Scientia Horticulturae, 25 (1985) 247--254 247 Elsevier Science Publishers B.V., A m s t e r d a m - Printed in The Netherlands

EFFECT OF SEED COAT ON PEACH SEED GERMINATION

HUSSEIN T. MEHANNA' and GEORGE C. MARTIN

Department of Pomology, University of California, Davis, CA 95616 (U.S.A.)

(Accepted for publication 16 October 1984)

ABSTRACT

Mehanna, H.T. and Martin, G.C., 1985. Effect of seed coat on peach seed germination. Scientia Hortic., 25: 247--254.

Various treatments to remove the seed coat surgically revealed its role in preventing germination of non-stratified peach seed. The peach embryo from a non-chilled seed is not dormant, as it germinates readily when the seed coat is completely removed. The peach seed coat provides a physical impediment -- commonly referred to as me- chanical resistance to germination. Incision through the seed coat around the seed at the cotyledonary gap allowed germination even though the seed coat remained in contact with the embryo. No role for inhibitors extracted from the seed coat could be elucidated.

Keywords: dormancy; inhibitors; Prunus persica; rest; seed coat.

ABBREVIATION

ABA = abscisic acid.

INTRODUCTION

Peach seeds require suitable chilling while moist in order to germinate and produce normal seedlings (Flemion, 1934). Several workers have been able to germinate seeds without chilling by removal (Tukey, 1933; Flemion, 1934; Lammerts, 1942) or puncturing (Chao and Walker, 1966) of the seed coat; yet in most of these investigations, resultant seedlings were stunted. Peach seedlings arising from chilled embryos with intact seed coats were taller and had a greater dry weight of shoots, leaves and roots than seedlings arising from non-chilled embryos without seed coats (du Toit et al., 1979). In non-chilled peach seed, investigators have suggested that a seed coat inhibitor prevents germination (Lipe and Crane, 1966; Sharma and Singh, 1980). Leaching peach seed in water, which is purported

'On sabbatic leave from the Pomology Department, Faculty of Agriculture, Cairo Uni- versity, A.R. Egypt.

0304-4238/85/$03.30 © 1985 Elsevier Science Publishers B.V.

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to remove the inhibitor (Lipe and Crane, 1966), will not necessarily improve germination (Chao and Walker, 1966).

The purpose of the present investigation was to test the hypothesis that germination of non-chilled peach seed was prevented by the physical im- pediment -- commonly referred to as mechanical resistance -- offered by the seed coat rather than to inhibitors present in the seed coat. This hypothesis does not exclude the internal chemical features regulating em- bryo growth; we sought to separate the role of the seed coat from that of the embryo. The test system included a long and short chilling peach seed and refined seed coat treatments which had previously been found to induce germination in non-chilled embryos.

M A T E R I A L S AND M E T H O D S

S e e d s o u r c e a n d g e r m i n a t i o n . - - Experiments were conducted with 'Nema- guard' peach seeds which require about 4 weeks chilling and 'Halford' which require about 12 weeks chilling at 7°C. Both cultivars were obtained from sources in California where cross-pollination was unlikely; it is pre- sumed tha t these seeds were self-pollinated. Seeds with the endocarp re- moved were soaked in water for 24 h at 20 -+ 2 ° C, and placed in a 2% Tersan 75 (E.I. du Pont de Nemours and Co., Inc., Biochemicals Department, Wilmington, Delaware, U.S.A.) fungicide solution for 1 h. Seeds were not chilled prior to experimentation. For each experiment, 3 samples of 10 seeds each were placed in petri dishes on filter paper, moistened with 5 ml distilled water and held at 20 + 2 ° C in the dark. Germination was indexed by radicle protrusion through the seed coat; data were recorded every 3 days for a 15-day period.

