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This is to certify that the

thesis entitled

THE EFFECTS OF PLANT AGE, PHOTOPERIOD, VERNALIZATION, AND TEMPE’\TURE ON THE GROWTH AND DEVELOPMENT OF AQUTLEGIA AND GAURA.

presented by

Leslie Marie Finical

has been accepted towards fulfillment of the requirements for

M. S. degree in Horticulture

Major professor

Date December 9 , 1998

0.7 639 MS U is an Affirmative Action/Equal Opportunity Institution _— 'Q—C- _.___...‘ Av W A v - W”.

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1!” WWW“ THE EFFECTS OF PLANT AGE, PHOTOPERIOD, VERNALIZATION, AND TEMPERATURE ON THE GROWTH AND DEVELOPMENT OF AQUILEGIA AND GAURA.

By

Leslie Marie Finical

A THESIS

Submitted to Michigan State University in partial fulfillment of the requirements for the degree of

MASTER OF SCIENCE

Department of Horticulture

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Of 0‘ THE EFFECTS OF PLANT AGE, PHOTOPERIOD, VERNALIZATION, AND TEMPERATURE ON THE GROWTH AND DEVELOPMENT OF AQUILEGIA AND GAURA ABSTRACT

By

Leslie M. Finical

Studies were conducted to determine the effects of plant age, vemalization, temperature, and photoperiod on the growth and development of a select group of Aquilegia and one cultivar of Gaura. Aquilegia were cold treated as either plants established in 5" containers, or small seedlings. Gaura were cold treated only as small cuttings. Plants were cooled at 5 °C for 0, 3, 6, 9, 12, and 15 weeks then forced in a 20 °C greenhouse. The duration of cold treatment required for flowering varied for the remaining species and cultivars.

To determine effects of forcing temperature after cold treatment on flowering.

After a 12-week cold treatment, ten plants of each species and cultivar were grown at 14, 17, 20, 23, and 26 °C. Time-to-flower was faster at higher temperatures but plant and flower size was reduced. For plants of Aquilegia, leaf unfolding rates were measured at 17, 20, 23 and 26 °C, 2) plant age and number of leaves required before cold treatment for flowering, and 3) the effects of short and long photoperiods before and after cold treatment. Leaf unfolding rates were similar at all temperatures for each cultivar. Age, leaf number, temperature and photoperiod affected subsequent flowering. Plants of Gaura were also studied to determine response to photoperiod. After 0 and 15 weeks of cold, plants were grown under 10, 12, 13, 14, 16, 24 and NI lighting. CTFVK TABLE OF CONTENTS

LIST OF TABLES ...... v

LIST OF FIGURES ...... ix

CHAPTER I LITERATURE REVIEW ...... 1 Introduction ...... 2 History of Vemalization ...... 3 Vemalization Versus Breaking of Dormancy ...... 4 Requirements of the Vemalization Process ...... 5 Juvenility ...... 6 Vemalization and Photoperiod ...... 7 De-vemalization ...... 9 The Perennial Plant ...... 10 Site of Vemalization Perception ...... 11 Mechanisms and Genetics ...... 12 Conclusions ...... 15 Literature Cited ...... 17

CHAPTER II A COMPARATIVE STUDY OF THE EFFECTS OF PLANT SIZE AND COLD TREATMENT ON THE GROWTH AND FLOWERING OF AQUILEGIA FLABELLA TA ‘CAMEO’, AQUILEGIA FLABELLA TA ‘MINI STAR’, AQUILEGIA XHYBRIDA ‘BLUEBIRD’, ‘CRIMSON STAR’, ‘MCKANA’S GIANT’, AND ‘MUSIK- WHITE’...... 20 Introduction ...... 21 Materials and Methods ...... 26 Results and Discussion ...... 29 Conclusions ...... 36 Literature Cited ...... 43

CHAPTER III THE EFFECTS OF FORCING TEMPERATURE ON THE GROWTH AND FLOWERING OF AQUILEGIA CANADENSIS, AQUILEGIA FLABELLA TA ‘CAMEO’ AND ‘MINI STAR’, AND AQUILEGIA XHYBRIDA ‘BLUEBIRD’ . . . 57 Introduction ...... 53 Materials and Methods ...... 61 Results and Discussion ...... 64 Conclusions ...... 69 Literature Cited ...... 72

CHAPTER IV. THE EFFECTS OF TEMPERATURE, PLANT AGE, COLD TREATMENT, AND PHOTOPERIOD ON THE GROWTH AND FLOWERING OF AQUILEGIA FLABELLA TA ‘CAMEO’, AQUILEGIA XHYBRIDA ‘BLUEJAY’, ‘CRIMSON STAR’, ‘MCKANA’S GIANT’ AND ‘MUSlK-WHITE’ ...... 87 Introduction ...... 88 Materials and Methods ...... 91 Results and Discussion ...... 95 Conclusions ...... 102 Literature Cited ...... 105

CHAPTER V. THE EFFECTS OF PHOTOPERIOD. VERNALIZATION, AND FORCING TEMPERATURE ON THE GROWTH AND FLOWERING OF GAURA LINDHEIMERI ‘WHIRLING BUTTERFLIES’ ...... 144 Introduction ...... 145 Materials and Methods ...... 149 Results and Discussion ...... 152 Conclusions ...... 156 Literature Cited ...... 160

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Tab fiVe 12 It Table 7. The average crown diameter measured in millimeters for each species before cold treatment. The diameter of each seedling was measured using calipers before they were moved to the cooler. There were no significant trends in flowering associated crown diameter before cold for any of the species or cultivars studied...... 51

Table 8. The recommended cold treatment durations for all flowering species tested...... 52

CHAPTER III

Table 1. Ages, sowing dates, and transfer dates for the Aquilegia species used in the experiment. All of the seedlings were sown and grown commercially in 72-cell plug trays (each cell equals 50 ml volume). The seedlings were shipped to MSU on September 16, 1997. All seedlings were grown under a 16-h photoperiod at 20 °C until they reached the age of 16 weeks before they were transferred to a 5 °C cooler. The seedlings that were older than 16 weeks upon arrival were transferred to a 5 °C cooler the day after they were received...... 74

Table 2. Dates of forcing and the average temperature during forcing. . . . . 75

Table 3. Regrowth and flowering responses of Aquilegia flabellata ‘Cameo Blue and White’ at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod at temperatures of 14, 17,20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower...... 76

Table 4. Regrowth and flowering responses of Aquilegia flabellata ‘Mini star’ at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod at temperatures of 14, 17, 20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower...... 77

Table 5. Regrowth and flowering responses of Aquilegia canadensis at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod at temperatures of 14, 17, 20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower...... 78

Table 6. Regrowth and flowering responses of Aquilegia xhybn’da “Bluebird’ at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod

vi at temperatures of 14, 17, 20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower...... 79

Table 7. Parameters of linear regression analysis relating forcing temperature to rate of progress to visible bud (VB), to first open flower (FLW), and from VB to FLW in A. flabellata ‘Carneo’ and ‘Mini star', A. canadensis, and A. xhybn’da ‘Bluebird’. Intercept and slope were used to calculate base temperature (T t,) and degree-days (°days)...... 80

CHAPTER IV

Table 1. All seedlings were four weeks of age when they were received at MSU on October 13, 1997. 128 Seedlings of each cultivar were placed at 17, 20, 23, and 26 °C under 16-h photoperiods with HPS lamps. Half of the seedlings from each cultivar were moved directly into a 5 °C cooler on the dates listed below and cooled in the plug tray. The treatments for the remaining seedlings are listed in Table 2...... 106

Table 2. Average greenhouse temperatures for all cultivars during forcing, after cold treatment...... 106

Table 3. Leaf unfolding rates/day at each growing temperature before cold treatment ...... 108

Table 4. Plants of Aquilegia xhybn'da ‘Bluejay‘ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower...... 109

Table 5. Plants of Aquilegia flabellata ‘Cameo’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower...... 1 12

Table 6. Plants of Aquilegia xhybrida ‘Crimson Star’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower...... 1 15

Table 7. Plants of Aquilegia xhybn'da ‘McKana’s Giant’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial

vii

USl leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower...... 1 18

Table 8. Plants of Aquilegia xhybrida ’Musik White’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower ...... 121

Table 9. The overall average leaf number of both the short-day and NI treatments combined at each growing temperature before cold of each species studied. Also, the overall average leaf number of pot- and plug-cooled plants combined for each growing temperature...... 123

Table 10. The overall combined average leaf numbers at the ages each species studied was placed into the cooler...... 124

Table 11. The requirements for flowering of five cultivars of Aquilegia. The following recommendations apply to plants grown in 1.1-L or larger containers prior to cold treatment to achieve 100% flowering. Cold duration recommendations are taken from chapter 2. Force plants at temperatures between 14 and 20 °C (chapter 3)...... 125

CHAPTER V

Table 1. The effects of photoperiod and cold treatment on flowering cooled plugs of Gaura Iindheimen’ ‘Whirling Butterflies”. Plants were cooled for 0 or 15 weeks in a controlled environment chamber set at 5°C. After cold treatment, plants were grown in a greenhouse set at 20 °C ...... 162

Table 2. The effects of cold treatment on flowering cooled plugs of Gaura Iindheimen' ’Whirling Butterflies’. Plugs were cooled for 0, 3, 6, 9, 12, and 15 weeks in a controlled environment chamber set at 5 °C. After cold treatment, plants were forced in a greenhouse set at 20°C under 16 hours of light from HPS lamps...... 163

Table 3. The effects of forcing temperature on flowering of cooled plugs of Gaura Iindheimen' ‘Whirling Butterflies’. Plugs were cooled for 12 weeks in a controlled environment chamber set at 5 °C...... 164

Table 4. Parameters of linear regression analysis relating forcing temperature to rate of progress to visible bud (VB), to first open flower (FLW), and from VB to FLW in Gaura Iindheimen' ‘Whirling Butterflies‘. Intercept and slope were used to calculate base temperature (Tb) and degree-days (°days)...... 164

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CHAPTER II Page

Figure 1. Effect of cold duration at 5 °C on percent flowering, days to visible bud, and days to flower on plants of Aquilegia flabellata ‘Cameo Blue and White’, A. flabellata ‘Mini star’, A. xhybrida ‘Bluebird’, A. canadensis, and A. chrysantha. Plants were cooled either in 1.1-L (5" US.) square pots (O) or in 72-cell plugs (A)...... 54

Figure 2. Effect of cold duration at 5 °C on total plant height, final leaf number, and flower number on plants of Aquilegia flabellata ‘Cameo Blue and White’, A. flabellata ‘Mini star’ , A. xhybrida ’Bluebird’, A. canadensis, and A. chrysantha. Plants were cooled either in 1.1-L (5" US.) square pots (O) or in 72-cell plugs (A)...... 56

CHAPTER III

Figure 1. The effects of forcing temperature on the flowering percentage of A. canadensis, ‘Cameo’, ‘Mini star’, and ‘BIuebird’. Plants that did not flower within 100 days were considered non-flowering...... 81

Figure 2. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates of progress to each event for A. flabellata 'Cameo‘. The points represent the actual data points and the curves and straight lines were calculated from regression analysis. . 82

Figure 3. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates of progress to each event for A. flabellata 'Mini star“. The points represent the actual data points and the curves and straight lines were calculated from regression analysis. . 83

Figure 4. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates of progress to each event for A. canadensis, The points represent the actual data points and the curves and straight lines were calculated from regression analysis. . 84

Figure 5. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates of progress to each event for A. xhybrida ‘Bluebird‘. The points represent the actual data points and the curves and straight lines were calculated from regression analysis. . 85 Figure 6. Influence of forcing temperature on number of flower buds, flower diameter, and plant height measured at first flower. Error bars show standard deviation...... 86

CHAPTER IV

Figure 1. The effects of temperature on leaf development of Aquilegia xhybrida ‘Bluejay’, A. flabellata ‘Cameo’, A. xhybrida ‘Crimson Star’, A. xhybrida ‘McKana’s Giant’, and A. xhybrida ’Musik—White’. The points indicate the actual number of leaves unfolded at each age. The straight line was calculated from linear regression analysis...... 126

Figure 2. The effects of initial leaf number, temperature before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs", on plants of ‘Bluejay’, ‘Cameo’, ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik-White’. Seedlings were grown at 17, 20, 23, and 26 °C before cold. (O) represents plants grown under SD, ([3) represents plants grown under NI...... 128

Figure 3. The effects of initial leaf number before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on flowering percentage of ‘Bluejay’, ‘Cameo’, ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik-White’. (O) represents plants grown under SD, (a) represents plants grown under Nl...... 131

Figure 4. The effects of temperature before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on days to visible bud for plants of ‘Bluejay’, ‘Cameo’, ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik— White‘. Seedlings were grown at 17, 20, 23, and 26 °C before cold. (0) represents plants grown under SD, (1:) represents plants grown under NI. . 133

Figure 5. The effects of temperature before cold treatment, and overall age of pot- and plug-cooled plants, on the days to visible bud for ‘Bluejay’, ‘Cameo’, ‘Crimson Star”, ‘McKana’s Giant’, and ‘Musik—White’. (a) = 17 °C, (0) = 20 °C, (X) = 23 °C, and (E1) = 26 °C...... 136

Figure 6. The effects of average leaf number before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on days to visible bud, days from visible bud to flower, and days to flower for plants of ‘Bluejay’, ‘Cameo’, ‘Crimson Star’, ‘McKana’s Giant‘, and ’Musik-White’. (0) represents days to visible bud, (a) represents days to flower...... 138

Figure 7. The effects of plant age in weeks on the flower number of “Pots” and “Plugs” of ‘Bluejay’, ’Cameo’, ‘Crimson Star”, ’McKana’s Giant’, and ‘Musik- White’. Data points represent total average of plants grown under both SD and NI lighting at each age...... 140 Figure 8. The effects of temperature before cold treatment, and photoperiod before cold for “Pots”, and photoperiod after cold for "Plugs" on plant height of ‘Bluejay', ’Cameo’, ‘Crimson Star', ‘McKana’s Giant’, and ‘Musik—White’. (0) represents plants grown under SD, (1:) represents plants grown under Nl. . 142

CHAPTER V

Figure 1. The effects of cold treatment and photoperiod on the flowering of Gaura Iindheimeri ‘Whirling Butterflies’. Percent flowering, days to visible bud, and days to flower for (A) year 1, 0 weeks of cold, (B) year 1, 15 weeks of cold, (C) year 2, 0 weeks of cold, and (D) year 2, 15 weeks of cold...... 165

Figure 2. The effects of cold treatment and photoperiod on node development of Gaura Iindheimeri ‘Whirling Butterflies’. Number of nodes at first flower for (A) year 1, 0 weeks of cold, (B) year 1, 15 weeks of cold, (C) year 2, 0 weeks of cold, and (D) year 2, 15 weeks of cold...... 166

Figure 3. The effects of cold treatment and photoperiod on the flower bud number and plant height of Gaura Iindheimeri ’Whirling Butterflies’. Flower bud number for (A) year 1 and (B) year 2. Total plant height for (C) year 1 and (D) year 2 ...... 167

Figure 4. The effects of different durations of cold treatment on the flowering, plant node development, flower number, and plant height of Gaura Iindheimeri ’Whirling Butterflies’. Flowering percentage, days to visible bud, and days to flower for (A) year 1 and (B) year 2. Number of nodes at first flower for (C) year 1 and (D) year 2. Number of flower buds at first flower for (E) year 1 and (F) year 2. Plant height at first flower for (G) year 1 and (H) year 2...... 168

Figure 5. Influence of forcing temperature on time and rate toward flowering. Lines represent predicted values from the regression equations. Base temperature (Tb) and cumulative thermal time (CTT) necessary to complete the indicated developmental stage were calculated from linear regression analysis...... 169

Figure 6. Influence of forcing temperature on number of flower buds, flower diameter, and plant height of Gaura Iindheimeri ’Whirling Butterflies'. Each measurement was taken at first flower. The points indicate the actual data points and the straight lines were calculated from linear regression analysis. Error bars show standard deviation...... 170

xi CHAPTER I. LITERATURE REVIEW Introduction

Environmental signals such as photoperiod and temperature can strongly influence flowering patterns and induction of flowers or inflorescences in plants.

Plant response to these stimuli depends upon the species and genotype of the plant. For some plants, particularly those growing in northern latitudes, a period of exposure to low temperatures below 7 °C, which is referred to as vemalization is required to induce flowering (Lang 1965).

Vemalization has been defined as a cold treatmentthat promotes subsequent flowering when it is given to imbibed seeds, bulbs, or whole plants

(Vince-Prue, 1975). The word vemalization, named by Russian scientist

Lysenko, originates from the Latin word “vemum” meaning spring (Chouard,

1960). Interest in the concept of vemalization originated from early agricultural experience with winter and spring cereals. Winter cereals needed to be planted before the end of winter in order to fruit within 12 months of sowing, whereas spring cereals flowered soon after spring sowing (Chouard, 1960). Early research concentrated on the chilling of germinated seeds of cereal grains.

