Planta
Planta 147, 444-450(1980)
9 by Springer-Verlag 1980
Fruit-set of Unpollinated Ovaries of Pisum sativum L. Influence of Vegetative Parts Juan Carbonell and Jos6 L. Garcia-Martinez* Instituto de Agroqulmica y Tecnologla de Alimentos, C.S.I.C., Jaime Roig 11, Valencia-10, Spain
Abstract. The influence of removing the apical shoot and different leaves above and below the flower on the fruit-set of unpollinated pea ovaries (Pisum satirum L. cv. Alaska) has been studied. Unpollinated ovaries were induced to set and develop either by topping or by removing certain developing leaves of the shoot. Topping had a maximum effect when carried out before or on the day of anthesis, and up to four consecutive ovaries were induced to set in the same plant. The inhibition of fruit-set was due to the developing leaves and not to the apex. The third leaf above the first flower, which had a simultaneous development to the ovary, had the stronger inhibitory effect on parthenocarpic fruit-set. The application of different plant-growth regulators (indoleacetic acid, naphthylacetic acid, 2,4-dichlorophenoxyacetic acid, gibberellic acid, benzyladenine and abscisic acid) did not mimic the negative effect of the shoot. Keywords: Abscisic acid Auxin - Cytokinin Decapitation - Fruit-set -- Gibberellin - Parthenocarpy - Pisum.
Introduction Fruit-set has been defined as the changeover from the static condition of the flower ovary to the rapidly growing condition of the young fruit (Leopold and Scott, 1952). There is a general assumption that this change is initiated by hormones either furnished by * Present address: Plant Growth Laboratory, Wickson Halt, University of California, Davis, CA 95616, USA Abbreviations: CCC = (2-chloroethyl)trimethylammoniumchloride;
MH=maleic hydrazide; IAA=indole-3-acetic acid; N A A = l-naphthaleneacetic acid; 2,4-D=2,4-dichlorophenoxyacetic acid; GA3=gibberellic acid; 6-BAP=benzyladenine; ABA=abscisic acid
the pollen, or produced by the developing seeds, or present in the ovules or pericarp in parthenocarpic varieties (Nitsch, 1970). Parthenocarpic fruit development can be induced by the application of plantgrowth regulators (Crane, 1964) or under certain environmental conditions (Gustafson, 1942). Vegetative development competes with fruit development and fruit-set can be increased by removing vegetative parts. The developing seeds seem to attract nutrients to the ovary and produce a sink which is able to compete with vegetative sinks (Bollard, 1970). Eeuwens and Schwabe (1975) showed that a close relationship exists between changes in extractable hormones and growth rates of pea pod walls and seeds. They used plants which had been topped immediately above the second flowering node. When we used the same model of plant in experiments with emasculated flowers we observed that the ovaries always set irrespective of the kind of treatment applied to the flowers. The aim of the present work was to study the influence of the vegetative parts on the development of the unpollinated ovary. We have shown that parthenocarpic fruit development may be induced by topping pea plants and that the inhibitory effect of the shoot on fruit-set can not be substituted by different plant-growth regulators.
Materials and Methods Plant Material
Seeds of Pisum sativum L. cv. Alaska were germinated and grown for 7 days in "coarse" sand (consisting of 60% (w/w) of 1-2 mm and 26% (w/w) of 0.5-1 mm diameter particles) at or below 25 ~ C under sunlight conditions and irrigated with deionized water. Seedlings were selected for uniformity and transferred into 11.5 cm diameter pots containing "fine" sand (consisting of 58% (w/w) of 0.5-1 mm and 30% (w/w) of 0.25-0.5 mm diameter particles). Both types of sand were usually treated with 1 M HCI for a week
0032-0935/80/0147/0444/$01.40
J. Carbonell and J.L. Garcia-Martinez: Fruit-set of Unpollinated Ovaries
LEAVES ~ FLOWERS 2
a
1
2
~
1
-1
-2
445
Table 1. Values of i and j Position of leaves removed
j value
i value
None 1 2 3 12 13 23 123 123etc.