T r e a t m e n t o f s e e d coa t s . - - Different seed tissues and portions of seed tissues were removed to assess their role in germination (Table I). A puncture

T A B L E I

Effect of seed tissue on the percentage of peach seed germinating 15 days after treat- menP

Seed t r e a t m e n t 2 ' N e m a g u a r d ' ' H a l f o r d '

Seed coat a t t a ched 20 a Entire seed coat r em oved 100 c Chalazel hal f of each c o t y l e d o n r em oved 100 c One entire cotyledon r em oved 97 c Seed coat cover ing mic ropy la r end of e m b r y o r em oved 97 c P u n c t u r e through seed coa t at m ic ropy la r end 37 b Rinsing in wate r for 72 h w i th seed coat a t t a c h e d 13 a

0 a 90 b 93 b 87 b 90 b 1 3 a

7 a

~Mean separation in co lumns b y Duncan's multiple range test, 5% level. 2Following treatment, seeds were soaked in wa te r for 24 h.

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with a pin was made through the seed coat at different locations before or after soaking in water to determine the effect of location of the puncture on germination (Table II).

Zero, 25, 50, 75 and 100% of the seed coat was removed from around the micropylar or chalazel end of the seed to determine the role of the seed coat in germination. In another experiment, incisions were made through the seed coat with a razor blade to determine the possible me- chanical restrictions of the seed coat on germination. These treatments included the following: (a) seed coat removed except for a piece encircling the seed at the cotyledonary gap; (b) an incision made along the c otyledonary gap completely around the seed; (c) one-half of the longitudinal section removed from the seed coat; (d) an incision made at right angles to the micropylar end; (e) control; (f) a section removed at right angles to the micropylar end (Fig. 1).

T A B L E II

Effect of seed coat puncturing on the percentage of peach seed germinating 15 days after treatment I

T r e a t m e n t 2 ' N e m a g u a r d ' 'Ha l fo rd '

Control 7 a Puncture chalazel end before soaking 3 a Puncture chalazel end after soaking 3 a Puncture micropylar end before soaking I0 a Puncture micropylar end after soaking I0 a Puncture at micropylar tip (apex of seed) before soaking 30 b Puncture at micropylar tip (apex of seed) after soaking 30 b

0 a 3 a 3 a 3 a

10 a 1 0 a 1 0 a

~Mean sepa ra t i on in co lumns b y D u n c a n ' s mul t ip le range test , 5% level. 2Before or a f t e r t r e a t m e n t , seeds were soaked in wa te r for 24 h. All p u n c t u r e s were m a d e t h r o u g h t h e seed coat .

E f f e c t o f the seed coat e x t r a c t or A B A on germinat ion . - - Seed coats were removed from non-chilled seeds of each cultivar and extracted with cold methanol for 48 h at 4°C. The methanol was evaporated, and the extract was diluted with distilled water to provide solutions of extracts equivalent to a given number of seed coats per ml. Three replicates of 5 ml of extract and 10 or 100 mg 1-1 ABA were placed on filter paper in petri dishes and 10 non-chilled seeds of each cultivar with seed coats removed were added to each dish and incubated at 18°C in the dark for germination.

In all experiments, data were evaluated by Duncan's multiple range test.

RESULTS

E f f e c t o f seed tissues on germinat ion . - - Little or no improvement in seed germination occurred by rinsing in water for 72 h compared to germination

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a b

C d

Fig. 1. Illustrations of incisions made on seed coats to relieve mechanical resistance. (a) Seed coat removed except for a piece encircling the seed at the cotyledonary gap; (b) an incision made along the cotyledonary gap completely around the seed; (c) one- half of the longitudinal section removed from the seed coat; (d) an incision (see arrows) made at right angles to the micropylar end; (e) control; (f) a section removed at right angles to the micropyl.ar end.

of controls which had an intact seed coat (Table I). Removing the entire seed coat, the chalazel half of each cotyledon, 1 entire cotyledon or the seed coat covering the micropylar end led to nearly 100% germination for both cultivars (Table I).

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Effect o f seed coat puncturing location on germination. -- None of the seed coat puncture treatments increased germination percentage over that of controls for 'Halford' (Table II), but there was a significant increase in 'Nemaguard' germination of 30%, which occurred with puncturing at the micropylar tip only (Table II).