Treating seeds with thermoinductive temperatures was practiced frequently in

Russia during the early 19003 to quicken the development of grain crops.

Modern day uses of vemalization treatments include forcing of herbaceous perennials for use in the garden along with potted flowering plant production such as Easter lilies. History of Vemalization

Many early vemalization experiments dealt with chilling germinating seeds of cereal crops such as spring and winter wheat. Much of the early work in the area of seed vemalization is credited to Klebs and Gassner in the early 1900‘s.

One of the earliest references of the chilling of seed is that of Klippart in 1857 and even earlier records go back to the 1830s. Klippart experimented with spring and winter wheat and stated that "To convert winter into spring wheat, nothing more is necessary than that the winter wheat should be allowed to germinate slightly in the fall or winter, but kept from vegetation by a low temperature or freezing, until it can be sown in the spring...”. Klippart explained his procedure for soaking and freezing the seeds, and presented the results of his experiment. Lysenko, however, is credited for reviving the interest in temperature and for defining the technique known as vemalization. The basic

principles of seed vemalization have not changed since these early experiments, although new methods have been developed for the sterilization and handling of

many different types of seeds.

The vemalization requirement for wheat and other cereal grains may be fulfilled while the kernels are still developing in the ear, although, in the early

stages of embryo development, the vemalization response seems to be non-

existent or very low. Hoogendoom (1984) found vemalization of kernels while

still on the ear advanced flower initiation faster than vemalization of kernels with

a fully developed endosperm. (Hoogendoom, 1884, cited in Krekule, 1987). For

example, wheat plants with developing ears were treated with thermoinductive

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Vemalization Versus Breaking of Dormancy

Although the terms “vemalization” and “cold treatment” are frequently used interchangeably, two different phenomenons may be taking place.

Vemalization is a cold treatment, but a cold treatment isn’t necessarily vemalization. Vemalization, as defined above, is a cold treatment that promotes or induces subsequent flowering when given to imbibed seeds, bulbs, or whole

plants. The return to warmer temperatures after the requirement for cold is fulfilled is also a necessary step in the vemalization process.

Cold temperatures promote flowering in many plants by breaking dormancy of existing buds when reproductive structures are present prior to the onset of low temperatures. Many woody perennials and flowering bulbs such as

azaleas, hydrangeas, and tulips for example, form their flower buds in the

summer or fall. These plants then remain dormant throughout the winter. The

floral primordia of Paeonia are initiated soon after anthesis of the current year’s

flowers and require a minimum of four weeks at 5.6 °C to break dormancy

(Byme and Halevy, 1986). Full reproductive development, or anthesis, will not

occur without a subsequent cold treatment. This would be considered a

breaking of dormancy, not vemalization (Thomas and Vince-Prue, 1984).

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Dicentra spectabilis, flower differentiation occurs during the cold, or cool temperature treatment (Thomas, Vince-Prue, 1984). In Allium cepa (onion), and his cv. Wedgewood, low temperatures are also required during floral initiation. In

Matthiola incana, low temperatures must continue until the buds are visible in order for development to continue upon return to warmer temperatures (Kohl,

1957). For some plants, a cold treatment will speed the process of flowering but is not necessary to initiate flowering. This is termed a facultative, or quantitative response to cold. Exposure of some species to low temperatures will induce subsequent initiation and development of floral primordia. For plants that require a vemalization treatment, flower buds will not form until the requirement for cold is fulfilled and the plants are returned to warmer temperatures.

Requirements of the Vemalization Process

For plants that absolutely require a vemalization period, no flowering will occur until the qualitative or obligate requirement for a cold treatment as been fulfilled (Vince-Prue, 1975). Cold treatments may be administered to seeds, seedlings, or older plants, depending on the species. When treating seeds, there must be sufficient imbibition of water to allow vemalization to proceed.

Lang (1965) claimed that the most effective temperatures for vemalization range from 1 to 7 °C. Duration of the cold treatment in conjunction with temperature is also a factor in the achievement of vemalization. Some species will achieve a stable vemalized state with only two or three weeks of cold treatment (Jedel et al, 1986). The vemalization requirement for some cultivars such as spring wheat is four weeks (Jedel et al, 1986), whereas the requirement for some cultivars of Astilbe is 10 or more weeks (Beattie, 1983).

For plants with a vemalization requirement, the lack of an adequate period of cold results in delayed, or complete absence of flowering (Thomas and Vince-

Prue, 1984).

Response to vemalization treatments also varies between different cultivars of the same species. This may be due to the differentiation of cultivars as ecotypes which have evolved under different climatic conditions. Thus, some cultivars may require lower or higher temperatures to satisfy their vemalization requirement. In a study performed by Doi, Takeda and Asahira (1984), four cultivars of Gypsophila paniculata L. were examined for their response to thermoinductive temperatures. ‘Perfecta’ was sufficiently vemalized at 10°C or below, ’Diamond’,‘FIamingo’ and ‘Red Sea’ all required temperatures below 5

°C.

Juvenility

For many herbaceous and woody perennial plants, a certain size or age must be reached before any treatment with thermoinductive temperatures is effective. The period of time during which plants are not responsive to external environmental stimuli is called the juvenile phase. Juvenility may be defined as the ear1y phase of growth during which flowering cannot be induced by any treatment (Thomas and Vince-Prue, 1984). Thus, cold treatments will not induce floweflnq draflor

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flowering until the plant has made the transition to its mature phase. The duration of the juvenile phase varies from species to species. The juvenile phase may be quite short in some herbaceous perennials, lasting only days or weeks (Thomas and Vince-Prue, 1984), whereas for woody plants, the juvenile phase may last years. Apples, for instance, remain juvenile for five to seven years before becoming reproductive.

Some plants, such as Hedera helix L., have several distinct differences

between juvenile and mature phases;

Juvenile characters Adult characters

Three or five lobed palmate leaves Entire, ovate leaves

Alternate phyllotaxy Spiral phyllotaxy

Anthocyanin pigmentation of you No Anthocyanin leaves and stems

Stems pubescent Stem glabrous

Climbing and plagiotropic growth Orthotropic growth habit habit Shoots show unlimited growth and Shoots show limited growth lack terminal buds terminated by buds with scales

Absence of flowering Presence of flowers

(Thomas and Vince-Prue, 1984-adapted from Wareing and Frydman,1976). In winter rye, a vemalization treatment is only effective after approximately 22

leaves have developed (Purvis,1961).

Vemalization and Photoperiod Some plants respond more strongly to a vemalization treatment if a specific photoperiod is provided in conjunction with the cold (Thomas and Vince-

Prue, 1997). For example, some plants prefer long days or 24-hour lighting such as Coreopsis verticillata, whereas some prefer short-days, such as

Chrysanthemum, before a cold treatment (Napp-Zinn, 1984).

The requirement for vemalization is commonly, but not only, found in long day plants (Thomas and Vince-Prue, 1997). Many plants that have a cold requirement also require subsequent treatment with long days, such as

Coreopsis ‘Sunray’. In some cold-requiring plants, such as Campanula medium, short days may substitute for, enhance, or be required instead of cool temperatures in order to achieve anthesis (Roberts et al.,1988). Flower bud initiation for young plants of Campanula medium to was one month faster when plants were exposed to short-days than when exposed to cool temperatures

(Wellensiek, 1960). This phenomenon is sometimes referred to as short-day vemalization. The low temperature requirement for flowering in Japanese winter wheat varieties Norinn 27 and 8 is completely replaced by short-day treatments

(Krekule, 1987). Providing certain photoperiods in conjunction with vemalization can also influence the effectiveness of the cold treatment. For some species, providing short-days plus night-interruption lighting, long days, or 24- hour lighting during the cold treatment, can increase the effectiveness of the vemalization (Napp-Zinn,1984).

Conversely, In Poa pratensis and celery plants, long—days during vemalization inhibited flower induction (Vince-Prue,1975:Thomas and Vince-Prue, 1984).

8 The process by which short-days induce flowering seems to be different than that of cold induction. Short days seem to be additive to sub-optimal vemalization but are usually inhibitory to fully vemalized material. An experiment that displays this inhibitory phenomenon was conducted on Triticum aestivum L.

(Krekule, 1964). Sprouted seeds of three varieties of wheat were chilled at 2-3 °

C for 10, 20, or 30 days. After treatment, the seedlings were placed under long days (14.5-16 hr) or short days (8 hr). In all three varieties, the days to ear emergence (30 days under short day conditions) increased and dry weight decreased with increased number of short days. In two of the varieties, growth remained prostrate for the plants treated with short days. Days to ear emergence for the San Pastore variety increased from 95 days under long day conditions to 115 days under short day conditions (both treatments received 20 days of chilling) (Krekule, 1987).

De- Vemalization

De-vemalization is the reversal, by environmental conditions, of the vemalized state of a plant. The primary cause of de-vemalization seems to be high temperatures, at or above 30 °C, but short days may also substitute for high temperatures (Wellensiek, 1965). Conditions such as high temperatures and short days are most effective in causing de-vemalization when provided immediately after vemalization has occurred. Once a plant has received its’ cold requirement, the vemalized condition is usually stable. Hyoscyamus plants which had been vemalized and subsequently kept under short days for more tf than 190 days flowered readily once placed back into long days (Lang,1986).

However, if the plant has received a less than Optimal period of cold treatment, subsequent high temperatures or short days may revert the plant to a non- vemalized state (Thomas and Vince-Prue, 1984). The stabilization of wild lettuce Lactuca sem'ola is dependent upon or associated with growth during normal temperatures and light during the chilling period. The same can be said for Arabidopsis (Napp-Zinn,1960).

The Perennial Plant

If the vemalized condition can be transferred to all parts of the plant including buds, how does a perennial plant ensure its’ perennial habit? Would not a vemalized bud go on to produce flowers and the plant remain monocarpic?

This would seem logical, but perennial plants have developed mechanisms to ensure that their seeds and subsequent seedlings must be exposed to a vemalization period before flowering (Thomas and Vince-Prue,1984). In

Chrysanthemum mon'folium, some of the vemalized buds become de-vernalized during the summer. The shoots which arise from the base of the plant at the end of summer are also non-vemalized (Thomas and Vince-Prue, 1984). In some perennial grasses, the vemalized condition is not perpetuated through all cell divisions. This enables new tillers which are produced in the summer to be non- vemalized (Purvis, 1961).

10 Site c

(Lang

leave:

1937)

therm

treatrr

when

expen

plants

tempe

leaves

"DOtSl

roots \

Vemal

bUtsc

QTOWir

aCtIOn

EXpefir ’yplcal l Site of vemalization perception

There are two established theories for the perception of vemalization

(Lang, 1965; Purvis, 1966): the original theory was that vemalization affects leaves and results in a hormonal balance in favor of flowering (Melchers 1936,

1937). The second, more widely accepted theory, states that cold therrnoinduces apical cells directly (Schwabe, 1954). Localized cooling treatments applied to certain biennial and perennial plants caused flowering when only the stern apex was chilled independent of the temperature experienced by the rest of the plant (Thomas and Vince-Prue, 1984). For other plants, it is apparent that the shoot apex is not the only site where cold temperatures are perceived. Vemalization perception in both the apex and leaves was observed in Pisum sativum L. by Reid and Murfet (1975).

Experiments on potential perception of thermoinductive temperatures by roots of Chinese cabbage were conducted by Reitze and Wiebe (1988). The roots were warmed in order to observe any inhibition or retardation of the vemalization process. Warm roots had no influence on the vemalization effect but soil heating decreased the effect of vemalization by heat transfer into the growing point (Riete and Wiebe, 1988). In this experiment, roots were not shown to be a primary site of cold perception.

Wellensiek (1964) proposed that dividing cells are necessary for the action of low temperatures, no matter where in the plant they occur.

Experiments were conducted by Wellensiek on Lunaria biennis L., which is a typical cold-requiring plant. The procedure involved taking leaf and root cuttings

11 from the plant and placing them under a range of cold treatments. On some of the leaf cuttings, the lower 0.5 centimeter of the petiole was removed from the cutting. This area of the petiole contains meristematic tissue, thus dividing cells.

Flowering was not observed in the regenerated shoots on leaf cuttings without petioles. Flowering was observed in the regenerated shoots of the leaf cuttings with intact petioles. Success also seemed to rely on the age of leaves used as cuttings. The vemalization treatments were more effective for younger leaf cuttings than for fully expanded, more mature leaves (Wellensiek, 1963).

The apex, or shoot tip was found to be the site of perception of thermoinductive temperatures in the species Thlaspi arvense L. (Metzger, 1988).

However, when untreated leaf cuttings from mature induced mother plants were examined, the leaf cuttings generated flowering shoots. The lower 1 to 2 centimeters of the petiole had been removed in order to remove any meristematic tissue from the leaf. The result of the experiment may suggest that different types of cells other than dividing cells may be capable of perceiving thermoinductive temperatures. The possibility remains, however, that there may have been meristematic tissue elsewhere in the leaf that was not removed before treatment, or that there are basic differences between species.

Mechanisms and Genetics

The term ‘vemalization’ can have at least two different meanings: on the one hand it is a cold treatment that induces or promotes flowering, and on the

other it is the specific biochemical processes that occur during cold temperatures

12 all< flc

allowing the development of the so-called “internal conditions” that lead to

flowering (Napp—Zinn, 1987). These internal conditions are controlled by both genetic and environmental factors.

Evidence suggests that some substance is formed by plants during exposure to thermoinductive temperatures. This substance can apparently be transmitted within the plant. Vemalized seeds and shoot tip may transport this substance to cells which form after the inductive period has been completed.

One theory suggests that the hypothetical substance, ’vemalin’, may be a gibberellin (Thomas and Vince-Prue, 1997). There are arguments for and against this theory. In Arabidopsis, practically all vemalization-requiring genotypes flower faster when treated with any gibberellin from GA1 to GA9

(Napp-Zinn, 1963). On the other hand, gibberellin treatment of winter wheat and other grasses does not replace their chilling requirement (Napp-Zinn, 1987).

It has been suggested that many factors such as temperature and light can enhance or delay vemalization in plants by effecting gene expression or repression. For instance, photoperiod alters the expression of the FRIGIDA gene, which is contained in certain phenotypes of Arabidopsis plants (Lee and

Amasino, 1995). Arabidopsis is normally considered a quantitative long-day plant. The presence of the FRIGIDA gene causes Arabidopsis to behave as an

“obligate” LDP, since the primary meristem of most un-vemalized FRIGIDA- containing plants fail to produce a flowering stalk under short days (Lee and

Amasino, 1995).

13 (A

ll:

91

CE

5)

Ve ler

De Ver Vemalization requirements may be caused by dominant alleles, as in

Pisum, or by recessive alleles, as in Triticum and Arabidopsis (Stankov, 1972). It is possible that the dominant alleles cause the synthesis of a substance which is not formed by recessive alleles. Dominant alleles which provoke a vemalization requirement may initiate the synthesis of a flower inhibitor (Napp-Zinn, 1987).

This process may be reversed in plants where recessive alleles cause a vemalization requirement.

Several genes involved in vemalization have been identified. In both

Secale cereale (Purvis, 1939), and Hyoscyamus niger (Lang, 1986) there is only one gene that is responsible for determining the vemalization requirement. Only one gene separates spring and winter wheat while up to 24 genes are responsible for the vemalization requirement in Arabidopsis plants.

Low temperatures may alter the sensitivity Of the apex or shoot tip to

floral initiation. Low temperatures may also inhibit the expression of certain genes which may allow for the vemalization requirement to be fulfilled. In the case of pea plants, low temperature or light can deactivate a gene that codes for a flowering inhibitor, thus allowing the induction of flowering to occur. rRNA synthesis and concentration in the plumula of winter wheat (Triticum aestivum L.

) plants were found to be significantly different depending on the stage of vemalization (Paldi and Devay, 1982). Certain rRNA is synthesized at low temperatures but decomposes when temperatures reach 25 °C (Paldi and

Devay, 1983). The rRNA is necessary for transcription and transfer of the vemalized condition through meiotic divisions. The raising of temperatures

14 be ve' the res as: cul rec azz anl res me ray

flo'

Cc

ar

ih

r e of before rRNA has coded for the vemalized condition may then lead to incomplete vemalization or de-vemalization (Paldi and Devay, 1983). They proposed that the control of gene expression involves DNA methylation in vemalization responses. Methylation of cytosine in the promoter region of a gene may be associated with a lack of transcription of that gene. In Arabidopsis and cell cultures of Nicotiana plumbaginigolia, cold treatments resulted in a substantially reduced level of methylation. Treatment with the de-methylation agent 5- azacytidine hastened flowering in Thlaspi arvense and late-flowering ecotypes and mutants of Arabidopsis, but had no effect on flowering in mutants that do not respond to vemalization (Furn et al,, 1993). Gamma rays are a presumed de- methylating agent and, in winter wheat, exposure to 5-azacytidine or to gamma rays partially substituted for the cold treatment and significantly hastened

flowering (Brock and Davidson, 1994).