1 2 3 4 5 6 7 8 9
0 1 0 2 0 1 0 1 034 024 014 04 0
2 3 3 2
3 4 4 4 4
'-3 , ~
9 Fig. 1. Numeration of leaves and flowers in a pea plant
and then washed repeatedly with deionized water; the "'fine" sand was then equilibrated with nutritive solution for a week by daily changing the solution. The nutritive solution was Hoagland n ~ 1, containing the oligoelements of Hewitt (1966), and was automatically supplied to the plants each hour. Plants were grown through the year in a heated glasshouse, except in January, July, August, and December. Natural light was supplemented with Sylvania "daylight" fluorescent tubes (about 2,000 lx at the level of the first flowers) for a 16 h photoperiod. The temperature fluctuated according to the environment, but extremes were never higher than 30 ~ C or lower than 12~ C. The first flowers opened between 28 and 32 days after sowing. Differences in the development of the plants were observed, the more vigorous being obtained between the begining of March and the end of May. The fruit length and percentage of fruit-set appeared to depend to a small degree on light intensity and temperature, so the same experiment was repeated in different seasons. The results given in this paper come usually from the Spring experiments. Fruits shorter than 25 mm 12 days after anthesis were occasionally obtained but they were not taken into account because we had observed previously that they degenerated within a few9 days. Figure 1 gives a scheme of the positions of leaves and flowers in the plants. Flowers were numbered according to the sequence in which they appeared. Leaves were numbered by taking the leaf placed at the node of the first flower as a reference (leaf 0) ; upper leaves were numbered positively and lower leaves negatively. Except in the experiments corresponding to Figs. 3 and 4, we always used only the first flower of each plant. Other flowers were removed as soon as possible. Unpollinated flowers were obtained by removing petals and stamens two days before anthesis. At this stage the petals had approximately the same length as the sepals. Topping was carried out by cutting the internode indicated in each experiment. Axillary shoots and new flowers were always removed as they appeared to eliminate competition problems.
0.1% Tween 80: AMO- 1618 (Calbiochem), (2-chloroethyl)trimethylammonium chloride (CCC) (Cyanamid), and maleic hydrazide (MH) (MH-30, Naugatuk Chemical). Indoleacetic acid (IAA) (Merck), naphthaleneacetic acid (NAA) (Fluka), 2,4-dichlorophenoxyaeetic acid (2,4-D) (Fluka), gibberellic acid (GA3) (Fluka), benzyladenine (6-BAP) (Fluka) and abscisic acid (ABA) (Sigma) were dissolved in a small amount of methanol or ethyl acetate and dispersed in melted anhydrous lanolin at 60 ~ C. The lanolin paste was applied with a glass rod to the cut surface of the topped stem.
Statistical and Mathematical Methods Statistical treatments of the data were made by using the analysis of variance and the Tukey test for comparison of mean values after calculating the least significant difference (LS.D.). In order to determine the effect of each leaf above the first flower on fruit-set, it was supposed that these leaves could have a positive or activator effect (A~) and a negative or inhibitor effect (It) where i indicates the leaf position and takes values from 0 to 4 (values 0 and 4 correspond to all the leaves under leaf i and above leaf 3, respectively). By assuming that these effects are additive, we obtained an equation system for each day of the experiment shown in Fig. 7. The algebraic sum of activator and inhibitor effects of each non-removed leaf was set equal to the number of non-degenerated ovaries (Ni) that day; j represents each of the 9 possibilities obtained when no leaf was removed, or one, or several leaves above leaf n ~ 0 (Fig. 7). The general equation for each day is:
(A,-Ie)+SVj=5~ i=y(j)
A,>=O j = l . . . 9 I i =>0 i =f(j) = 0... 4
Nj_>0 svj~0 where SVj is a slack variable which represents all kinds of errors present in the experiment and i is a function of j, according to Table 1. Solutions for each equation system were obtained by minimizing the associated lineal function formed by the sum of the absolute values of the slack variables, that is, the sum of errors. The lineal programs formed were solved by a computer.
Results Application o f Plant Growth Regulators Plant-growth retardants and inhibitors were applied by dipping the top part of the plant, including leaf n ~ 1 in the solution. The following compounds were used in aqueous solution containing
Topping of plants above the first flower between days -2 and 8 after anthesis induced parthenocarpic f r u i t - s e t o f u n p o l l i n a t e d p e a o v a r i e s ( F i g . 2, i n s e r t ) .
446
J. Carbonell and J.L. Garcia-Martinez: Fruit-set of Unpollinated Ovaries
~Ioo~ 80-
.-_
,0-1 I
-2
70-
~
I x
I
topping
1
)4 6 8
0 2 4 Day of topping
80- ~
day of
~......~
e~
/
~
70-
~
_ ~o.,~-I , - e~'-'~ >'0
~60E 50r
./
60.
o
50"
3 z.
u. 30-
20.
/ # ///o/
/ / / / ' / / / . oo z//I/// e
I I
!:!:!:!:!:!:!::
~;i~iiiiiiiiil
~o__O_oO:%0 i
-
.............
