. In almond, Grigorian (1972) and Garcı´a-Gusano et al. (2004) examined the relationship between the flowering time of a cultivar and the stratification requirements of seeds for germination. These authors indicated a great variability in the stratification requirements of seeds for germination, probably due to their origin (open pollination). In addition, Kester (1969) and Grigorian (1972) indicated an effect of the pollinizer on germination, establishing a correlation between pollinizer time of blooming and the stratification requirements of germination, the genotype of the embryo being responsible for these requirements. The objective of this work was to study the influence of pollinizer flowering time (chilling requirements of flower buds for developing) on the stratification period required for almond seed germination.
2. Materials and methods 2.1. Almond cultivars The 11 cultivars studied covered a range of flowering times (chilling requirements for flowering) and shell hardness (Table 1). 2.2. Experimental design Stratification requirements for germination were studied in seeds from 17 crosses between the almond cultivars assayed (Table 2). These crosses, made by hand crosspollination in the almond breeding program of CEBAS-CSIC in Murcia, Spain, were performed between 1998 and 2000: - Lauranne crossed with different male parents (Desmayo Largueta, Marcona, Lauranne, open pollination, Primorskii, and S5133), then stratifying the seed with endocarp in 1998.
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Table 1 Almond cultivars assayed (origin, pedigree, date of flowering, and shell hardness) a
Cultivar
Origin
Pedigree
Flowering
Desmayo Largueta Peraleja S2332 Marcona Marta Anton˜ eta Lauranne A2198 Primorskii S5133 R1000
Spain Spain Spain Spain Spain Spain France Spain Ukraine Spain France
Unknown Unknown Peraleja Tuono Unknown Ferragne`s Tuono Ferragne`s Tuono Ferragne`s Tuono Tuono Genco Nikitskii Princesse Primorskii Garrigues Tardy Nonpareil Tuono
Early (29) Middle (38) Middle (41) Middle (42) Late (46) Late (47) Late (52) Late (54) Late (55) Very late (58) Very late (66)
Shell hardnessb Very hard (28) Very hard (29) Very hard (29) Very hard (26) Hard (32) Hard (35) Hard (35) Hard (35) Soft (55) Semi-hard (37) Very hard (24)
a
In parentheses: average number of Julian days (days after January 1st) for full blooming (50% flowers opened) for 3 years. b In parentheses: [weight of seed/weight of (seed + endocarp)] 100.
- S5133 crossed with different male parents, then stratifying the seeds with endocarp in 1999 (crossed with Marta, Anton˜ eta, Lauranne, and R1000), or without endocarp in 2000 (crossed with Desmayo Largueta, A2198, and R1000). - Reciprocal crosses (Peraleja and S2332, Primorskii and Lauranne, S5133 and Lauranne), stratifying the seeds with endocarp, in 1998. Mature nuts from the different crosses were collected during the summer and kept at room temperature. In winter, seeds (with or without endocarp) were treated with a 2% TMTD1 (tetramethilthiuram disulfide) fungicide solution for 30 min, and placed in plastic mesh bags in moist vermiculite in a chamber at 7 8C. The particle size of the vermiculite was 2 mm 2 mm. Seed germination was defined as radicle emergence. For each treatment during the stratification period, the number of germinated seeds was recorded weekly until germination was completed. 2.3. Statistical analysis To analyse the differences between the families studied, an analysis of variance of the data for each assay was carried out, and Duncan’s multiple range test applied to identify significant differences between means (SAS Institute, 1989).
3. Results 3.1. Stratification requirements of almond seeds with endocarp The stratification period required for almond seeds with endocarp differed among crosses of Lauranne and S5133 with different pollinizers (Table 3). In Lauranne crossed with several male parents, the mean stratification requirements ranged between 7.0 and 8.4
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Table 2 Mean period in stratification (weeks) required for germination of seeds from different crosses Mean perioda
C.V.