Conclusions

As it became clear that the vemalization responses observed in cereals may also be at work in other types of plants, the interest in treating seedlings and young plants with thermoinductive temperatures expanded to include other commercial crops. The seed experiments concentrated mainly on grain crops for the purpose of increasing crop yield and ensuring that a crop finished in a minimum amount of time. Experimentation with whole plants began after researchers felt that the area of seed vemalization had been fully explored. This opened the door to a much larger variety of plants such as potted and

15 omamen plants ha results ca

ornamental bedding plants. Over the last thirty to fifty years, many perennial plants have been studied for their responses to cold temperatures. These results can be used by commercial growers to produce plants in flower.

16 Bea‘

Bror

Byrr

Chc

Doi

Elli

Hr:

Je

K6

Kll

Kr La Literature Cited

Beattie, DJ. and E.J. Holcomb. 1983. Effects of chilling and photoperiod on forcing Astilbe. HortScience; 18: 449-450.

Brock, R. and J. Davidson. 1994. 5-Azacytidine and gamma rays partially substitute for cold treatment in vemalizing winter wheat. Env. Exper. Bot. 34:195-199.

Byme, T. and H. Halevy. 1986. Forcing herbaceous peonies. J. Amer. Soc. Hort. Sci. 11:379-383.

Chouard, P. 1960. Vemalization and its relation to dormancy. Ann. Rev. of Plant Physiol. 11: 191-238.

Doi, M. Y. Takeda, and T. Asahira. 1984. Differences in flowering response to low temperature among cultivars of Gypsophila paniculata L. and among vegetative lines of cv. Bristol Fairy. Mem. Coll. Agric., Kyoto Univ. 124: 27-34.

Ellis, R. H. and R. J. Summerfield.1988. Effects of temperature, photoperiod and seed vemalization on flowering in faba bean Vicia faba. Ann. of Bot. 61: 17-27.

Hoogendoom, J. 1984. A comparison of different vemalization techniques in wheat Triticum aestivum L.. J. of Plant Physiol. 116:11-20.

Jedel, P. E., L. E. Evans, and R. Scarth. 1986. Vemalization responses of a selected group of spring wheat (Triticum aestivum L.) cultivars. Can. J. of Plant Sci. 66:1-9.

Ketellapper, H. and A. Barbaro.1966. The role of photoperiod, vemalization and gibberellic acid in floral induction in Coreopsis grandiflora Nutt. Phyton 23: 33-41.

Klippart, J.H. 1857. Section 12, p. 562-816. Ann. Rept. Ohio State, Bd. Agr.

Krekule, J. 1987. Vemalization in wheat, p. 159-169. In; J. G. Atherton (ed.) Manipulation of Flowering. Butterworths, London.

Lang, A. 1965. Physiology of flower initiation, p. 1380-1536. In: J.G. Atherton (ed). Handbuch der Pflanzenphysiologie. Springer-Verlag, BerIin.

17 Lang.l

Lee,L

MBIChl

Meug

Mumh

Nappa

Nappé

Nappa

PadLl

Phnce

PUnfis,

Pun/lg, Reid, J. Lang, A. 1986. Hyoscyamus niger, p. 144-186. In: A. Halevy (ed.). CRC Handbook of Flowering. CRC Press, Boca Raton, FL.

Lee, I. and R. M. Amasino. 1995. Effect of vemalization, photoperiod, and light quality on the flowering phenotype of Arabidopsis plants containing the FRIGIDA gene. Plant Physiol.108:157-162.

Melchers, G. 1936. Biol. Zentr., 56: 567-570.

Metzger, JD. 1988. Localization of the site of perception of thermoinductive temperatures in Thlaspi arvense. Plant Physiol. 88:424-428.

Murphy, L. A. and R. Scarth 199. Vemalization response in spring Oilseed rape (Brassica napus L.) cultivars. Can. J. of Bot. Sci. : 275.

Napp-Zinn, K. 1963. Uber den Einfluss von Genen und Gibberellinnen auf die Blutenbildung von Arabidopsis thaliana. Berichte der deutschen botanischen Gesellschaft. 76: 77-89.

Napp-Zinn, K. 1964. Uber genetische und entwicklungsphysiologische Grundlagen jahreszeitslicher Aspekte von Pflanzengesellschaften, p. 33 - 49. In: K Kreeb (ed.). Beitrage zur Phytologie. E. Ulmer, Stuttgart.

Napp-Zinn, K..1987. Vemalization-environmental and genetic regulation, p. 123- 132. In: J. G. Atherton (ed.). Manipulation of flowering. Bodmin. Cornwall, UK.

Paldi, E. and M. Devay. 1983. The changes in the nature of rRNA synthesis in the plumula of seedlings during vemalisation. Plant Sci. Lett. 30:69-75

Prince, T. A. and. MS. Cunningham. 1990. Response of Easter Lily Bulbs to Sphagnum Peat Moisture Content and the use of Sphagnum Peat or of Polyethylene-lined Cases during Handling and Vemalization. J. Amer. Soc. Hort. Sci. 115.

Purvis, O. 1939. Studies in vemalization of cereals. V. The inheritance of the spring and winter habit in hybrids of Petkus rye. Ann. Bot. 32719-729.

Purvis, O. 1961. The physiological analysis of vemalization, p. 76-122. In: W. Ruhland (ed.). Encyclopedia of Plant Physiology volume 16. Springer- Verlag, Berlin.

Reid, J. B. and I. C. Murfet 1975. Flowering in Pisum: the sites and possible mechanisms of the vemalization response. J. of Exp. Bot. 26: 860-867.

18 Rietze, E ve

Roberts, i Ph of

Salisbury ge Gr

Schwabe

Talz, L, a Re

Teraoka,

Rietze, E. and H. J. Wiebe 1988. The influence of soil temperature on vemalization of Chinese cabbage. J. of Hort. Sci. 63: 83-86.

Roberts, E. H., R. J. Summerfield, R.H Ellis, and K.A. Stewart. 1988. Photothermal time for flowering in lentils (Lens culinaris) and the analysis of potential vemalization responses. Ann. of Bot. 61: 29-39.

Salisbury, P. A., G. J. Berry, and G. Halloran. 1979. Expression of vemalization genes in near-isogenic wheat lines: methods of vemalization. Can. J. Genet. Cytol. 21:429-434.

Schwabe, W.W. 1956. Factors controlling flowering in the Chrysanthemum. IV. The site of vemalization and translocation of the stimulus. J. Exp. Bot. 5: 389-400.

Stankov, I.K. 1972. Study of F1 hybrids between Tn'ticum sphaerococcum Perc. and the species T. durum Desf. And T. dicoccum Schrank. Genetika i Selektsiya. 25:199-210.

Taiz, L. and E. Zeiger. 1991. Plant physiology. Benjamin/Cummings Publishing, Redwood City, California.

Teraoka, H. 1967. Proteins of wheat embryos in the period of vemalization.Plant & Cell Physiol. 8: 87-95.

Thomas, B. and D. Vince-Prue.1984. Juvenility, photoperiodism and vemalization, p. 408-439. In: M. Wilkins (ed.). Advanced Plant Physiology. Pitman, London.

Vince-Prue, D. 1975. Photoperiodism in Plants. McGraw-Hill, Maidenhead.

Wang, S., R.W. Ward, J.T. Ritchie, R.A. Fischer, and U. Schulthess. 1995. Vemalization in wheat: I. A model based on the interchangeability of plant age and vemalization duration. Field Crops Res. 41 :91-100.

Wellensiek, SJ. 1964. Dividing cells as a pre-requisite for vemalization. Plant Physiol. 39: 832-5.

19 Ii -.

ELL THI FLA

CHAPTER II.

THE EFFECTS OF PLANT SIZE AND VERNALIZATION ON THE GROWTH

AND FLOWERING OF AQUILEGIA FLABELLA TA ‘CAMEO’, AQUILEGIA

FLABELLATA ‘MINI STAR’, AQUILEGIA XHYBRIDA ‘BLUEBIRD’, ‘CRIMSON

STAR’, ‘MCKANA’S GIANT’, AND ‘MUSIK—WHITE’.

20

35

DC

I

T1.

I‘i

«N.

.

,.

Ve

IS

fir:

Introduction

The genus Aquilegia is made up of about 70 species, commonly known as columbine, that are native to the temperate and mountain regions of the northern hemisphere and are (Bailey, 1976). Aquilegia are herbaceous perennials that bloom in spring and early summer and are best known for their dramatic spurred flowers. Aquilegia caemlea, A. longissima, A. chrysantha, A. canadensis, and A. vulgan's have been interbred resulting in numerous popular hybrids of many shapes, sizes and colors (Shedron and Weiler, 1982). The diversity of the genus provides commercial producers with many opportunities to adapt Aquilegia to pot-plant culture.

Vemalization has been defined as a cold treatment that promotes subsequent flowering when given to imbibed seeds, bulbs, or whole plants

(Vince-Prue, 1975). For plants that require a vemalization treatment, flower buds will not form until the cold requirement is fulfilled and the plants are returned to warmer temperatures. For some plants, a cold treatment will speed the process of flowering but is not necessary to initiate flowering. This response is termed a facultative, or quantitative response to cold. Lang (1965) suggests that the most effective temperature range for vemalization of most plants is 1 to

7 °C.

It is well known that most Aquilegia species have an obligate vemalization requirement in order to produce flowers (Shedron and Weiler,

1982). As a result, most Aquilegia species and cultivars will not flower during the

first season from seeds when planted in summer and fall. The vemalization

21

“a! require

‘%.'-

. 9.1. a and the

Weiler 1

leaves,

’0) . A

atrata, I

unheau

before 1 found tl A. xhyb

Cold tre

COIOl’ac

from th havela

(White

requirement has only been studied for a few species and cultivars of Aquilegia, and the duration required for each species and cultivar may differ. Shedron and

Weiler (1982) found that 50 to 100% of ’McKana’s Giant’ plants, averaging 17 leaves, flowered after being given eight and ten weeks of cold, respectively (4.5

°C) . Apparently, three weeks at 5 °C was adequate to induce flowering in A. atrata, A. chrysantha, and A. vulgan's, although plants were grown in an unheated greenhouse and it is unclear if additional chilling may have occurred before or after the 5 °C treatment (Masvidal et al., 1989). Merritt et al. (1997) found that two selections from the Musik series (Ernst Benary, Hanover F .R.G.),

A. xhybrida ‘Musik—Blue White’ and ‘Musik—Red White’ did not flower without a cold treatment.

The Weddle’s Songbird series was released by Weddle Seed in Palisade

Colorado a number of years ago, and has since become very popular. Hybrids from the series, such as ’Bluebird’, ’Blue Jay’, ‘Dove’, ‘Cardinal’, and ‘Robin’, have large showy flowers, some of which are fragrant. In some reports, selections from the Weddle's Songbird series flowered without a cold treatment

(White et al., 1990; Zhang et al., 1991 ). However, these studies suggest that seedlings were grown outdoors in Colorado until mid-October, possibly receiving some natural cooling before the scheduled cold treatments. One hundred percent of ’Bluebird’, ’Cardinal’, ‘Dove’, and ’Robin’ plants flowered when first grown under long days from sowing until they reached 15 to 18 leaves, then cooled for six weeks (0°C) (White et al., 1989). However, only 85% and 72% of these cultivars flowered after five and four weeks of cold treatment, respectively

22 BI pl; lee

far

al.

an

we we ne

ple

Ire

Onl

The in fa (White et al., 1989). ‘Blue Jay’ and ‘Dove’ did not flower without a cold treatment when grown under any of four different day/night temperature regimens (Merritt etaL,1997)

For Aquilegia, cold treatments typically have not induced flowering until plants have made the transition from the juvenile to mature phase (Shedron and

Weiler, 1982). Juvenility has been defined as the early phase of growth during which flowering cannot be induced by any treatment (Thomas and Vince-Prue,

1984). Plant age, or maturity, has typically been defined by the number of leaves on a given plant (Bemier et al., 1981 ). Crown fresh weight may also be a factor in determining when the transition from juvenile to adult occurs (White et al., 1990b). The duration of the juvenile phase varies from species to species and may be quite short in some herbaceous perennials, lasting only days or weeks (Thomas and Vince-Prue, 1984) but usually longer for Aquilegia. Eight- week-old plants of A. xhybrida ‘McKana’s Giant’, with an average of 7 leaves, never reached 100% flowering after cold treatment. However, 12-week-old plants, averaging 12 leaves, reached 100% flowering after ten weeks of cold treatment at 4.5 °C (Shedron and Weiler, 1982). Twelve-week-old plants of A. xhybrida ‘Fairyland’ averaging 12 leaves per plant never reached 100% flowering

, but sixteen-week-old plants averaging 15 leaves reached 100% flowering after only four weeks of cold treatment (Shedron and Weiler, 1982).

There are two established theories for the perception of vemalization :

The first being that vemalization affects leaves and results in a hormonal balance in favor of flowering (Melchers et al., 1936, 1937), secondly, that cold

23

the

IIE

9X

pl;

te

ml

001 be thermoinduces apical cells directly (Schwabe et al., 1954). Localized cooling treatments applied to certain biennial and perennial plants caused flowering when only the stern apex was chilled independent of the temperature experienced by the rest of the plant (Thomas and Vince-Prue, 1984). For other plants, it is apparent that the shoot apex is not the only site where cold temperatures are perceived. Vemalization perception in both the apex and leaves was observed in Pisum sativum L. (Reid and Murfet, 1975). For

Aquilegia, however, leaves are not necessary. In Aquilegia, the central growing point, not the leaf tissue, appears to perceive the cold temperatures (Shedron and Weiler, 1982). Plants of ‘McKana’s Giant’ were cooled both with leaves under light and without leaves in darkness (Shedron and Weiler, 1982). The percent of flowering plants was not significantly different between the plants cooled with or without leaves (Shedron and Weiler, 1982).

The ability to cool small plugs of Aquilegia, as opposed to overwintering plants in pots, is an attractive alternative for many commercial producers. For crops that experience a long juvenile phase, such as Aquilegia, commercial producers must increase the vegetative growth (bulking) of the plants before they can administer cold treatments. In some cases, Aquilegia species are planted from plug trays to larger containers in the fall season and over wintered in the container. In cold climates, the plants in containers can be grown in unheated or minimally heated greenhouses where they receive natural cooling in the container. In mild climates, however, the average winter temperature may not be low enough to provide adequate cooling to induce flowering for many

24 herbaceo’

environm

environm

constant c

production

and maxirr

in plug tray response c well docun

plants in or

Th1

0f differing

Plant heigl Aquilegia 1 mmpare tl

plants COO

"lost COm; were Chos. popular hy herbaceous perennials and it may be necessary to cool plants in a controlled- environment chamber or facility. The ability to cool plants in a controlled- environment chamber allows the producer to provide more accurate and constant cooling and to cool plants at different times of year for staggered production schedules. A grower could shorten production time, reduce labor, and maximize space utilization if the plants could be cooled as young seedlings in plug trays as opposed to larger containers. However, the cold-temperature response of plants in plug trays versus that of plants in containers has not been well documented, nor has a direct comparison been made to overwintering the plants in containers.

The main objectives of this experiment were to 1) measure the influence of differing cold durations on flower timing, flower number, final leaf number, and plant height, 2) establish and compare cold-duration requirements for different

Aquilegia selections and establish the scope of species and cultivar responses 3) compare the influence of differing cold durations on plants cooled in pots versus plants cooled in plug trays, and 4) determine the cold duration that induce the most complete and rapid flowering for each selection. The species and cultivars were chosen to provide a comparison between a wide range of species and popular hybridized Aquilegia cultivars.

25 Material:

Plant ma

Se

A. ch/ysa

and ‘Mini

xhybrida !

xhybn'da 5 grown fror and growr

were ship

the seedlir

into 1.1-L (

16 weeks L

weeks of a!

each remai

396dlings a

with HPS I 8

TWO r

seedlings in

8’5 QC (Tat Count on See seedling Was Materials and Methods Plant material.