Effect o f the amount and location o f seed coat removed on germination. -- Removing 25% of the seed coat from the micropylar end of the seed led to near maximum germination percentage for both cultivars (Table III). Removal of larger portions of the seed coat did not increase germination percentage for either cultivar (Table III). In contrast, removing 25% of the seed coat from the chalazel end did not increase germination for either cultivar (Table III): Removal of 50, 75 and 100% of the seed coat from the chalazel end led to a steady increase in germination percentage. For most treatments, germination percentage of 'Nemaguard' was slightly greater than that of 'Halford' (Table III).

TABLE III

Effect of amount and location of seed coat removed on the percentage of peach seed

germination

Seed coat removed (%) 'Nemaguard' 'Halford'

Micropylar Chalazel Micropylar Chalazel

0 0 0 0 0 25 95 0 80 10 50 95 50 90 20 75 95 80 80 80

100 95 95 90 90

T A B L E IV

Ef fec t o f seed coa t inc i s ion o n peach seed g e r m i n a t i o n pe rcen tage '

T r e a t m e n t 2 ' N e m a g u a r d ' 'Ha l fo rd '

Control (seed coat attached) 20 a Incision at right angle to embryonic axis 30 a Section of seed coat removed at right angle to

embryonic axis 25 a Incision through seed coat along cotyledonary

gap completely around seed 95 b Removed one-half longitudinal section of seed coat 95 b Removed seed coat except piece encircling seed

at cotyledonary gap 90 b

4 a 20 b

17 b

85 b 80 c

7 5 c

1Mean s e p a r a t i o n in co l um ns b y D u n c a n ' s mu l t i p l e range test , 5% level. ~Following t r e a t m e n t , seeds were soaked in wa te r for 24 h.

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E f f e c t o f seed coat incision on germinat ion . - - Neither the incision at right angles to the embryonic axis nor removal of a section of seed coat at the same location significantly increased germination percentage for either cultivar (cf. Fig. 1 with Tabel IV). A significant increase in germination for both cultivars occurred when an incision was made along the cotyl- edonary gap encircling the seed, one-half of the longitudinal section of the seed coat was removed, or the seed coat was removed except for the thin piece encircling the seed at the cotyledonary gap (cf. Fig. 1 with Table IV).

E f f e c t o f seed coat ex t rac t s or A B A on germinat ion . - - Abscisic acid applied at 10 mg 1-1 and methanol extracts of either 'Halford' or 'Nemaguard' seed coats applied to embryos at a ratio (seed coat extract:seed) of 1:1 inhibited germination of both cultivars (Fig. 2). Extracts taken from either cultivar were effective on the other cultivar but greater inhibition resulted from 'Halford' extracts, and 'Halford' was more sensitive to extracts from either source. Abscisic acid applied at 100 mg 1-1 or 'Halford' extract applied at a ratio of 2:1 completely prevented germination (Fig. 2).

,oo

8O

6 0 o

~ 4o

20

'NEMAGUARD'

'HALFORD'

CONT 2:1 I:1 2:1 I : I ABA ABA H. EX H.EX N.EX N. EX I0 I00

mg I-I TREATMENTS

Fig. 2. Effect of 'Nemaguard' or 'Halford' seed coat extracts or ABA on the germina- tion of seeds which have their seed coat removed. 'Halford' extract (H.EX), 'Nemaguvxd' extract (N.EX), extracts from 2 seed coats applied to I embryo (2:1), and extracts from 1 seed coat applied to I embryo (1:1).