7.2cd 7.0d 8.4a 7.4bc 8.2a 7.7b
15.7 16.1 16.6 16.5 17.8 16.1
6.3b 7.7a 7.5a 7.4a
25.5 27.7 23.2 21.3
A female parent with different male parents (seeds without endocarp) S5133 Desmayo Largueta 26 2–11 A2198 187 2–11 R1000e 17 3–9
5.6a 5.9a 5.2a
47.0 41.2 44.0
Reciprocal crosses (seeds with endocarp) Peraleja S2332 S2332 Peraleja
Female
Male
N
Range of weeks
A female parent with different male parents (seeds with endocarp) Lauranne Desmayo Largueta 279 5–11 Marcona 107 5–11 Lauranne 50 6–10 Open pollination 177 5–11 147 6–11 Primorskiib S5133c 68 5–10 S5133
Marta Anton˜ eta Lauranned R1000e
180 123 97 137
5–12 5–12 5–12 5–11
59 37
5–10 5–7
6.6a 5.5b
13.7 11.0
Primorskii Lauranne
Lauranne Primorskiib
73 147
4–11 6–11
6.9b 8.2a
23.4 17.8
Lauranne S5133
S5133c Lauranned
68 63
5–10 4–11
7.7a 6.6b
16.1 26.1
Number of seeds stratified (N), range of weeks in stratification (minimum–maximum), mean period in stratification, and coefficient of variation (C.V.). a Values followed by the same letter show no significant differences by the Duncan multiple range test (P < 0.05). b Same data. c Same data. d Same cross in a different year. e Same cross with and without endocarp.
weeks (Table 2). Stratification duration required for germination in the early flowering cultivars (Desmayo Largueta and Marcona) was shorter than for late flowering cultivars (Lauranne, Primorskii, and S5133). Moreover, almond seeds from the crosses with Desmayo Largueta needed only 0.5 weeks less to germinate than the seeds from the cross with S5133, despite the fact that the flowering dates of these cultivars differ by 4 weeks. In contrast, seeds from the cross with Lauranne (selfing) showed longer stratification requirements than those from the cross with S5133, with a date of blooming 6 days later. In cultivar S5133 pollinated with several male parents, the mean period required for germination ranged between 6.3 and 7.7 weeks. Duncan’s test identified only two groups, a group of faster germination (with Marta as pollinizer) and a second group with the other three late flowering pollinizers (Table 2). This grouping was not related to flowering date of the pollinizers. It is noteworthy that there were no significant differences in the
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Table 3 Analysis of variance of the mean period in stratification (weeks) required for germination of seeds from different crosses F-value
Pr > F
Lauranne with different male parents (seeds with endocarp) Families 36.44 Error 1.53
23.73
0.0001
S5133 with different male parents (seeds with endocarp) Families 59.79 Error 3.02
19.75
0.0001
S5133 with different male parents (seeds without endocarp) Families 4.35 Error 6.02
0.72
0.4862
Reciprocal crosses (Peraleja x S2332) (seeds with endocarp) Families 28.19 Error 0.64
43.43
0.0001
Reciprocal crosses (Primorskii Lauranne) (seeds with endocarp) Families 91.30 Error 2.28
39.88
0.0001
Reciprocal crosses (Lauranne S5133) (seeds with endocarp) Families 35.47 Error 2.24
15.81
0.0001
Source
Mean square
stratification requirements of seeds from the crosses with the cultivars Anton˜ eta, Lauranne, and R1000, with very different flowering dates. 3.2. Stratification requirements of almond seeds without endocarp In seeds without endocarp, no differences were observed between S5133 with different pollinizers, in spite of the great range of flowering dates of the pollinizers (Table 3). In general, endocarp removal reduced the mean stratification requirements. In the crosses between S5133 and R1000, with and without endocarp, endocarp removal reduced the mean stratification requirements by 2.2 weeks (Table 2). 3.3. Reciprocal crosses The results of three reciprocal crosses showed significant differences between each pair (Tables 2 and 3). Seeds from crosses with late-flowering cultivars as the female parent (Primorskii with Lauranne, and S5133 with Lauranne) showed shorter stratification requirements for germination. 3.4. Variability of almond seed stratification requirements Seeds exhibited a range of stratification requirements between 4 and 12 weeks in tests with endocarp, and between 2 and 11 weeks in tests without endocarp (Table 2). The
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coefficients of variation reflected this large variability. These coefficients were smaller in the cultivars with a hard endocarp (11% from the cross between S2332 and Peraleja) and larger in the tests without endocarp (up to 47%) (Table 2). In the cross S5133 Lauranne, we measured a difference in the mean stratification requirements of almost 1 week (6.6 and 7.5 weeks, respectively) between the 2 years studied (Table 2).