Seedlings of Aquilegia alpine L., A. caemlea James. , A. canadensis L.,

A. chrysantha A. Gray. , A. flabellata Siebold & Zucc. ‘Cameo Blue and White’ and ‘Mini star', A. xhybrida Sims ‘Bluebird’ (Weddle Seed, Palisade, Colo.), A. xhybrida Sims ’Crimson Star’, A. xhybrida Sims ’McKana’s Giant, and A. xhybrida Sims ‘Musik White’ (Ernst Benary, Hanover F .R.G) (Bailey, 1976) were grown from seed in 72-cell plug trays (52-ml volume) during the summer of 1997, _____!

and grown outdoors by a single commercial producer (Table 1). All seedlings t were shipped to MSU in plug trays on September 16, 1997. Upon arrival, half of the seedlings of A. alpine, A. caemlea, and ‘McKana’s Giant’ were transplanted

into 1.1-L (5” US.) square pots. These three were the only species older than

16 weeks upon arrival. The rest of the species and cultivars were grown to 14 weeks of age. On the transplant dates listed in Table 1, half of the plugs from

each remaining species were transplanted into 1.1-L (5" US.) square pots. All

seedlings and potted plants were grown at 20 °C under 16-hour days provided with HPS lamps before cold treatments.

Two weeks after transplanting, the plants in 1.1-L pots and the remaining

seedlings in plug trays were transferred to a controlled environment chamber set

at 5 °C (Table 1). Because plants were cutback before shipping, the initial leaf

count on each seedling was not recorded. Instead, crown diameter for each

seedling was measured before they were transferred to the cooler.

26 Cold

enviro

trays h

0800 II

(VHOF

sensor

were we

water ac

seedling

then all 1

There we

Greenhol

Te 20 :0. Ai mmdiamr

Scientific,

SSconds a mmmem each Speci

Cold treatments.

Plants of each species or cultivar were cold-treated in a controlled-

environment chamber at 5 °C for 0, 3, 6, 9, 12, or 15 weeks in the 72-cell plug trays in which they were received or in 1.1-L pots. The chamber was lit from

0800 to 1700 HR at 10 /.¢mol-m'2-s‘1 from cool-white fluorescent lamps

(VHOF96T12; Philips, Bloomfield, N.J.), as measured by a LICOR quantum

sensor (model LI-189; Ll-COR, Inc., Lincoln, NE). While in the cooler, plants

were watered approximately two to three times per week as needed with well has

. ‘12-. water acidified (H2804) to an approximate pH of 6.0. After each cold treatment, .1

seedlings that were cooled in the plug tray were transplanted into 1.1-L pots, then all potted plants were transferred immediately to a 20 °C greenhouse.

There were 10 plants of each species or cultivar per treatment.

Greenhouse environment.

Temperature control. All plants were grown in glass greenhouses set at

20 °C. Air temperatures on each bench were monitored with 36-gauge (0.013-

mm-diameter) type E thermocouples connected to a CR10 datalogger (Campbell

Scientific, Logan, UT). The datalogger collected temperature data every 10

seconds and recorded the hourly average. Actual average daily air temperatures

from the beginning of forcing to the average date of flowering were calculated for

each species and the average temperature varied from 19.9 to 21.5 °C, with

most averaging 20.5 1 0.4 °C.

27 pressur was lit

50am<

COR.

high-y

apprc

dropl 2-1 '3.

Met

COI

prc

Pr: Lighting. All plants were grown under natural light plus light from high- pressure sodium lamps to extend the photoperiod to 16 hours. The greenhouse was lit from 0700 to 0800 HR and from 1700 to 2300 HR at approximately

50umol-m'2-S'1 as measured by a LICOR quantum sensor (model Ll-189; LI-

COR, lnc., Lincoln, NE), yielding a 16-h photoperiod. From 0800 to 1700 HR, high-pressure sodium lamps provided a photosynthetic photon flux (PPF) of approximately 50 umOI°m'2'S" at plant level when the ambient greenhouse PPF dropped to 200 pmoI-m'Z-S'1 ,and discontinued when PPF reached 400 umoI-m'

2~s".

Media and watering

Plants were grown in a commercial soilless medium composed of composted pine bark, horticultural vermiculite, Canadian sphagnum peat moss, processed bark ash, and washed sand (MetroMix 510, Scotts-Sierra Horticultural

Products Company, Marysville, Ohio). Plants were top-watered daily as needed with well water acidified (two parts H3PO4 plus one part H2S04, which provided

22.5 mol P-m3) to a titratable alkalinity of approximately 130 mg calcium bicarbonate per liter and fertilized with 14N-0P-6K20 (mol-m‘a) from potassium nitrate (14-0p-55K20) (Vicksburg Chemical Co., Vicksburg, MS) and ammonium nitrate (34N-0P-0KZO) (Cargill, Lexington, KY). Fertilization and acidification rates were adjusted in response to weekly soil test results, so regimens varied during experiments. The target range for soil pH was 5.8 to 6.2 and, and 0.5 to

1.2 for soil EC.

28 Dana tans bud : have

nonf

rnan wsk:

Dan

and

lies

flow

afie

The

the

the

be

em Data collection and analysis.

Crown diameter was measured with calipers when the plants were transferred to the cooler. For each plant that flowered, the date of the first visible bud and date of the first fully-expanded flower were recorded. Plants that did not have visible buds after 15 weeks of forcing were discarded and considered nonflowen'ng. At flowering, the visible flower bud number, the leaf number on the main stem below the first flower, and total plant height were measured. Days to a ’21-’75? visible bud, days from visible bud to flower, and days to flower were calculated. I

Data were analyzed using SAS‘s (SAS Institute, Cary, NC) analysis of variance '2- and general linear models procedures.

Results and Discussion

A. caerulea, ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik White’ did not

flower under any treatment. Only 10% of A. alpine pot-cooled plants flowered after each cold treatment, while no plug-cooled plants of A. alpine flowered.

Therefore, no data for these five selections are presented. Based on the ages of these seedlings and information from previously cited studies, we expected these plants to flower and we have no explanation for why they did not. It may be possible that the sowing dates received from the commercial producer were erroneous.

29 Flowering F ’Can a cold treat without COII well and he of ‘Bluebirc (I ables 4, percentage pots or in r

Flov

Plug—coole

Cooled pla

1)- Pot-cc weeks of ,

FIOWering

Canaden5 but neVer

S’gnIfiCan

C’"ysé‘mtl Cold, an d Only 600/, Flowering Percentage

’Cameo’, was the only cultivar tested that attained 100% flowering without a cold treatment (Table 2, Figure 1). Thirty percent of ’Mini star’ plants flowered without cold. We presumed that some of the un-cooled plugs would flower as well and have no explanation for why they did not (Table 3, Figure 1). No plants of ‘Bluebird’, A. canadensis, or A. chrysenthe, flowered without a cold treatment

(Tables 4, 5, 6, Figure 1). With increased cold treatment duration, flowering percentage increased for all species studied whether cold was given to plants in pots or in plugs.

Flowering percentage for all species was higher for pot-cooled than for plug-cooled plants. The difference in flowering percentage between the pot- cooled plants and plug-cooled plants was greatest for plants of ‘Bluebird’ (Figure

1). Pot-cooled plants of ‘Bluebird’ reached 100% flowering after only three weeks of cold treatment, while plug-cooled plants never reached 100% flowering.

Flowering percentage for both pot-cooled and plug-cooled plants of A. canadensis increased significantly as the duration of cold treatment increased, but never reached 100% (Table 5, Figure 1). Flowering percentage was significantly higher for the pot-cooled than for the plug-cooled plants of A. chrysanthe. Pot-cooled plants of A. chrysentha reached 40% after six weeks of cold, and 80% after nine weeks of cold, whereas the plug-cooled plants reached only 60% after fifteen weeks of cold (Table 6, Figure 1).

30 Flower I

D: treatmen cooled p were hig flowering bud and and plug points w example

(Figure '

70% rec as the d

Figure ‘l

flower in

'Bluebir decreas

Cooled : treatme

E transple treame Flower Timing

Days to visible bud and days to flower significantly decreased as cold treatment duration increased from 0 to 15 weeks for both pot-cooled and plug- cooled plants for all species. In all cases, the linear contrasts for the decrease were highly significant. The most dramatic change in time to visible bud and

flowering was for plants of A. canadensis. For A. canadensis, the days to visible

bud and days to flower decreased by more than four weeks for both pot-cooled and plug-cooled plants (Table 5, Figure 1). However, in some cases the data

points were based on only one or a small number of flowering plants. For example, only 10% of plug-cooled plants flowered after 6 and 12 weeks of cold

(Figure 1). For plants of ’Cameo’, ’Mini star’, and ‘Bluebird’, there was a 50 to

70% reduction in days to visible bud, and e 30 to 50% reduction in days to flower as the duration of cold treatment increased from 0 to 15 weeks (Tables 2, 3, 4,

Figure 1). Days to visible bud were reduced by about 17 to 21 days, and days to

flower were reduced by about 14 to 21 days for ’Cameo’, ‘Mini star’, and

‘Bluebird’ (Tables 2, 3, 4, Figure 1). Time-to-flower for plants of A. chrysentha

decreased by only two weeks with increasing cold treatment duration for pot-

cooled plants and less than one week for plug-cooled plants with increasing cold- treatment duration (Table 6, Figure 1).

Days to visible bud and days to flower were generally lower for plants

transplanted and grown in pots for two weeks prior to the beginning of the cold

treatment (Tables 2, 3, 4, 5, 6, Figure 1). Pot-cooled plants of A. canadensis

consistently bloomed about 17 days earlier than the plug-cooled plants,

31 independent of cold duration (Table 5, Figure 1). Pot-cooled plants of both

’Bluebird’ and ’Cameo’ consistently flowered about 7 days faster, while pot- cooled plants of A. chrysentha bloomed only a few days faster than plug-cooled plants. There was little, if any, difference in timing for plants of ‘Mini star’ , particularly after long cold treatment durations (Table 4).

Days from visible bud to flower _ .‘ . 1’“

There were statistically significant differences between days fromvisible .. “In-" bud to flower for ‘Mini star’, ‘Bluebird’, A. canadensis, and A. chrysentha in IL..- response to cold treatment, although the actual numerical decreases were small and no trends were observed (Tables 3-6). Time from visible bud to flower ranged from 10 to 16 days for plants of ‘Cameo’ and ‘Mini star’ to 15 to 20 days for A. canadensis and A. chrysentha. In the case of ’Cameo’, days from visible bud to flower increased with increased cold duration and there were significant linear and quadratic contrasts (Table 2). For un-cooled plants of ‘Cameo’ the

first flower bud opened at the base of the leaf axis, whereas with increasing cold there was increased peduncle elongation prior to flower Opening. This physiological response to cold may have accounted for the increase in days from visible bud to flower for ’Cameo’.

There were no significant differences in days from visible bud to flower between the pot-cooled and plug-cooled plants of ‘Cameo’, ’Mini star', A. canadensis, and A. chrysentha.

32 There was a highly significant effect on days from visible bud to flower due to interaction between plant size (pot or plug) and weeks of cold for plants of ’Mini ster’ and ’Bluebird’, but actual numerical differences were slight and no trends were observed (Tables 3 and 4).

Flower Number

In general, cold increased flower number for all species and cultivars studied (Figure 2). The total flower number for plants of ‘Cameo’ increased 2 to

3 times for both pot-cooled and plug-cooled plants as the duration of cold Fifi treatment increased from 0 to 15 weeks (Table 2, Figure 2). There was no significant increase in flower number for plants of ‘Mini star’ (Table 3). The

flower number for ’Bluebird’ and A. chrysentha increased approximately 160-

180% for pot-cooled plants, and 120-130% for plug-cooled plants as the duration of cold increased from 3 to 15 weeks (Table 4 and 6, Figure 2). Flower number of the plug-cooled plants of A. canadensis increased by 320% (Table 5, Figure

2). Conversely, flower number for the pot-cooled plants of A. canadensis decreased by 25% as cold treatment increased from 3 to 15 weeks (Table 5 and

Figure 2). However, this decrease is complicated the low number of flowering plants, particularly at shorter cold treatment durations (Figure 2).

The effects of plant size and cold on flower number varied between the species. Overall, pot-cooled plants had higher flower numbers. The most dramatic difference between pot- and plug-cooled plants was observed for

‘Cameo’, ’Mini star’, and A. chrysentha.

33 For plants of A. canadensis and ‘Bluebird’, there were no significant differences in flower number between the pot-cooled and plug-cooled plants (Tables 4 and

5).

Final leaf number

Cold had a significant effect on final leaf number for all selections tested

(Tables 2-6). In some cases, however, the actual differences were small. Final leaf number for plants of ’Cameo’ and ‘Bluebird’ decreased by about one to two leaves for both the pot-cooled and plug-cooled plants (Tables 2 and 4, Figure 2).

For plants of ‘Mini star’, the final leaf number for pot-cooled plants decreased by three leaves and by seven leaves for plug-cooled plants as cold increased from 0 to 15 weeks (Table 3, Figure 2). Final leaf number for pot-cooled plants of A. chrysentha and A. canadensis decreased by eight and ten leaves, respectively.

Final leaf number for the plug-cooled plants of A. canadensis decreased by two or three leaves but did not change at all for plug-cooled plants of A. chrysentha as cold treatment increased from 0 to 15 weeks (Tables 5 and 6, Figure 2).

Leaf number at first flower was Often lower for the plug-cooled plants, but the actual differences between plug- and pot- cooled plants were small. For plants of ’Cameo’ and ’Bluebird’, the pot-cooled plants consistently had one to two more leaves at flowering than the plug-cooled plants (Tables 2 and 4). Pot- cooled plants of A. canadensis had a significantly higher leaf number then plug- cooled plants, although some of the data points are based on a low number of

flowering plants (Table 5). The decrease in final leaf number of A. chrysentha

34 10111 an!" big

followed a quadratic pattern as the cold duration increased (Table 6). The biggest difference between pot-cooled and plug-cooled plants was observed for

’Mini star’ (Table 3 and Figure 2). After 12 and 15 weeks of cold, pot-cooled

’Mini star’ plants had almost 50% more leaves than the plug-cooled plants (Table

3, Figure 2).

Plant height

Cold treatment duration had a significant but moderate effect on plant height at first flower. Height tended to decrease as the duration of cold increased, but the differences were generally small. Increased cold treatment reduced plant height by about two to three centimeters for ’Cameo’ and ‘Mini star’ plants (Tables 2 and 3, Figure 2). For plants of ‘Bluebird‘ and A. chrysentha, increased cold treatment reduced height for pot-cooled plants by 6 and 10 centimeters, respectively. Cold reduced the height of plug-cooled plants of ‘BIuebird' and A. chrysentha by 3 or 4 centimeters (Tables 4 and 6, Figure

20). For plants of A. canadensis, a longer cold treatment increased plant height

(Table 5).

On average, the pot-cooled plants of ‘Cameo’ and ‘Mini star' were two to three centimeters taller than the plug-cooled plants (Tables 2 and 3, Figure 2).

There was no significant difference in height between the pot-cooled and plug-

cooled plants of A. xhybrida ‘Bluebird’ and A. chrysentha and there was no

significant interaction between plant size (pot or plug) and weeks of cold for

either species (Table 4 and 5, Figure 2). Statistically, there was a difference

35 be dil

-vv..-..u between the pot-cooled and plug-cooled plants of A. chrysentha, although the difference was only noted after the 12-week cold treatment. For plants of A. canadensis, there were variations in height between cold treatments.

When only the 9-, 12-, and 15 week cold treatments are compared, the pot- cooled plants were consistently taller than the plug-cooled plants.

Crown diameter

No trends were observed for crown diameter in relationship to flowering

(averages in Table 7). Some plants with seemingly small crown diameters

flowered, whereas some with larger crown diameters did not flower. No other inforrnetion on crown diameter of Aquilegia was found in the literature. Crown diameters were measured because the plugs had been cutback before shipping by the grower, thus making it impossible to count the leaves.

Conclusion

Not only is there great diversity in plant and flower morphology between species of the genus Aquilegia, but there is also extensive diversity in the types of habitats that they occupy, including warm-temperate forests, extremely high altitude alpine zones, and desert springs (Hodges and Arnold, 1994a). Thus, it is not surprising that there should be differences in how different species respond to environmental signals such as cold temperatures. A vemalization treatment is not required for flowering of ‘Cameo’ or ‘Mini’, but is horticulturally beneficial for both cultivars. Both ‘Mini star’ and the ‘Cameo’ series (released

36 I '5? Irv 2

.

.

.L

IM.II.I.-

._ u *2": .

during 1998 by Benary Seed), are cultivars of the species Aquilegia flebellete, native to the alpine regions of Japan (Bailey, 1976). It was unexpected that an alpine species such as A. flabellate would flower without vemalization, whereas

A. chrysentha, a species native to much warmer areas such as Arizona and New

Mexico, required a lengthy cold treatment in order to flower. A. canadensis, a species that has a broad habitat range from Nova Scotia to Texas, also had an obligate cold requirement.