Planta 147, 444-450(1980)
9 by Springer-Verlag 1980
Fruit-set of Unpollinated Ovaries of Pisum sativum L. Influence of Vegetative Parts Juan Carbonell and Jos6 L. Garcia-Martinez* Instituto de Agroqulmica y Tecnologla de Alimentos, C.S.I.C., Jaime Roig 11, Valencia-10, Spain
Abstract. The influence of removing the apical shoot and different leaves above and below the flower on the fruit-set of unpollinated pea ovaries (Pisum satirum L. cv. Alaska) has been studied. Unpollinated ovaries were induced to set and develop either by topping or by removing certain developing leaves of the shoot. Topping had a maximum effect when carried out before or on the day of anthesis, and up to four consecutive ovaries were induced to set in the same plant. The inhibition of fruit-set was due to the developing leaves and not to the apex. The third leaf above the first flower, which had a simultaneous development to the ovary, had the stronger inhibitory effect on parthenocarpic fruit-set. The application of different plant-growth regulators (indoleacetic acid, naphthylacetic acid, 2,4-dichlorophenoxyacetic acid, gibberellic acid, benzyladenine and abscisic acid) did not mimic the negative effect of the shoot. Keywords: Abscisic acid Auxin - Cytokinin Decapitation - Fruit-set -- Gibberellin - Parthenocarpy - Pisum.
Introduction Fruit-set has been defined as the changeover from the static condition of the flower ovary to the rapidly growing condition of the young fruit (Leopold and Scott, 1952). There is a general assumption that this change is initiated by hormones either furnished by * Present address: Plant Growth Laboratory, Wickson Halt, University of California, Davis, CA 95616, USA Abbreviations: CCC = (2-chloroethyl)trimethylammoniumchloride;
MH=maleic hydrazide; IAA=indole-3-acetic acid; N A A = l-naphthaleneacetic acid; 2,4-D=2,4-dichlorophenoxyacetic acid; GA3=gibberellic acid; 6-BAP=benzyladenine; ABA=abscisic acid
the pollen, or produced by the developing seeds, or present in the ovules or pericarp in parthenocarpic varieties (Nitsch, 1970). Parthenocarpic fruit development can be induced by the application of plantgrowth regulators (Crane, 1964) or under certain environmental conditions (Gustafson, 1942). Vegetative development competes with fruit development and fruit-set can be increased by removing vegetative parts. The developing seeds seem to attract nutrients to the ovary and produce a sink which is able to compete with vegetative sinks (Bollard, 1970). Eeuwens and Schwabe (1975) showed that a close relationship exists between changes in extractable hormones and growth rates of pea pod walls and seeds. They used plants which had been topped immediately above the second flowering node. When we used the same model of plant in experiments with emasculated flowers we observed that the ovaries always set irrespective of the kind of treatment applied to the flowers. The aim of the present work was to study the influence of the vegetative parts on the development of the unpollinated ovary. We have shown that parthenocarpic fruit development may be induced by topping pea plants and that the inhibitory effect of the shoot on fruit-set can not be substituted by different plant-growth regulators.
Materials and Methods Plant Material
Seeds of Pisum sativum L. cv. Alaska were germinated and grown for 7 days in "coarse" sand (consisting of 60% (w/w) of 1-2 mm and 26% (w/w) of 0.5-1 mm diameter particles) at or below 25 ~ C under sunlight conditions and irrigated with deionized water. Seedlings were selected for uniformity and transferred into 11.5 cm diameter pots containing "fine" sand (consisting of 58% (w/w) of 0.5-1 mm and 30% (w/w) of 0.25-0.5 mm diameter particles). Both types of sand were usually treated with 1 M HCI for a week
0032-0935/80/0147/0444/$01.40
J. Carbonell and J.L. Garcia-Martinez: Fruit-set of Unpollinated Ovaries
LEAVES ~ FLOWERS 2
a
1
2
~
1
-1
-2
445
Table 1. Values of i and j Position of leaves removed
j value
i value
None 1 2 3 12 13 23 123 123etc.