4. Discussion 4.1. Effect of the pollinizer Although seeds with endocarp of a cultivar with different pollinizers showed different germination rates after stratification, these differences were not related with pollinizer flowering date. No differences were measured for seeds without endocarp, probably due to the washing out of the germination inhibitors of the integument. In contrast, Kester (1969) assessed that there was an important influence of pollinizer. However, re-analyzing his data, although differences between families with different pollinizers were observed, these differences were frequently not related to the date of flowering of pollinizer. Only when Kester (1969) crossed almond (in general, earlier flowering) by peach (later flowering), this relationship was narrower. We believe that in this case, the high requirements of the seeds to germinate could be partially due to its interspecific nature. Thus almond peach seeds germinate later than almond almond. However, among the almond almond families, data did not show a strong influence of pollinizer. Kester (1969) explained these differences as being due to an incapacity of certain cultivars to transmit this characteristic. Grigorian (1972) attempted to confirm the results of Kester (1969), but his interpretation of results was questionable. Again, differences were observed between families, but most of the time, they were not related to pollinizer flowering date. To explain the numerous exceptions, Grigorian (1972) assessed that some cultivars could have low chilling requirements to flower, but high for seed germination. Differences in the stratification requirements for germination appear to be due to differences in the endocarp–embryo interaction (shell hardness, shell seal, permeability, exchange of water and gases, genotype, etc.). The pollinizer could affect the nature of the endocarp and thus be a factor affecting seed germination. In this respect, Kumar and Das (1996) described an effect of the pollinizer (xenia) on the nut size and ripening date. Other factors affecting endocarp properties could be the position and the characteristics of the branch where the different crosses were carried out. 4.2. Consanguinity Seeds from the self-pollination of Lauranne showed the longest stratification requirements. This could be explained as collateral effects of the endogamy (inbreeding), which would produce a delay in the development of the embryo. Similar problems, including deficiencies in the endosperm, low germination rate, and less radicle growth,
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have been reported previously in almond (a typically self-incompatible species) seeds resulting from self-pollination, and could be related to the accumulation of unfavourable genes in the inbred descendants (Grasselly and Olivier, 1988; Oukabli et al., 2001). 4.3. Reciprocal crosses The significant differences observed in the tests from reciprocal crosses indicated unequal effects of the female and male parents on this characteristic. If the influences of the female and the male were the same, there would have been no differences in the reciprocal crosses, as Dicenta and Garcı´a (1993) previously indicated for other traits in almond, such as flowering date. In our crosses, the stratification requirements were different in the direct and the inverse crosses, although it is necessary to point out the different hardness of the shell in both cases. The results seem to indicate that the seeds of a female parent with a softer shell germinated faster, with less stratification requirements, independent of the chilling requirements for flowering. 4.4. Variability of stratification requirements The large variability observed within cultivars could be due to different concentrations of germination inhibitors in the seed coat, different genetic nature of the embryo of each seed, or to differences in shell hardness or permeability to water and gases. A relationship has been described between the chilling requirements of vegetative and floral buds for sprouting, and the stratification requirements of seeds for germination in peach and almond (Powell, 1987; Garcı´a-Gusano et al., 2004). However, the differences in the requirements of buds are larger than the stratification requirements for germination. Almond cultivars with differences in flowering time of around 1 month only showed differences of 1 week in the stratification requirements for germination. The endocarp could have a mechanical effect, preventing almond seed imbibition and the washing out of the seed coat hormonal inhibitors (Du Toit et al., 1979), thus modifying the stratification period required for germination and contributing to the large variability observed within cultivars. Gradziel and Martı´nez-Go´ mez (2002) described great differences in the shell seal of nuts and the lignin density (hardness) in different samples from the same almond cultivar. These differences could affect the exchange of water and gases, and have a different mechanical effect, affecting seedling radicle growth. In all cases, elimination of the endocarp reduced the stratification requirements for germination. Similar results have been observed in other almond cultivars (Grigorian, 1972; Garcı´aGusano et al., 2004).
5. Conclusions Different stratification requirements were measured for germination in the different crosses. A great variability in the stratification requirements for germination between seeds from the same cross was observed also. However, the flowering time of the pollinizer
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(chilling requirements of flower buds for sprouting) did not appear to influence these stratification requirements, what seems to be determined mainly by the female parent. To deepen our knowledge of the great variability observed and the influence of the pollinizer in the stratification requirements of almond seed germination, further studies must be performed on interaction of endocarp end embryo.