Aquilegia species remain largely inter-fertile, which suggests that, overall, they may be genetically very similar (Hodges and Arnold, 19943). The name A. xhybrida was originally given to hybrids of A. canadensis and A. vulgan's, but is more often used for long-spurred garden hybrids with a parentage involving A. caerulee, A. chrysentha, and possibly others (Bailey, 1976). In this study we were not able to determine cold requirements of the alpine species A. caerulee, except that it must require cold because no plants flowered in our experiment. A. chrysentha also had an obligate requirement for cold and needed at least six weeks of cold for flowering. One would expect that the traits for vemalization response would be heritable to offspring. A. xhybrida ‘Bluebird’, a hybrid with parentage involving A. caerulee and A. chrysentha, required only three weeks of cold for 100% flowering.

According to previous studies by Shedron and Weiler (1982), White, et al.

(1990), and Zhang et al. (1991), and based on the presumed ages of our

seedlings, we felt confident that plants of all species in the study would flower.

However, five of the species flowered poorly or did not flower at all. According to

37 Shedron and Weiler (1982), 100% of ‘McKana’s Giant’ plants flowered after ten weeks of cold with an average of 12 leaves before cold, and 100% of “Crimson

Star' plants flowered after eight weeks of cold with an average of 15 leaves before cold. Both cultivars were 16 weeks of age when cooled in this study.

Based on the literature, some plants of ‘McKana’s Giant’ and ‘Crimson Star’ in the present study should have flowered based on the age of the plants, 24 and

16 weeks, respectively. No plants of the cultivar ‘Musik—White’ flowered under any cold treatment, and there is little information published on selections from the Musik series (Benary, Hann. Muenden, F .R., Germany). Merritt et al. (1997) chilled seeds of ’Musik- Blue White’ and ‘Musik- Red White’, then forced the seedlings under four different day/ low-night temperature regimens. No plants of either cultivar flowered in their experiment.

If juvenility is not the cause for poor flowering of selections in this study, one possible hypothesis is that defoliated plugs of Aquilegia may not respond similarly to chilling. However, this seems unlikely due to evidence from previously cited work by Shedron and Weiler (1982) where cooling had no deleterious effects on subsequent flowering of defoliated plants. Another possible hypothesis is that high temperatures while plugs were grown outdoors may have been detrimental to the seedlings. High temperatures before cold treatment reduced flowering for plants studied in chapter 3.

Based on studies by White et al. (1989, 1990) and Zhang et el. (1991), we expected plants from the Weddle’s Songbird series to flower without a cold treatment. White and co-workers reported in 1990 and 1991 that plants of

38 ‘Bluebird’, ’Dove’, ‘Purple’, and ’Robin’ flowered five months after seeding without exposure to low temperatures. However, in our experiments, no plants of

’BIuebird’ flowered without a cold treatment. We also received seedlings of

’Cardinal’ and ‘Dove‘ for this experiment but there were only enough plants of each cultivar for one treatment, so the plants were grown without a cold treatment. After 100 days, no plants of either ‘Cardinal’ or ‘Dove’ flowered. Our seedlings were sown in the summer months of 1997 and received on September

16, 1997. The seedlings were then placed in our greenhouses set at 20 °C under long-day photoperiods of 16 hours. The seedlings used in the studies by

White and Zhang had been grown outdoors and were shipped to Pennsylvania

State University in mid-October. It is possible that the plants used in the studies by White and Zhang may have received some natural cooling before forcing

There is little published information on flower timing for species and cultivars of Aquilegia. According to Shedron and Weiler (1982), 50% of

‘McKana’s Giant’ plants flowered after an average of 165 days without a cold treatment under a 10-h photoperiod and after an average of 118 days under 18-h of light. After ten weeks of cold, 100% of plants flowered under short days after average of 76 days and after 70 days under long-days. Days from visible bud to

flower for plants of ‘McKana’s Giant’ generally decreased as the duration of cold treatment increased, but was variable in all experiments (Shedron and Weiler,

1982). According to White et al. (1989), ’Bluebird’ seedlings cooled for a total of six weeks at temperatures from -1.5 °C and 9 °C, then forced at a greenhouse temperature of 15.5 °C under 24 hours of light from HPS lamps, reached visible

39 bud after 29 days and flowered after 45 days. When the cooled plugs were forced at the same temperature under natural short-days in December, plants reached visible bud after 40 days and flowered after 51 days. As the duration of cooling increased, the days from visible bud to flower remained about the same

(White et al., 1989). The timing for ’Bluebird’ in White’s study is similar to the timing of ‘Bluebird’ seedlings in our experiment that were cooled for six weeks.

Our seedlings reached visible bud after 24 days and flowered after 40 days.

White et al. (1990) reported that increased duration of cold actually delayed the days to visible bud for plants of ‘Bluebird’ and ‘Robin’. However, their plants had already initiated flower buds before they were cold treated, thus, the cold temperatures were causing bud abortion or delayed further flower bud development rather than affecting flower initiation. Zhang et al. (1990) reported that there was no effect of cold on days to visible bud or flower for plants of

‘Purple’ and ‘Dove’, however, plants had already initiated flower buds before they were cold treated, therefore, no comparative flower timing can be extrapolated from these two studies. Overall, increased cold durations decreased the time to visible bud and first flower for all flowering species in the present study. Cold duration recommendations for each flowering species and cultivar in the present study are listed in Table 8.

There is little information on the effects of vemalization duration on flower

number, leaf number, and plant height of Aquilegia at the time of flowering.

According to White and Zhang, there was a decrease in flower bud number and

no effect on plant height of Aquilegia plants with increasing durations of cold

40 treatment. However, because flower buds initiated before cold in both of these studies, the results cannot be compared to our results. Our experiments showed an overall increase in flower bud number and an overall decrease in plant height at flowering with increased cold duration, and, to our knowledge our plants had not initiated flower buds before they were cold-treated.

All the flowering species performed better when cooled in pots as opposed to plugs in this study. Pot size in conjunction with bulking time could be an important factor in flowering Aquilegia. One explanation for why Shedron and

Weiler (1982) were able to flower Aquilegia cultivars such as ’McKana’s Giant’ and ’Crimson Star’, and we were not, may be pot size. The plug-cooled plants in our experiment were grown to the same age (16 weeks) as those in Shedron and

Weiler’s study, but our seedlings were grown in 50 ml cells (72-cell plug trays).

Our pot-cooled plants were transplanted from plug trays into 1.1-L (5” US.) square pots and grown for only two weeks before cold treatment. The plants in

Shedron and Weiler’s study were transplanted into 7.5 or 10 cm diameter

(approx. 350 to 470 ml) pots and grown for 4, 8, and 12 additional weeks before cold treatment. It is possible that not only age or leaf number determines the change from juvenile to mature stage in Aquilegia, but also root growth and increased nutrient storage prior to vemalization.

Shedron and Weiler (1982) proposed production of Aquilegia in small cell packs (48 cells per 28 x 53-cm flat). Plants would be transplanted directly into the cell pack, grown to maturity, vemalized, forced, and sold in the same pack.

Due to the low flowering percentages of the plug-cooled plants of most species

41 tested in the present study, the production of these cultivars in 72-cell trays prior to forcing does not seem feasible, at least not at the ages we tested. The exception would be ‘Cameo', which could be cooled in 72-cell trays, then transplanted into pots, and grown to flowering for sale on specific dates. It may be that the other species tested could be produced from cooled plugs as well, perhaps if they were grown in a larger plug size or for a longer period of time before a cold treatment.

42 Literature cited

Bailey Hortorium Staff. 1976. Hortus Third. Macmillan. New York.

Ball, Vic. 1991. Culture by Crop, pp. 329-331, 625-651. In: V. Ball, (ed.) Ball Red Book. 15’h ed. Ball Publishing, Chicago.

Bemier, G., J.M. Kinet, and RM. Sachs. 1981. The Physiology of Flowering. Vol. 1. CRC Press, Boca Raton, Florida.

Chouard, P. 1960. Vemalization and its relation to dormancy. Ann. Rev. of Plant Physiol. 11:191-238.

Hodges, S., and M. L. Arnold. 1994a. Columbines: A geographically widespread flock. Proc. Natl. Acad. Sci. USA. 91 :5129-5132.

Hodges, S. and M. L. Arnold. 1994b. Floral and ecological isolation between Aquilegia fonnosa and Aquilegia pubescens. Proc. Natl. Acad. Sci. USA. 91 :2493-2496.

Lang, A. 1965. Physiology of flower initiation, p. 1380-1536. In: J.G. Atherton (ed). Handbuch der Pflanzenphysiologie. Springer-Vedag, Berlin.

Larson, R.A., W. J. Garrison 8 R. W. Carlson. 1990. Differential responses of alpine and non-alpine Aquilegia species to increased ultraviolet-B radiation. Plant, Cell and Environ. 13:983-987.

Masvidal, L., and D. Lopez. 1989. The adaptation to protected cultivation of three aquilegia species in the Mediterranean areas. Acta Hort. 246:355.

Merritt, R.H., T. Gianfagna, R.T. Perkins Ill, and JR. Trout.1997. Growth and development of aquilegia in relation to temperature, photoperiod and dry seed vemalization. Scientia Hort. 69:99-106.

Reid, J. B., and I. C. Murfet.1975. Flowering in Pisum: The sites and possible mechanisms of the vemalization response. J. of Exp. Bot. 26: 860-867.

Schwabe, W.W. 1956. Factors controlling flowering in the Chrysanthemum. IV. The site of vemalization and translocation of the stimulus. J. Exp. Bot. 95: 389-400.

Shedron, K. G., and T. C. Weiler. 1982. Regulation of growth and flowering in Aquilegia xhybrida Sims. J. Amer. Soc. Hort. Sci. 107: 878-882.

43 ‘I’IAL‘ID

‘I. Vino

Thor

Well ‘ -—- .n-r'm Whi‘ ‘I"’ '.

h.

Zha Vince-Prue, D. 1975. Vemalization, p. 262-291. In: Photoperiodism in Plants. Maidenhead, McGraw-Hill.

Thomas, B. and D. Vince-Prue. 1984. Juvenility, photoperiodism and vemalization, p. 408-439. In: Advanced Plant Physiology. Pitman Publ., London.

Weiler, T. C., and K.G. Shedron. 1986. Aquilegia xhybrida, p. 18-21. In: A.H. Halevy (ed.). CRC Handbook of Flowering. Vol. V. CRC Press, Boca Raton, Florida.

White, J W., D.J. Beattie, and E. J. Holcomb. 1989. Flowering studies with aquilegia cultivars. Acta Hort. 252:219-226.

White, J.W., J. Chen, and DJ. Beattie. 1990a. Gibberellin, light, and low- temperature effects on flowering of Aquilegia. HortScience.25:1422-1424.

White, J.W., H. Chen, X. Zhang, D.J. Beattie, and H. Grossman. 1990b. Floral initiation and development in Aquilegia. HortScience 25: 294-296.

Zhang, X., J.W. White, and DJ. Beattie. 1991. Regulation of flowering in Aquilegia. J. Amer. Soc. Hort. Sci. 116:792-797.

44 Table 1. Ages, sowing dates, transplant dates, and transfer dates for the Aquilegia species used in the experiment. All of the seedlings were grown in 72- cell plug trays (each cell equals 50ml volume). The seedlings were shipped to MSU on September 16, 1997 and grown under a 16-h photoperiod at 20°C. On the transplant dates below, half of the plugs from each species were transplanted into 1.1-L (5" US.) square pots. Two weeks later, the remaining seedlings in the 72-cell plug trays and the plants in the 5" pots were transferred to a 5 °C cooler for cold treatment. The plants that were older than 14 weeks upon arrival were transplanted the day after they were received.

Date Age of plants Species Date transplanted Cold treatments when moved to sown to pots started cooler

Aquilegia alpine 4/9/97 9/17/97 9/30/97 25 wks

Aquilegia caerulee 5/21/97 9/17/97 9/30/97 18 wks

Aquilegia canadensis 6/8/97 9/23/97 10/08/97 16 wks

Aquilegia chrysentha 6/18/97 10/3/97 1 1/17/97 16 wks

Aquilegia flabellata ’Cameo 8/3/97 1 1/20/98 12/5 /97 16 wks Blue 8 White’ Aquilegia flabellate ‘Mini star' 6/17/97 10/3/97 1 1/17/97 16 wks Aquilegia xhybrida ‘Crimson 6/18/97 10/3/97 1 1/17/97 16 wks Star’

Aquilegia xhybrida ’McKana 4/9/97 9/17/97 9/30/97 25 wks Giant"s g Aquilegia xhybrida Musik 6/11/97 9/26/97 10/12/97 16 wks White’ Aquilegia xhybrida Songbird 7/1/97 10/16/97 11/1/97 16 wks ‘Bluebird’

45

TlnthCuCll..-

Table 2. Regrowth and flowering responses of Aquilegia flabellate ’Cameo Blue and White’ following 0 to 15 weeks at 5 °C. Plants were cooled in 72-cell plug trays or in 1.1-L (5" US.) square pots. Following cold treatment, plants were grown at 20 °C under 16-h photoperiods. Flower number, final leaf number, and final plant height were measured at first open flower.

Days to Days from Final plant Weeks of 5 °C Percent flowering ‘ visible visible bud to Days to Flower Final leaf height bud flower Flower number number (cm)

Pots 0 100 29 10 38 4 11 13 3 100 17 12 29 5 10 9 6 1 00 13 1 1 24 6 10 1 1 9 100 1 1 14 25 8 10 1 1 12 100 7 1 5 23 12 1 1 1 1 15 100 8 15 23 12 9 10 Plugs 0 60 35 10 45 3 9 10 70 23 1 1 34 5 8 7 90 23 12 35 5 9 9 100 18 14 33 6 9 9 12 100 15 13 28 7 9 8 15 100 15 15 31 a a 9__

Significance Pot or plug m NS m m . m Weeks of cold m m m m .. m Pot or plug x Weeks of cold NS NS NS ' NS NS

Contrasts 0, 3, 6, 9, 12, and 15 weeks 50 P W m m m m . NS

P m m m m m .. . NS, *, **, *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively 1 Plants that were not in bud after 100 days were considered nonflowering.

46 Table 3. Regrth and flowering responses of Aquilegia flabellate ‘Mini star' following 0 to 15 weeks at 5 °C. Plants were cooled in 72-cell plug trays or in 1 - 1-L ( 5" US.) square pots. Following cold treatment, plants were grown at 20 ° C under 16-h photoperiods. Flower number, final leaf number, and final plant height were measured at first open flower.

Days to Days from Final plant Weeks of 5 °C Percent flowering ' visible visible bud to Days to Flower Final leaf height bud flower Flower number number (cm)

Pots 0 3O 28 14 42 3 18 14 3 80 29 1 1 40 5 18 15 6 100 19 14 33 8 16 15 9 100 19 13 32 7 12 14 12 100 18 16 34 9 15 15 15 100 10 14 24 7 15 15 Plugs 0 0 - - - - - 60 36 1 1 51 4 17 15 80 23 15 44 4 15 12 80 22 13 35 5 10 14 12 90 17 1 1 28 3 1 1 15 70 10 13 24 5 10

Significance Pot or plug NS NS NS '“ ’" NS Weeks of cold m .. m NS m m

Pot or plug x Weeks of cold NS '“ " NS “ "

Contrasts 0, 3, 6, 9, 12, and 15 weeks SC p u" m NS m m .. NS

P om,“ “' NS “' ’ “ NS NS, *, **, *** Nonsignificant or significant at P_<_0.05, 0.01, or 0.001 , respectively ‘ Plants that were not in bud after 100 days were considered nonflowering.

47

Si

Table 4. Regrowth and flowering responses of Aquilegia xhybrida ’Bluebird’ following 0 to 15 weeks at 5 °C. Plants were cooled in 72-cell plug trays or in 1 - 1 -L (5" US.) square pots. Following cold treatment, plants were grown at 20 ° C under 16-h photoperiods. Flower number, final leaf number, and final plant height were measured at first open flower.

Days to Days from Final plant Weeks 015 °C Percent flowering‘ visible visible bud to Days to Flower Final leaf height bud flower Flower number number (cm)

Pots 0 0 ------3 100 30 15 46 10 17 41 6 100 16 19 35 14 17 37 9 100 17 16 32 15 15 33 12 100 12 13 24 15 15 32 15 100 9 15 25 16 16 35 Plugs 0 0 ------10 34 17 50 1 1 15 39 30 24 16 40 15 12 35 9 40 26 17 43 15 15 31 12 30 17 16 33 15 14 33 1 5 20 17 15 31 1 1 13 36

Significance Pot or plug “' NS “' NS “' NS Weeks of cold m m m .. . m Pot or plug x Weeks of cold NS “ NS NS " NS

Contrasts 0, 3, 6, 9, 12, and 15 weeks 5C P m, "' NS “‘ NS NS NS

P W.,, ”" NS '" NS NS NS NS, *, **, *** Nonsignificant or significant at P_<_0.05, 0.01 , or 0.001 , respectively 1 Plants that were not in bud after 100 days were considered nonflowering.