1 2 3 4 5 6 7 8 9
0 1 0 2 0 1 0 1 034 024 014 04 0
2 3 3 2
3 4 4 4 4
'-3 , ~
9 Fig. 1. Numeration of leaves and flowers in a pea plant
and then washed repeatedly with deionized water; the "'fine" sand was then equilibrated with nutritive solution for a week by daily changing the solution. The nutritive solution was Hoagland n ~ 1, containing the oligoelements of Hewitt (1966), and was automatically supplied to the plants each hour. Plants were grown through the year in a heated glasshouse, except in January, July, August, and December. Natural light was supplemented with Sylvania "daylight" fluorescent tubes (about 2,000 lx at the level of the first flowers) for a 16 h photoperiod. The temperature fluctuated according to the environment, but extremes were never higher than 30 ~ C or lower than 12~ C. The first flowers opened between 28 and 32 days after sowing. Differences in the development of the plants were observed, the more vigorous being obtained between the begining of March and the end of May. The fruit length and percentage of fruit-set appeared to depend to a small degree on light intensity and temperature, so the same experiment was repeated in different seasons. The results given in this paper come usually from the Spring experiments. Fruits shorter than 25 mm 12 days after anthesis were occasionally obtained but they were not taken into account because we had observed previously that they degenerated within a few9 days. Figure 1 gives a scheme of the positions of leaves and flowers in the plants. Flowers were numbered according to the sequence in which they appeared. Leaves were numbered by taking the leaf placed at the node of the first flower as a reference (leaf 0) ; upper leaves were numbered positively and lower leaves negatively. Except in the experiments corresponding to Figs. 3 and 4, we always used only the first flower of each plant. Other flowers were removed as soon as possible. Unpollinated flowers were obtained by removing petals and stamens two days before anthesis. At this stage the petals had approximately the same length as the sepals. Topping was carried out by cutting the internode indicated in each experiment. Axillary shoots and new flowers were always removed as they appeared to eliminate competition problems.
0.1% Tween 80: AMO- 1618 (Calbiochem), (2-chloroethyl)trimethylammonium chloride (CCC) (Cyanamid), and maleic hydrazide (MH) (MH-30, Naugatuk Chemical). Indoleacetic acid (IAA) (Merck), naphthaleneacetic acid (NAA) (Fluka), 2,4-dichlorophenoxyaeetic acid (2,4-D) (Fluka), gibberellic acid (GA3) (Fluka), benzyladenine (6-BAP) (Fluka) and abscisic acid (ABA) (Sigma) were dissolved in a small amount of methanol or ethyl acetate and dispersed in melted anhydrous lanolin at 60 ~ C. The lanolin paste was applied with a glass rod to the cut surface of the topped stem.
Statistical and Mathematical Methods Statistical treatments of the data were made by using the analysis of variance and the Tukey test for comparison of mean values after calculating the least significant difference (LS.D.). In order to determine the effect of each leaf above the first flower on fruit-set, it was supposed that these leaves could have a positive or activator effect (A~) and a negative or inhibitor effect (It) where i indicates the leaf position and takes values from 0 to 4 (values 0 and 4 correspond to all the leaves under leaf i and above leaf 3, respectively). By assuming that these effects are additive, we obtained an equation system for each day of the experiment shown in Fig. 7. The algebraic sum of activator and inhibitor effects of each non-removed leaf was set equal to the number of non-degenerated ovaries (Ni) that day; j represents each of the 9 possibilities obtained when no leaf was removed, or one, or several leaves above leaf n ~ 0 (Fig. 7). The general equation for each day is:
(A,-Ie)+SVj=5~ i=y(j)
A,>=O j = l . . . 9 I i =>0 i =f(j) = 0... 4
Nj_>0 svj~0 where SVj is a slack variable which represents all kinds of errors present in the experiment and i is a function of j, according to Table 1. Solutions for each equation system were obtained by minimizing the associated lineal function formed by the sum of the absolute values of the slack variables, that is, the sum of errors. The lineal programs formed were solved by a computer.
Results Application o f Plant Growth Regulators Plant-growth retardants and inhibitors were applied by dipping the top part of the plant, including leaf n ~ 1 in the solution. The following compounds were used in aqueous solution containing
Topping of plants above the first flower between days -2 and 8 after anthesis induced parthenocarpic f r u i t - s e t o f u n p o l l i n a t e d p e a o v a r i e s ( F i g . 2, i n s e r t ) .
446
J. Carbonell and J.L. Garcia-Martinez: Fruit-set of Unpollinated Ovaries
~Ioo~ 80-
.-_
,0-1 I
-2
70-
~
I x
I
topping
1
)4 6 8
0 2 4 Day of topping
80- ~
day of
~......~
e~
/
~
70-
~
_ ~o.,~-I , - e~'-'~ >'0
~60E 50r
./
60.
o
50"
3 z.
u. 30-
20.
/ # ///o/
/ / / / ' / / / . oo z//I/// e
I I
!:!:!:!:!:!:!::
~;i~iiiiiiiiil
~o__O_oO:%0 i
-
.............