Acknowledgements The authors thank Mariano Gambı´n for technical assistance in the experimental work presented in this paper. The work has been financed by the project AGL2001-1054-C03-01 entitled ‘‘Mejora Gene´ tica del Almendro’’ from the Spanish Ministry of Science and Technology. The authors also acknowledge the support of the Spanish Ministry of Science and Technology for the contract of Dr. P. Martı´nez-Go´ mez.
References Chao, L., Walker, D.R., 1966. Effects of temperature, chemicals, and seed coat on apricot and peach seed germination and growth. J. Am. Soc. Hort. Sci. 88, 232–238. Dicenta, F., Garcı´a, J.E., 1993. Reciprocal crosses in almond. Plant Breeding 110, 77–80. Du Toit, H.G., Jacobs, G., Strydom, D.K., 1979. Role of various seed parts in peach seed dormancy and initial seedling growth. J. Am. Soc. Hort. Sci. 104, 490–492. Frisby, J.W., Seeley, S.D., 1993. Chilling of endodormant peach propagules: comparison between seeds and seedlings. J. Am. Soc. Hort. Sci. 118, 269–273. Garcı´a-Gusano, M., Martı´nez-Go´ mez, P., Dicenta, F., 2004. Breaking seed dormancy in almond (Prunus dulcis (Mill.) D.A. Webb). Sci. Hort. 99, 363–370. Gonza´ lez-Cepeda, I.A., 1975. Dormancy in almond seeds: a study in relation to stratification temperature and growth regulator levels. M.Sc. Dissertation. University of California, Davis, USA, 40 pp. Gradziel, T.M., Martı´nez-Go´ mez, P., 2002. Shell seal breakdown in almond is associated with the site of secondary ovule abortion. J. Am. Soc. Hort. Sci. 127, 69–74. Grasselly, C., Olivier, G., 1988. Phenomenes d’inbreeding dans les descendances issues d’autofecundation chez l‘amandier, 7 Colloque du GREMPA (Reus), Junio 1987, pp. 73–78. Grigorian, V., 1972. L’embryogene`se chez l’Amandier (Prunus amygdalus Batsch) e´ tude compare´ de la dormances des graines et de la dormances des bourgerons ve´ ge´ tatifs. PhD Dissertation. University of Bourdeaux, Bourdeaux, France, 144 pp. Kester, D.E., 1969. Pollen effects on chilling requirements of almond and almond-peach hybrid seeds. J. Am. Soc. Hort. Sci. 94, 318–321. Kumar, K., Das, B., 1996. Studies on xenia in almond (Prunus dulcis (Miller) D.A. Webb). J. Hort. Sci. 71, 545– 549. Martı´nez-Go´ mez, P., Dicenta, F., 2001. Mechanisms of dormancy in seeds of peach (Prunus persica (L.) Batsch) cv. GF305. . Sci. Hort. 91, 51–58. Mehanna, H.T., Martin, G.C., Nishijuma, C., 1985. Effects of temperature, chemical treatments and endogenous hormone content on peach seed germination and subsequent seedling growth. Sci. Hort. 27, 63–73. Mehlenbacher, S.A., Voordeckers, A.M., 1991. Relationship of flowering time, rate of seed germination, and time of leaf budbreak and usefulness in selecting for late-flowering apples. J. Am. Soc. Hort. Sci. 116, 565–568. Oukabli, A., Lansari, A., Loudiyi, W., Abousalim, A., 2001. Effets endogamiques sur la germination et la croissance de semis du cultivar autocompatible Tuono (Prunus dulcis). Fruits 56, 197–205.
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99
Powell, L.E., 1987. Hormonal aspects of bud and seed dormancy in temperate-zone woody plants. Hort. Sci. 22, 845–850. SAS Institute, 1989. SAS/STAT User’s Guide, SAS Inst., Cary, NC. Seeley, S.D., Ayanoglu, H., Frisby, J.W., 1998. Peach seedling emergence and growth in response to isothermal and cycled stratification treatments reveal two dormancy components. J. Am. Soc. Hort. Sci. 123, 776–780.