48 Table 5. Regrowth and flowering responses of Aquilegia canadensis following 0 to 15 weeks at 5 °C. Plants were cooled in 72-cell plug trays or in 1.1-L (5" US.) square pots. Following cold treatment, plants were grown at 20 °C under 16-h photoperiods. Flower number, final leaf number, and final plant height were measured at first open flower.

Days to Days from Final plant Weeks of 5 °C Percent flowering ' visible visible bud to Days to Flower Final leaf height bud flower Flower number number (cm)

Pots 0 0 ------3 10 46 17 63 20 24 15 6 10 38 17 55 17 19 22 9 40 29 19 48 16 12 49 12 70 25 17 42 14 13 47 15 7O 17 16 33 15 14 38 Plugs 0 0 ------20 60 17 77 5 15 41 10 56 17 73 9 16 38 20 47 18 65 16 12 34 12 10 49 16 65 14 12 34 15 40 31 15 4,5 16 13 31

Significance Pot or plug m NS m . .1. . Weeks of cold m m m .. m . Pot or plug x Weeks of cold '“ NS “" m t "

Contrasts 0. 3, 6, 9, 12, and 15 weeks 5C P m m NS m m m NS

NS no m, NS P anaemic NS NS NS, *, **, *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively 1 Plants that were not in bud after 100 days were considered nonflowering.

49 Table 6. Regrth and flowering responses of Aquilegia chrysentha following 0 to 15 weeks at 5 °C. Plants were cooled in 72-cell plug trays or in 1.1-L (5" US) square pots. Following cold treatment, plants were grown at 20 °C under 16-h photoperiods. Flower number, final leaf number, and final plant height were measured at first open flower.

Days to Days from Final plant Weeks of 5 °C Percent visible visible bud to Days to Flower Final leaf height flowering ' bud flower Flower number number (cm)

Pots 0 0 ------3 0 ------6 40 37 20 57 15 28 48 9 60 27 17 43 15 24 45 12 80 25 20 45 20 19 39 15 80 24 15 39 27 20 38 Plugs 0 0 ------

40 32 19 51 15 19 45 12 40 29 20 48 17 19 52 15 60 27 18 44 18 19 5]

Significance Pot or plug m NS m m . m Weeks of cold m m m m .. m

Pot or plug x Weeks of cold NS NS NS ' NS NS

Contrasts 0, 3, 6, 9, 12, and 15 weeks 50 P u" m m m ... . NS

p M“ m m m m .. . HS, *, **, *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively 1 Plants that were not in bud after 100 days were considered nonfiowering.

50 Table 7. The average crown diameter measured in millimeters for each species before cold treatment. The diameter of each seedling was measured using calipers before they were moved to the cooler. There were no significant trends in flowering associated crown diameter before cold for any of the species or cu Itivars studied.

Cold breatrfrtients IV6I'I a 6f ' ' ' ' ' méaei’e’aea’zggts Aquilegia Aquilegia #:5417613: flaggllglteag‘llaini 22:11:31]: canadensrs chrysentha ‘Cameo’ star’ ‘Bluebird’

Pots 0 9.7 9.8 10.6 7.8 13.4

3 11.6 8.5 9.9 9.1 11.1

6 10.4 10.0 9.7 9.1 10.8

9 8.7 9.6 9.5 8.1 11.4

12 9.5 9.6 9.3 8.6 11.1

15 8.8 9.2 9.1 8.1 10.8

Plugs 0 9.7 9.2 8.9 8.2 12.9

3 10.8 9.5 9.1 8.6 12.5

6 9.8 8.7 9.0 7.8 10.6

9 10.6 9.7 8.9 8.5 10.5

12 9.3 8.9 9.3 8.9 9.4

15 9.8 9.4 9.2 8.3 10.9

51 Table 8. The recommended cold treatment durations for all flowering species tested.

Species Shortest duration of cold Recommendations required for 100% flowering

Plug-cooled Pot-cooled

Aquilegia flabellate ‘Cameo 9-12 weeks at 5 °C in pots or Blue 8 White’ 9 weeks 0 weeks plugs. Cold not required, but will quickened flowering and increased flower number

Aquilegia flabellata ’Mini 100% 9 weeks at 5 °C star’ flowering 6 weeks in pots . Cold increased never uniformity and quickened time reached to flower

Aquilegia xhybn'da 100% 3 to 6 weeks at 5 °C ‘Bluebird’ flowering 3 weeks in pots.. 3 weeks of cold never was enough for uniform reached flowering, but a longer duration quickened flowering

Aquilegia canadensis 100% 100% 3 12 weeks at 5 °C flowering flowering in pots. Flowering % never never increased with increased reached reached duration of cold treatment, and flowering time decreased

Aquilegia chrysentha 100% 100% _>_ 12 weeks at 5 °C flowering flowering in pots. Flowering % never never increased with increased reached reached duration of cold treatment, and flowering time decreased

52

Figure 1. Effect of cold duration at 5 °C on percent flowering, days to visible bud, and days to flower on plants of Aquilegia flabellata ‘Cameo Blue and White’, A. flabellata ’Mini star’, A. xhybrida ‘Bluebird’, A. canadensis, and A. chrysentha. Plants were cooled either in 1.1-L (5" US.) square pots (O) or in 72-cell plugs (A).

53

8 Bud

A81

Vblblo to

3

Pmntflowirlng Days

:- E 5 g o I m.. 33 i s

n. o

o o 3 a 9 12 15 1e WeeksotCold

100 A s c c c 60 'Bluoblnl' 'Bluobll'd' :2 75 . A- ...... E 40 g 350-“ ~ at 3

E 2 20 . a 3. . 25 .. - 2. m o

'2 o e 9 12 15 15 o WooltsofCoId

100 so A. canadensis gE , Em- 1:S I; — - g 40 8 a i 5 8 6 20 , o 20 e

A n 0 o 6 9 12 1a 0 5 9 12 15 1e 1 :1 6 9 12 15 1a WooluofCold WeeksofCoId WnhofCold

so so A. chrysentha , ‘- chm-nth- A. chrysentha

Ego , _ 60 »

.LW... :40, ,. g“), A, 8 8 ------$20 . 5 20 . a 1:1

0 . . . o ._ e 9 12 o 6 9 12 15 18 o :1 e 9 12 15 1e WuktofCold WeeksofCoId WeeksofCold

Figure 1 .

’11

Figure 2. Effect of cold duration at 5 °C on total plant height, final leaf number, and flower number on plants of Aquilegia flabellate ’Cameo Blue and White’, A. flabellata ‘Mini star', A. xhybrida ‘Bluebird’, A. canadensis, and A. chrysentha. Plants were cooled either in 1.1-L (5" US.) square pots (O) or in 72-cell plugs (A).

55

'Cameo' <5

2:.- E :1 10 K} I T i i E as E= . 5~ 1! - 5 . _ .-_.. g 0- 2 E II. 0 0 . i o" 4._ A # t 3 6 9 12 15 18 o 3 12 1 18 o 3 6 9 12 15 15 Weeks of Cold Weeks of Cold Weeks of Cold 20 25 15 'Mlnlstar' ‘Mlnlstar' 'Mlnlster‘ d ‘ 3 20g. . . , .

E L E g 15 . g .2 :1 :r. ‘ ’5 z E .1 10 . g 3 E T 5 ' ’ E 5 t "

o .

0 3 6 9 12 15 18 Weeks ofCold 25 'Blueblrd' i 20 E i imber. "’ , a ite’, A. g ~ 3 1o - —- » 2 rtha. g "'3 1o .7 ...... , E 5» " 5 .. ~ . — - lugs

o - -4- . o - o O 6 9 12 15 1B 0 3 6 9 12 15 18 0 3 6 9 12 15 18 Weeks of Cold Weeks of Cold Weeks of Cold 60 30 30 A. canadensis A. canadensls A- censdensls S l H I- ‘3: 40 b . g 20 > g 20 I z a E ‘5 T 2

2n. 20 .. 3- 10 .. . .2 10 .. E 2 u

0 0 . . . . . L o ( 3 6 9 12 15 13 0 3 6 9 12 15 18 ( Weeks of Cold Weeks of Cold

40 40 60 A. chryssnm A. chrysentha l

- 3’ 3° : 30 . e“ *5 . b I 3z 20 . 3E 20 .. v5 z .. a 20 ...... i i E 10 .> _ 1o IL IL

0 0 0 6 9 12 15 18 o s 9 12 15 18 r Weeks of Cold Weeks of Cold

Figure 2.

56

«aw CHAPTER III.

THE EFFECTS OF FORCING TEMPERATURE ON THE GROWTH AND

FLOWERING OF AQUILEGIA CANADENSIS, AQUILEGIA FLABELLA TA

‘CAMEO’ AND ‘MINI STAR’. AND AQUILEGIA XHYBRIDA ‘BLUEBIRD’

57 Introduction

Aquilegia, or columbine, are herbaceous perennials belonging to the

Ranunculaceae family. There are about 70 species of Aquilegia native to temperature regions around the globe. Aquilegia are well know wild flowers, and have also been intensely cultivated for use in the perennial border, as potted

flowering plants, and as cut flowers (Ball, 1991). Aquilegia caerulea, A.

Iongissima, A. chrysentha, A. canadensis, and A. vulgaris have been interbred resulting in many popular hybrids of various shapes, sizes and colors. The diversity of the genus provides commercial producers with many opportunities and challenges to adapt Aquilegia to pot-plant culture.

It is well known that most Aquilegia species require some duration of cold in order to produce flowers (Shedron and Weiler, 1982). Most Aquilegia species and cultivars will not flower during the first season from seeds planted in the fall.

For most species of Aquilegia, cold treatments will not induce flowering until the plant has made the transition from its juvenile to mature phase (Shedron and

Weiler, 1982). The duration of the juvenile phase varies from species to species and may be quite short in some herbaceous perennials, lasting only days or weeks (Thomas and Vince-Prue, 1984), but can be quite long for Aquilegia. For example, seedlings of A. xhybrida ‘McKana’s Giant’ required an average of 12 leaves, about 12 weeks of growth at 20 °C, before uniform flowering occurred

(Shedron and Weiler, 1982).

For most herbaceous perennials, forcing temperature plays a significant role in the rate of flower induction and production time (Yuan et al., 1998,

58 Whitman et al., 1996). Many physiological processes, such as flowering, are quickened as temperatures rise until an optimum temperature is reached. This optimum temperature may be different for different physiological processes and different species. At temperatures below a species-specific minimum (Tb), the time to flower (f) is infinite and at temperatures above a ceiling value (Tom)

flowering is delayed (Roberts and Summerfield, 1987). When developmental time is converted to a rate by taking the reciprocal, the relationship between mean temperature and rate of development is often linear in the range between

TD and Top, (Roberts and Summerfleld, 1987). Thus, the relationship between the rate of development toward flowering (1/DTF, where DTF is the days to flower) and temperature can be described as follows

1/DTF=b0+b1 *T [1]

(Yuan et al., 1998). Using the constants b0 and b1, the base temperature, Tb, and degree-days (°days) can be calculated as follows:

Tb = -b0/b, [2]

°days = 1/b1 [3]

(Yuan et al., 1998). Below the base temperature, the rate of progress toward flowering is zero. Degree-days represent the thermal time required for flowering

(Yuan et al., 1998).

Forcing temperature also has an effect on flowering percentage. When seedlings of Pharbitis nil were grown under several different day/night temperature regimens, it was found that the percent of flowering plants

progressively increased as the day temperature increased from 12 to 30 °C

59 (Reese and Emin, 1997). Percent of flowering and flower bud number of

Pharbitis nil were greatest when seedlings were induced with a 24 or 30 °C day temperature and a 30 °C night temperature (Reese and Erwin, 1997).

There is limited information on the effect of forcing temperature on growth and flowering of Aquilegia species and cultivars. Selections from the Weddle’s

Songbird Series reached visible bud and flowered faster with night temperatures of 15 °C than with night temperatures of 10 °C, but no data was given for the

daytime temperatures (White et al., 1989). Day temperatures of 16-19 °C with

average night temperatures between 9 °C and 12 °C induced earlier flowering

than day temperatures greater than 24 °C for several selection of Aquilegia

xhybrida Sims, suggesting heat delay at higher temperatures (Merritt et al.,

1997)

The objectives of our study were to 1) establish and compare the effect of

forcing temperature on the flowering of several Aquilegia species and cultivars

and to establish the scope of species and cultivar responses, 2) measure the

influence of forcing temperature on flower number, flower diameter, and plant height, and, 3) establish the optimum forcing temperature to induce the most complete and rapid flowering for each selection. The species and cultivars were chosen to provide a comparison between a wide range of species and popular hybridized cultivars.

60 Materials and Methods

Plant material.

Species and cultivars studied were Aquilegia alpina L., A. caerulee

James. , A. canadensis L., A. chrysentha A. Gray. , A. flabellata Siebold & Zucc.

‘Cameo Blue and White’ and ‘Mini star’, A. xhybn'da Sims ‘Bluebird’ (Weddle

Seed, Palisade, Colo), A. xhybrida Sims ‘Crimson Star’, A. xhybn'da Sims

‘McKana’s Giant, and A. xhybrida Sims ‘Musik White’ (Ernst Benary, Hanover

F.R.G) (Bailey Hortorium Staff, 1976). Seeds were sown and seedlings were produced by a single commercial producer and shipped to MSU on September

16, 1997. Upon arrival, the seedlings of A. alpine and ‘McKana’s Giant’ were placed directly into a controlled environment chamber set at 5 °C for 12 weeks of cold treatment. The seedlings of the remaining species were grown at 20 °C under 16 hours of extended day light from high-pressure sodium lamps until they

reached 16 weeks of age. Upon reaching 16 weeks of age, the seedlings were transferred to a controlled environment chamber set at 5 °C for 12 weeks. Dates that the seedlings were transferred to the greenhouse are included in Table 1.

Because plants had been cut back before shipping, we were not able to take accurate initial leaf counts on each seedling. There were 10 plants of each

Species or cultivar per treatment.

Cold treatments

Plants of each species or cultivar were cold-treated for 12 weeks in the

72-cell plug trays in which they were received in a controlled-environment

61 chamber at 5 °C. The chamber was lit from 0800 to 1700 HR at 10 umol-m'Z-S'1 from cool-white fluorescent lamps (VHOF96T12; Philips, Bloomfield, N.J.), as

measured by a LICOR quantum sensor (model LI-189; LI-COR, lnc., Lincoln,

NE). While in the cooler, plants were watered approximately two to three times

per week as needed with well water acidified (H2804) to an approximate pH of

6.0.

Greenhouse environment.

Temperature treatments. After cold treatment, the seedlings were

transplanted into 1.1-L (5" US.) square pots and transferred immediately into

glass greenhouses set at 14, 17, 20, 23, and 26 °C. Air temperatures on each

bench were monitored with 36-gauge (0.013-mm-diameter) type E

thermocouples connected to a CR10 datalogger (Campbell Scientific, Logan,

UT). The datalogger collected temperature data every 10 seconds and recorded

the hourly average. Actual average daily air temperatures from the beginning of

forcing to the average date of flowering were calculated for each species (Table

2).

Lighting. All plants were grown under natural light plus light from high- pressure sodium lamps to extend the photoperiod to 16 hours. The greenhouse was lit from 0700 to 0800 HR and from 1700 to 2300 HR at approximately 50

umol-m'2 s‘1 as measured by a LICOR quantum sensor (model Ll-189; Ll-COR,

|nc., Lincoln, NE), to deliver the correct photoperiod. From 0800 to 1700 HR,

high-pressure sodium lamps provided a photosynthetic photon flux (PPF) of

62 approximately 50 umoI-m'z s‘1 at plant level when the ambient greenhouse PPF dropped to 200 umoI-m'2 s“, and discontinued when PPF reached

400 umol-m'z 5".

Media and watering

Plants were grown in a commercial soilless medium composed of

composted pine bark, horticultural vermiculite, Canadian sphagnum peat moss,

processed bark ash, and washed sand (MetroMix 510, Scotts-Sierra Horticultural

Products Company, Marysville, Ohio, year 1). Plants were top-watered daily with

well water acidified (two parts H3PO4 plus one part H2804, which provided z2.5

mol P-m“) to a titratable alkalinity of approximately 130 mg calcium bicarbonate

per liter and fertilized with 14N-0P-6K20 (moi-m3) from potassium nitrate (14-Op-

55K20) (Vicksburg Chemical Co., Vicksburg, MS) and ammonium nitrate (34N-

OP-OKZO) (Cargill, Lexington, KY). Fertilization and acidification rates were

adjusted in response to weekly soil test results, so regimens varied during

experiments.

Data collection and analysis.

For each plant that flowered, the date of the first visible bud and date of

opening of the first flower were recorded. Linear regression analysis was used

to calculate rates of development for each event. At flowering, the number of

visible flower buds, diameter of the first open flower, number of leaves on the

main stem below the first flower, and total plant height were measured. Days to

63 visible bud, days from visible bud to flower, and days to flower were calculated.