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DISCUSSION

Clearly, as shown by others (Tukey, 1933; Lammerts, 1942) and in our own work, the peach embryo is capable of germination wi thout cold t reatment providing the seed coat is removed. As shown in our studies, both 'Halford' which has a high-chilling requirement, and 'Nemaguard' which has a low-chilling requirement, germinate wi thout cold t reatment if the seed coat is removed. The question remains: Is the seed coat a me- chanical or a chemical barrier to germination? If the inhibition of germina- tion were strictly chemical, then one would expect that leaching would remove the inhibitor -- were it water soluble, e.g. ABA (Chao and Walker, 1966; Lipe and Crane, 1966). We were unable to remove a water-soluble germination barrier with 72 h of leaching. An inhibitor not soluble in water could be present. However, the argument for an inhibitor still seems weak for the following reasons: removing 25% of the seed coat at the micropylar end led to near maximum germination, thus 75% of the seed coat con- taining a conjectured inhibitor was still present; removing 25% of the seed coat from the chalazel end of the seed did not improve germination; and, perhaps the best evidence against a seed coat inhibitor, the incision through the seed coat along the cotyledonary gap encircling the seed led to near maximum germination percentage. The latter t reatment left the seed coat in immediate contact with the embryo, thus allowing chemical movement from the seed coat to the embryo while relieving mechanical restriction at the micropylar end. The location of the incision was critical, i.e. along the cotyledonary gap around the seed where the radicle emerges. This t reatment reduced the barrier through which the radicle grows. By what mechanism could any of the above treatments reduce inhibitor content? In each of the critical treatments, the same amount of seed coat remained in close contact with the micropylar end of the seed. If a seed coat inhibitor were a factor, on what basis would its action be terminated by seed coat incision or puncture at the micropylar end of the seed?

As chilling takes place, the moist seed coat may degrade and offer less mechanical resistance to the radicle. Further, the capacity for embryo growth may be enhanced because of chemical transformations which occur in the embryo during chilling. These transformations could increase the capacity of the radicle to overcome mechanical restriction of the seed coat and ensure vigorous epicotyl development.

If a seed coat inhibitor were controlling germination of the non-chilled peach seed, then we would expect larger quantities of the inhibitor to be present in seed requiring longer chilling. In fact as tested, the methanol extract from long-chill-requiring 'Halford' reduced germination less in 'Nemaguard' than in 'Halford'. Similarly, the short-chilling 'Nemaguard' methanol extract reduced germination less in 'Nemaguard' than in 'Halford'. Clearly, non-chilled 'Halford' has a lower capacity for germination and is far more sensitive to unfavorable t reatment than 'Nemaguard'. Methanol

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extracts of tissues are diff icul t to interpret because of the wide range o f c o m p o n e n t s tha t are methanol-soluble and because prior to seed coat ex- t ract ion, compar tmen ta l i za t ion effect ively prevents m a n y chemicals f rom entering sensitive react ion sites.

While we have n o t el iminated the potent ia l role for seed coat inhibitors, we have posit ively presented the case for a mechanical resistance role o f the seed coat. We suggest tha t the seed coat offers mechanical resistance to radicle pro t rus ion and tha t subsequent to germinat ion, chemical factors in the e m b r y o prevent normal seedling growth. These exper iments reveal tha t little or none of the co ty l edon is necessary for peach seed germinat ion.

REFERENCES

Chao, L. and Walker, D.R., 1966. Effect of temperature, chemicals and seed coat on apricot and peach seed germination and growth. Proc. Am. Soc. Hortic. Sci., 88: 232--238.

Du Toit, H.J., Jacobs, G. and Strydom, D.K., 1979. Role of the various seed parts in peach seed dormancy and initial seedling growth. J. Am. Soc. Hortic. Sci., 104: 490--492.

Flemion, F., 1934. Dwarf seedling from non-after ripened embryo of peach, apple, and hawthorn. Contrib. Boyce Thompson Inst., 6: 205--209.

Lammerts, W.E., 1942. Embryo culture an effective technique for shortening the breed- ing cycle of deciduous trees and increasing germination of hybrid seeds. Am. J. Bot., 29: 166--171.

Lipe, W.N. and Crane, J.C., 1966. Dormancy regulation in peach seeds. Science, 153: 541--542.

Sharma, H.C. and Singh, R.N., 1980. Effect of stratification temperature and duration on the level of endogenous inhibitor and its relationship with dormancy in seeds of subtropical peach (Prunus persica stock) cv. Sharbati. Indian J. Plant Physiol., 23: 26--32.

Tukey, H.B., 1933. Effect of photoperiod and temperature on growth of embryo cul- tured peach seedlings. Am. J. Bot., 30: 707--711.