Data were analyzed using SAS’s (SAS Institute, Cary, NC) analysis of variance

and general linear models procedures.

Results and Discussion

No plants of A. caerulea, ‘Cn’mson Star’, ‘McKana’s Giant’, and ‘Musik

White’ flowered under any treatment. Only 10 to 20 % of A. alpine, and A.

chrysentha plants flowered under each temperature treatment. Therefore, no

data for these plants are presented. Based on the ages of these seedlings and the results of previously cited studies, we expected these plants to flower and we

have no explanation for why they did not. It may be possible that the sowing

dates received from the commercial producer were erroneous.

Flowering Percentage

‘Cameo’ and ‘Mini star' were the only two cultivars to reach 100%

flowering (Tables 3 and 4, Figure 1). Flowering percentage for both ‘Cameo’ and

‘Mini star’ plants decreased by 60% as the forcing temperature increased from

23 to 26 °C (Table 3 and 4, Figure 1). Flowering percentage for plants of A.

canadensis and ‘Bluebird’ reached only 40-50°/o and 80-90%, respectively

(Tables 5 and 6, Figure 1).

64 Flower Timing

Days to visible bud and days to flower generally decreased as temperature increased, but heat-delay (increased time-to-flower at high temperatures) was observed at 26 °C for both A. canadensis and ‘Bluebird’

(Tables 5 and 6, Figures 4 and 5).

The most dramatic change in time to visible bud and flowering in response to increased temperature was observed for plants of A. canadensis (Table 5,

Figure 4). Days to visible bud and days to flower both decreased by thirty days as the forcing temperature increased from 14 to 23 °C. Flowering of A. canadensis was delayed, however, by about eight days as temperatures increased from 23 to 26 °C (Table 5, Figure 4). For plants of ‘Bluebird’, days to visible bud decreased by 16 days as the temperature increased to 23 °C, and increased by about a day at 26 °C (Table 6, Figure 5). These results suggest that 23 °C is near the optimum temperature, in reference to timing, for A. canadensis and ‘Bluebird’. For ‘Cameo’ and ‘Mini star’, the relationship between temperature and time to visible bud and time to flower followed a linear pattern, with no delay in flowering observed at 26 °C (Tables 3 and 4, Figures 2 and 3).

Days to visible bud decreased by 19 and 22 days for ‘Cameo’ and Mini star’ as temperature increased from 14 to 26 °C (Tables 3 and 4, Figures 2 and 3). Days to flower were reduced by 22 days for both ‘Cameo’ and ‘Mini star' plants as the forcing temperature increased from 14 to 26 °C (Tables 3 and 4, Figures 2 and

3).

65 Days from visible bud to flower was also significantly affected by forcing temperature, but the differences were relatively small in most cases. Days from visible bud to flower for A. canadensis and ‘Mini star’ decreased by five and four days, respectively, as the forcing temperature increased from 14 to 23 °C.

(Table 4 and 5, Figures 2 and 3). Days from visible bud to flower for plants of

‘Cameo’ and ‘BIuebird’ differed by only 3 days and 2 days respectively, over the temperature range tested.

There were significant linear relationships between temperature and rate of progress toward visible bud and flowering for plants of ‘Cameo’ (Table 7). The rate to visible bud and flowering continued to increase linearly as temperature increased from 14 to 26 °C, therefore, all data for ‘Cameo’ was included in the regression analysis (Table 7). Since the rate to flowering increases linearly with temperature only at sub-optimal ranges (Roberts and Summerfield, 1987), the data at 26 °C were excluded from regression analysis for A. canadensis and

‘Bluebird’. Surprisingly, the rate to visible bud for plants of ‘Mini star’ increased as the temperature increased from 23 to 26 °C. Because this response to temperature is atypical, the data for 26 °C was excluded from regression analysis.

The rate of progress from visible bud to flower followed a statistically significant linear pattern in relation to temperature only for A. canadensis (Table

7). The rate of progress from visible bud to flower for ‘Cameo’ increased with temperature (Table 7). Statistically, there was no relationship between the rate of visible bud to flower and temperature for plants of ‘Mini star’ and ‘Bluebird’.

66 Base temperatures and degree-days for each developmental stage of each species were determined using equations [2] and [3]. Base temperature and degree-days can be used to predict the flowering date in commercial greenhouse environments in which temperatures fluctuate (Yuan et al., 1998).

The base temperature for forcing to visible bud and forcing to flower the selections in the present study ranged from 4.2 to 10.6 and -3.9 to 4.7 respectively (Table 7). Base temperature for the rate of progress from visible bud to flower was estimated to be far below 0 °C for ‘Cameo’ and A. canadensis, suggesting that there may have been bud initiation while plants were still in the cooler. There was no significant linear relationship between rate of progress between days from visible bud to flower for ‘Mini star’ and ‘Bluebird’.

Final leaf number

As forcing temperature increased, final leaf number for each species in this experiment decreased linearly (Tables 3—6). Final leaf number for plants of

‘Cameo’, ‘Mini star’, and ‘Bluebird’ decreased by 50-60% as the forcing temperature increased from 14 to 26 °C (Tables 3 and 4).

Flower diameter and number

For all species studied, there was a dramatic decrease in flower diameter, measured across the open face of the flower, as the forcing temperature increased. Flower diameter decreased linearly for plants of A. canadensis and

‘Bluebird’, and quadratically for plants of ‘Cameo’ and ‘Mini star’ with increasing

67 temperature (Tables 3, 4, 5, and 6). For ‘Bluebird’, which featured the largest

flowers of all species studied in this experiment, flower diameter increased two- fold as the forcing temperature decreased from 26 to 14 °C (Table 6, Figure 6).

Flowers of ‘Bluebird’ plants grown at 14 °C measured more than 13 cm across on average. For all species tested, flowers at the highest temperature were also reduced in color, and overall quality, although no data to support these observations were recorded. No gross flower deformities or doubling were observed at 26 °C, though flowers at this temperature were much smaller than those at lower temperatures.

Reducing temperature from 26 to 14 °C promoted a three-fold increase in

flower number for ‘Cameo and ‘Bluebird’ plants (Tables 3 and 6, Figure 6).

There were no significant trends in flower number in relation to temperature for

plants of A. canadensis and ‘Mini star’ (Tables 4 and 5, Figure 6).

Plant height

The effect of forcing temperature on plant height at first flower was

species dependent. Height for A. canadensis plants was independent of forcing temperature (Table 5, Figure 6). Final plant height for plants of ‘Cameo‘ decreased by 25% as the forcing temperature increased from 14 to 26 °C (Table

3, Figure 6). Plant height for both ‘Mini star’ and ‘Bluebird’ decreased by nearly half as the forcing temperature increased from 14 to 26 °C (Table 4 and 6,

Figure 6).

68 Conclusion

Based on the presumed ages of our seedlings, we felt confident that plants of all species in this study would flower. However, three species and three hybrids flowered poorly or did not flower at all. According to Shedron and

Weiler (1982), 100% of ‘McKana’s Giant’ plants with an average of 12 leaves

flowered after ten weeks of cold, and 100% of ‘Crimson Star’ plants with an average of 15 leaves flowered after eight weeks of cold. In these experiments, both cultivars were 16 weeks of age when cooled. Based on this information, some plants of ‘McKana’s Giant’ and ‘Crimson Star‘ should have flowered based on the age of the plants, 24 and 16 weeks, respectively. In our experiment, no plants of the cultivar ‘Musik—White’ flowered under any cold treatment, and there is little information published on selections from the Musik series (Benary, Hann.

Muenden, FR, Germany). Merritt et al. (1997) chilled seeds, but not whole seedlings, of ‘Musik- Blue White’ and ‘Musik- Red White”, then forced the seedlings under four different day/ low-night temperature regimens. No plants of either cultivar flowered in their experiment.

Despite the striking diversity amongst Aquilegia species, there is still a great deal of inter-fertility which suggests that, overall, species of Aquilegia may be genetically very similar (Hodges and Arnold, 1994). The name A. xhybrida was originally given to hybrids of A. canadensis and A. vulgan's, but is more often used for long-spurred garden hybrids with a parentage involving A. caerulea, A. chrysentha, and possibly others (Bailey, 1976). We included parental species in this study for the expressed purpose of comparing the temperature to those of

69 the hybrids. In this study we were not able to determine the response to temperature of the alpine species A. caerulea, A. alpine, or A. chrysentha because no plants , or very few plants, flowered.

Fewer plants of the Aquilegia flabellata selections flowered when forced at

26 °C than when forced at lower temperatures, suggesting that 23 °C is near the optimum for this species. Alternatively, temperatures _>_ 23 °C may have de- vemalized the plants that did not flower. Flowering plants of the A. flabellata selections at 26 °C did not exhibit heat delay, in fact, flowering accelerated for plants of ‘Mini star’. High temperatures did not delay the plants that did flower, but reduced the number of plants that flowered.

For both ‘Cameo’ and ‘Mini star’, higher forcing temperatures greatly reduced leaf number at the time of first flower and the decrease was strongly correlated with faster flowering times for both cultivars.

Flower timing of A. canadensis, a species with a native range from Nova

Scotia to Texas, and A. xhybrida ‘Bluebird’, were most sensitive to temperature compared to A. flabelleta selections, but 100% flowering was never reached for either species and no trends were associated with temperature (Figure 1).

Temperature had a dramatic effect on flower size of all species tested and flower number for two species tested. Flower size, for a given species, can be affected by the rate of plant growth (Pearson et al., 1995). A higher plant growth rate, thus a shorter period of growth, is most likely responsible for a smaller final flower size. Pearson et al. (1995) showed that flower size was reduced at higher temperatures, and further, that the flowers were sensitive to

70 high temperature from the time of induction to anthesis. In two varieties of tomato, heat-sensitive Pusa Ruby and heat-tolerant CL-1311, many floral

anomalies such as stigma exertion without anthesis, empty flowers, and

persistent flowers without fruit-set were observed in when grown under a 35/ 30

°C day/night temperature regimen (Lohar and Peat, 1998). Abnormalities such

as flower doubling occurred for plants of Campanula ‘Birch Hybrid’ when grown

at constant day night temperatures of 29 °C (Frane, unpublished data). No

flower abnormalities were observed in this study for plants of Aquilegia grown at

26 °C, but flower size and color were both reduced.

Overall, plants grown at lower temperatures had more and larger flowers

and were taller, but took longer to reach anthesis than plants grown at lower

temperatures. Plants of all species grown at the highest temperature were too

weak and diminutive in size for 1.1-L (5" US.) pots, whereas the plants of A.

canadensis and ‘Bluebird’ grown at the coolest temperature may have been too

large for them. Settings of 14 to 20 °C are recommended to force A.

canadensis, A. flabellete ‘Cameo’ and ‘Mini star', and A. xhybrida ‘Bluebird’.

Plants flowered slightly slower at temperatures between 14 and 20 °C than those

grown at higher temperatures, but they were much more attractive.

71

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White, J. W. , J. Chen, and DJ. Beattie. 1990. Gibberellin, light, and low- temperature effects on flowering of Aquilegia. HortScience. 25:1422-1424.

72 Whitman, C.M., R.D. Heins, A.C. Cameron, and CH. Carlson.1996. Cold treatments, photoperiod, and forcing temperature influence flowering of Levendula angustifolia. HortScience. 31 :1 150-1 153.

Whitman, C.M., R.H. Heins, A.C. Cameron, and W.H. Carlson. 1997. Cold treatment and forcing temperature influence flowering of Campanula carpatica ‘Blue Clips’. HortScience. 32:861-865.

Yuan, M., W.H. Carlson, R.D. Heins, and AC. Cameron. 1998. Effect of forcing temperature on time to flower of Coreopsis grandiflora, Gail/ardia xgrandiflora, Leucanthemum xsuperbum, and Rudbeckia fulgida. HortScience. 33:663-667.

Zhang, X. , J.W. White, and D.J. Beattie. 1991. Regulation of flowering in Aquilegia. J. Amer. Soc. Hort. Sci. 116:792-797.

73 Table 1. Ages, sowing dates, and transfer dates for the Aquilegia species used in the experiment. All of the seedlings were sown and grown commercially in 72- cell plug trays (each cell equals 50 ml volume). The seedlings were shipped to MSU on September 16, 1997. All seedlings were grown under a 16-h photoperiod at 20 °C until they reached the age of 16 weeks before they were transferred to a 5 °C cooler. The seedlings that were older than 16 weeks upon arrival were transferred to a 5 °C cooler the day after they were received.

Age of plants Species Date Cold treatments when moved to sown started cooler

Aquilegia alpine 4/9/97 9/17/97 23 wks Aquilegia caerulee 5/21/97 9/30/97 16 wks Aquilegia canadensis 6/8/97 10l08/97 16 wks Aquilegia chrysentha 6/18/97 11/17/97 16 wks Aquilegia flabellate 8/3/97 12/5 /97 16 wks ‘Cameo Blue & White’ Aquilegia flabelleta 6/17/97 11/17/97 16 wks ‘Mini star’ Aquilegia xhybrida 6/18/97 11/17/97 16 wks “Crimson Star’ Aquilegia xhybrida 4/9/97 9/17/97 23 wks ‘McKana Giant’ Aquilegia xhybrida 6/11/97 10/12/97 16 wks Musik ‘White‘ Aquilegia xhybrida 7/1/97 11/1/97 16 wks Songbird ‘Bluebird’

74 Table 2. Dates of forcing and the average temperature during forcing.

Average temperature during Species Greenhouse forcing (°C)

temperature Photoperiod (hours) set pornt Date Of 164'" (HID)

during forcing forcing (°C) Force to VB to V8 Flower

Aquilegia flabellate ‘Cameo Blue & White’ 14 2/27/98 14.2 14.2 17 16.9 17.2 20 19.9 20.4 23 23.2 23.4 26 26.1 26.2

Aquilegia flabellata ‘Mini star' 14 2/11/98 14.3 14.4 1 7 1 7.2 1 7.7 20 20.4 19.7 23 23.9 23.1 26 26.4 26.3

Aquilegia canadensis 14 12/31/97 14.3 14.2 17 17.2 17.6 20 20.3 20.1 23 23.7 23.9 26 26.4 26.2

Aquilegia xhybrida ‘Bluebird’ 14 1/24/98 14.2 14.6 17 17.2 17.5 20 20.4 20.0 23 23.9 23.3 26 26.4 26.3

75 Table 3. Regrth and flowering responses of Aquilegia flabellata ‘Cameo Blue and White’ at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16- h photoperiod at temperatures of 14, 17, 20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower.

Final Forcing Temperature Days to Days from Flower plant (°C) Percentage visible visible bud Days to Flower diameter height Final leaf flowering 1 bud to flower Flower number (cm) (cm) number

14 100 27‘“ 15'" 42'” 7'“ 4"“ 8*“ 8'" 17 100 23 15 38 8 3.8 8 8 20 100 16 12 29 8 3.3 8 7 23 100 12 12 24 6 2.4 5 7 26 4p 8 12 20 5 1.7 6 A

Contrasts 14, 17, 20, 23, and 26 ‘0 p m m m m m m m eee

P mm NS NS NS m m m m

NS, *, **, *“ Nonsignificant or significant at P5005, 0.01, or 0.001, respectively 1 Plants that were not in bud after 100 days were considered nonflowering.

76 Table 4. Regrowth and flowering responses of Aquilegia flebelleta ‘Mini star’ at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod at temperatures of 14, 17, 20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower.

Forcing Temperature Days to Days from Flower :l'grai: (°C) Percentage visible visible bud Days to Flower diameter height Final leaf flowering ‘ bud to flower Flower number (cm) (cm) number 14 100 29... 14... 43... 8"“ 3.9... 9... 10"“

17 100 22 15 37 8 3.7 9 10 20 100 17 10 27 7 3.5 10 9 23 100 12 10 23 7 2.8 26 100 Z 13 20 4 1-6

Contrasts 14, 17, 20, 23, and 26 °C P W m .. m m m m m

p Mm NS m to NS m .. NS

NS, *, **, *** Nonsigniflcant or significant at P5005, 0.01, or 0.001, respectively ‘ Plants that were not in bud after 100 days were considered nonflowering.

77 Table 5. Regrowth and flowering responses of Aquilegia canadensis at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod at temperatures of 14, 17, 20, 23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower.

Forcing Days to Days from Flower 22:: Temperature Percentage visible visible bud Days to Flower diameter height Final leaf (°C) flowering ‘ bud to flower Flower number (cm) (cm) number

14 4o 50'" 20m 70m 10~s 3.6"" 36”s 17m 17 50 45 19 64 11 3.3 37 16 20 50 34 17 51 12 3.0 36 15 23 50 25 15 40 10 2.7 34 9 26 40 32 17 4a 9 2.1 35 L

Contrasts 14, 17, 20, 23, and 26 °C P w m m m . m NS m

P qwm m NS m NS . . n

NS, *, **, *" Nonsignificant or significant at P_<_0.05, 0.01, or 0.001, respectively ‘ Plants that were not in bud after 100 days were considered nonflowering.

78 Table 6. Regrowth and flowering responses of Aquilegia xhybrida ‘Bluebird’ at five different forcing temperatures. Plants were cooled in 72-cell plug trays for 12 weeks. Following cold treatment, plants were grown under a 16-h photoperiod at temperatures of 14, 17, 20,23, and 26 °C. Flower number, flower diameter, final leaf number, and final plant height were measured at first open flower.

Forcing Temperature Days to Days from Flower 32:: Final leaf (°C) Percentage visible visible bud Days to Flower diameter height number flowering ‘ bud to flower Flower number (cm) (cm)

14 80 33'“ 14“ 47'” 17"“ 13.3'“ 46'" 15*” 17 80 28 13 42 16 11.3 41 13 20 90 22 12 34 13 9.1 34 14 23 80 18 14 32 8 7.6 34 10 26 80 18 13 31 10 5-7 26 6

Contrasts 14, 17,20, 23, and 26 °C p w m NS m NS m m m

P W "' NS '" '“ NS NS NS

NS, *, ", *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively ‘ Plants that were not in bud after 100 days were considered nonflowering.

79 Table 7. Parameters of linear regression analysis relating forcing temperature to rate of progress to visible bud (VB), to first open flower (FLW), and from VB to FLW in A. flabellata ‘Cameo’ and ‘Mini star’, A. canadensis, and A. xhybrida ‘Bluebird'. Intercept and slope were used to calculate base temperature (TD, and degree-days (°days).

Developmental stage (d) Intercept (b0) Slope (b0) Tb (°C) °days r2 1/d (1 /d)/C

A. flabellete ‘Cameo’ Forcing to was -008035 1 0.011352 0.00756 1 0.00119 10.6 132 0.93 vs to FLW -000907 1 0.00224 0.0022 1 0.00023 -48.0 882 0.67 Forcing to FLW 0.05456 1 0.00436 0.00113 1 0.00046 4.0 453 0.97

A. flabellate ‘Mini star’ Forcing to VB -0.01305 1 0.00395 0.0049 3 0.00055 7.7 204 0.98

VB to FLW -0.01097 _t 0.00154 0.00232 5 0.00016 -67.0 1056 0.08

Forcing to FLW 0.01985 1; 0.01338 0.00347 1 0.00213 4.7 430 0.98

A. canadensis Forcing to VB -0.01305 3 0.00248 0.00217 1 0.00038 6.0 460 0.95

VB to FLW -0.00302 3; 0.00125 0.00114 1 0.00018 -15.0 606 0.95

Forcing to FLW 0.02521 3: 0.00182 0.00165 5 0.00026 2.6 874 0.95

A. xhybrida 'Bluebird' Forcing to VB -0.01204 1 0.00164 0.00286 1 0.00023 4.2 349 0.99

VB to FLW 0.00457 1 0.00135 0.00117 1 0.0002 -24.5 541 0.88

Forcing to FLW 0.04524 1 0.00258 0.00185 1 0.00067 -3.9 855 0.95

:Standard error.

80

100

V 5

1 a : : 7

‘4 0| I

l

V

S

.

Flowering

\ I .. u

... -; 25 1..- - E

Percent A. canadensis V

0 g 4

100 —e e e

l. t g 75 § 2IL 50 . 22 - - .._- __.-

a g i

3 25 r.____,_--.,_-..z W-,. _ ,_ ., ., _. . , W... W.,, .. __ _ - ._ W.W.”..- ,, .. n. 'Cameo' 0 f 1 1

on I: t 3 2

‘5 .

“- 'Mlnlstar‘ o . 4

100

E 75 O 3 2 u. 50 . _._ - - - H

U 5 . 0 E 25 .. .2 .. - _ _._ , 'Bluebird' o 1311818202224 26 Temperature (°C) F'Qure 1. The effects of forcing temperature on the flowering percentage of A. canadensis, ‘Cameo’, ‘Mini star’, and ‘Bluebird‘. Plants that did not flower within 00 days were considered non-flowering.

81 Forcing to Visible Bud Visible Bud to Flowering Forcing to Flowering

14181820222428 {41818202227428 14161820222426 Temperature (°C)

Figure 2. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates ot progress to each event for A. flabellata 'Cameo'. The points represent the actual data points and the curves and straight lines were calculated from regression analysis.

82 Forcing to Visible Bud Visible Bud to Flowering Forcing to Flowering

14101020222420 1410102022242014101020222420 Temperature (°C) Figure 3. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates of progress to each event for A. flabellate ‘Mini star". The points represent the actual data points and the curves and straight lines were calculated from regression analysis.

83 Forcing to Visible Bud Visible Bud to Flowering Forcing to Flowering

0.02

0.00-...%-. W 14101020222420 1410102022242014101020222420 Temperature (°C) Figure 4. The effects of forcing temperature on the days to visible bud, days from visible bud to flower, and days to flower, and the rates ot progress to each event for A. canadensis. The points represent the actual data points and the curves and straight lines were calculated from regression analysis.

84 Forcing to Visible Bud Visible Bud to Flowering Forcing to Flowering

1'41T61'820222426 1416182022242614161820222426 Tomporature(°C) Figure 5. The effects of forcing temperature on the days to visible bud, days fromvisible bud to flower, and days to flower, and the rates of progress to each event for A. xhybrida 'Bluebird'. The points represent the actual data points and the curves and straight lines were calculated from regression analysis.

85

first at 'Bluebird' measured height 14161620222426

D plant 20

and diameter,

canadensis

A. flower

buds, (°C) 14161620222426

flower of 'Mlnlstar’ Temperature number on

deviation.

F“

temperature 'o O

standard N 9 forcing

l of l show l i bars Influence

Error r 6. g l l i [E r 1416162022242614161020222426

l L

m l 8 '- 2 O v N O a c; O 3 a N Figure Jequmu pna (we) Jezemalp JOMOH (ma) zubleu weld flower.

86 CHAPTER IV

THE EFFECTS OF TEMPERATURE, PLANT AGE, COLD TREATMENT, AND

PHOTOPERIOD ON THE GROWTH AND FLOWERING OF AQUILEGIA

FLABELLA TA ‘CAMEO', AQUILEGIA XHYBRIDA ‘BLUEJAY’, ‘CRIMSON STAR',

‘MCKANA'S GIANT' AND ‘MUSIK-WHITE'.

87 Introduction

One of the most distinctive group of plants in the Ranunculaceae family is that of the genus Aquilegia, commonly known as columbine. There are about 70

Aquilegia species that are native to the temperate and mountain regions of the northern hemisphere (Bailey Hortorium Staff, 1976). Due to the wide geographical distribution of the genus Aquilegia, there are many environmental factors that influence the physiology of each species.

Leaf unfolding rate increases with increasing temperature to a maximum and then decreases with continued increased temperature (Karlsson, 1992). The ability to follow progression of leaf development would facilitate scheduling and timing of Aquilegia crops. Average daily temperature controls rate of leaf development in Easter lily, and leaf counting is used successfully to monitor and schedule the crop (Friend et al, 1962; Kansson et al., 1982; Tollenaar et al,

1979). Determining leaf unfolding rates for Aquilegia species and cultivars at a

range of daily average temperatures would promote proper scheduling.

Aquilegia typically do not flower during the first season of growth due to juvenility and a requirement for vemalization. Therefore, it is necessary to

determine an appropriate measurement of maturity, such as leaf number. Plant

age, or maturity, has typically been defined by leaf number for many plants

(White, 1990). Cold treatments will not induce flowering until the plant has made

the transition to its’ mature phase.

Published information provides only general recommendations for

determining plant age or maturity of Aquilegia. Shedron and Weiler (1982)

88 studied the juvenile stage of Aquilegia xhybrida ‘McKana’s Giant’, ‘Fairyland’, and

“Crimson Star’. Plants were chilled at three different ages; 8, 16, and 20 weeks.

‘McKana’s Giant” plants with an average of 6.7 leaves did not flower, nor did

‘Fairyland’ plants with and average of 7.8 leaves or ‘Crimson Star‘ plants with an average of 7.6 leaves (Shedron and Weiler, 1982). After 16 weeks of age and an average number of 12 to 15 leaves, all three cultivars flowered with varying percentages following 6 different cold treatments; 0, 4, 8, 10, and 12 weeks of cold treatment at 4.5 °C. One hundred percent of ‘McKana's Giant’ plants ”'l

flowered at the 12-leaf stage after ten weeks of cold, and all ‘Fairyland’ and 1‘2 ’2‘“

“Crimson Star’ plants flowered at the 15-leaf stage after eight weeks of cold treatment (Shedron and Weiler, 1982). However, because only average leaf numbers were given for each age group, it cannot be determined exactly what plants with a specific leaf number flowered or did not flower.

The information from Shedron and Weiler‘s study has served as the basis for much of the subsequent research on Aquilegia. Latter publications cite

Shedron and Weiler (1982) and have used plants with 12 to 15 leaves before administering treatments. However, this leaf number range may not to be valid for every species or cultivar. White (1990) investigated floral initiation and development in Aquilegia by scanning electron microscopy using selections from the Weddle Songbird series. Plants with a minimum of 12 to 14 leaves usually produced an average of only one flower (White, 1990) but it is not clear from the data presented if 100% of plants with 12 leaves initiated flower buds.

89 A cold requirement for flowering is often linked with a requirement for a long photoperiod after cold, and the requirement for vemalization is most commonly, but not only, found in long day plants (Thomas and Vince-Prue, 1997).

Aquilegia is considered a day-neutral plant after a cold treatment, and will flower under both long days and short days. However, Aquilegia could also be considered a facultative long-day plant because plants will flower faster under long-days (Shedron and Weiler, 1982). After a 12 week cold treatment, cultivars of Aquilegia xhybn'da reached visible bud approximately 20% faster when grown _ -LM‘J under 18-hour long days (7 to 18 pmol rn'2 sec‘1 incandescent lighting) as - 2.“. compared to those grown under short 9-hour days (Shedron and Weiler, 1982).

Days from visible bud to flower, however were not significantly affected by photoperiod.

There are morphological differences in plants grown under long days that may be considered undesirable. Leaf petioles were highly elongated compared to those of plants grown under short days and total plant height at flowering was also greater for plants grown under long days (Shedron and Weiler, 1982).

Plants grown under 10-hour days after 12 weeks of cold had an average height of

43 cm whereas plants grown under 18-hour days after 12 weeks of cold had an average height of 52cm (Shedron and Weiler, 1982).

The objectives of the present study were to 1) measure leaf unfolding development of five cultivars of Aquilegia and calculate rates of leaf development at four average daily temperatures; 17, 20, 23, and 26 °C , 2) determine the age or leaf number at which the plants make their transition from the juvenile to

90 mature stage of development, 3) determine if different temperatures could hasten the transition from the juvenile to mature stage of development, and 4) determine the effects of temperature during plug growth, and of photoperiod before and after

a cold treatment on the growth and flowering of each cultivar.

Materials and Methods

General. Seedlings of A. flabellata Siebold & Zucc. ‘Cameo Blue and

White’, A. xhybrida Sims ‘Bluejay’ (Weddle Seed, Palisade, Colo.), A. xhybrida “1 A.

Sims ‘Crimson Star’, A. xhybrida Sims ‘McKana’s Giant, and A. xhybn'da Sims _m

5... lb

“Musik White’ (Ernst Benary, Hanover F .R.G) (Bailey, 1976) were sown on the

same day and grown for four weeks in 128-cell plug trays (35-ml volume) by a

single commercial grower, then shipped to MSU on October 13, 1997. Upon

arrival, the seedlings were placed in a 20 °C greenhouse under natural days. On

October 15, 1997, the seedlings were placed into Expt 1.

Plants were grown in a commercial soilless medium composed of

composted pine bark, horticultural vermiculite, Canadian sphagnum peat moss,

processed bark ash, and washed sand (MetroMix 510, Scotts-Sierra Horticultural

Products Company, Marysville, Ohio, year 1). Plants were top-watered with well

water acidified (two parts H3PO4 plus one part H2804, which provided z2.5 mol

P-m‘3) to a titratable alkalinity of approximately 130 mg calcium bicarbonate per

liter and fertilized with 14N-0P-6KZO (mol-m‘3) from potassium nitrate (14-Op-

55K20) (Vicksburg Chemical 0., Vicksburg, MS) and ammonium nitrate (34N-0P-

OKZO) (Cargill, Lexington, KY). Fertilization and acidification rates were adjusted

91 in response to weekly soil test results, so regimens varied during experiments.

Our target range for soil pH was 5.8 to 6.2 and, and 0.5 to 1.2 for soil EC.

All cold treatments were delivered in a 5 i 0.5 °C controlled environment chamber lit for 9 h-d’1 at approximately 10 umol-m'Z-S'1 with cool-white fluorescent lamps (VHOF96T12; Philips, Bloomfield, N.J.), as measured by a LICOR quantum sensor (model Ll-189; Ll-COR, lnc., Lincoln, NE). Plants were cold- treated in the 128-cell plug trays in which they were received or the 1.1-L (5" 331 US.) square pots they were transplanted into. While in the cooler, plants were

watered two to three times a week as needed with well water acidified (H2804) to

IN". E

an approximate pH of 6.0.

Leaf unfolding rates (Expt. 1). On October 15, 1997, 128 plant of each

Cultivar were grown in plug trays at each of four different temperatures; 17, 20,

23, and 26 °C, to three ages; 12 (group1), 16 (group 2), and 20 (group3) weeks.

New, fully expanded leaves were counted at 5, 6, 7, 8 and 12 weeks of age for

each individual plug in group 1 at each temperature. Leaf counts were taken at 5,

6, 7, 8, 12, and 15 weeks of age for group 2, and 5, 6, 7, 8, 12, 16, and 20 weeks for group 3. New leaves were marked with a dot of liquid paper correction fluid.

Leaf un-folding rates were calculated for each temperature. Linear regression analysis was used to determine and describe the rate of leaf unfolding within the range of temperatures studied. Linear regression functions were developed for the rate of leaf unfolding per day for each cultivar studied.

92 All plugs were grown under natural light plus light from high-pressure sodium lamps at PPF levels of 50 umol-m'2 s" to extend the photoperiod to 16 hours. From 0800 to 1700 HR, high-pressure sodium lamps provided a photosynthetic photon flux (PPF) of approximately 50 umol-m'2 s‘1 at plant level when the ambient greenhouse PPF dropped to 200 umol-m‘2 s‘1 and discontinued when PPF reached 400 umol-m'2 s‘1 as measured by a LICOR quantum sensor

(model Ll-189; Ll-COR, lnc., Lincoln, NE). "1 -_'-n -' .L

Effects of temperature, plant age, and leaf number before cold, and the effects of p... 6 ‘. photoperiod before and after cold. (Expt. 2).

For Expt. 2, a third of the plugs of each cultivar from groups 1, 2, and 3 in

Expt. 1 were removed from the greenhouse at 12, 16, and 20 weeks of age.

One-half of these plugs were placed directly into a 5 °C cooler for 12 weeks. The other half were transplanted directly into 1.1-L (5" US.) square pots, then grown to the ages of 20 (group 4), 24 (group 5), and 28 (group 6) weeks. Half of these potted plants were grown for the eight week period under a 4-hour night interruption (Nl) from 2200 to 0200 HR and the other half under 9 hours of continuous natural light before cold treatment. Potted-plants were then transferred to a 5 °C cooler for 12 weeks.

After the cold treatment, plug-cooled seedlings from groups 1, 2, and 3 were transplanted into 1.1-L (5"U.S.) square pots. Half of the plugs from each group were grown under NI from 2200 to 0200 HR and the other half under 9 hours of continuous natural light in a greenhouse set at 20 °C. The Ni

93 photoperiod was completed with incandescent lamps at 1 to 3 umol-m‘z-s". High-

pressure sodium lamps provided a photosynthetic photon flux (PPF) of

approximately 50 umol-m'2 8'1 at plant level when the ambient greenhouse PPF

dropped to 200 limol-m'2 s‘1 and discontinued when PPF reached 400 umol-m'2 s‘1

as measured by a LICOR quantum sensor (model Ll-189; Ll-COR, lnc., Lincoln,

NE).

After cold treatment, the pot-cooled plants from groups 4, 5, and 6 were

grown in a greenhouse set at 20 °C, under 9 hours of natural light plus light from

high-pressure sodium lamps at a PPF level of 50 umol-m'2 s‘1 to extend the

photoperiod to 16 hours. From 0800 to 1700 HR, high-pressure sodium lamps

provided a photosynthetic photon flux (PPF) of approximately 50 umol-m'z s‘1 at

plant level when the ambient greenhouse PPF dropped to 200 umol-m'2 s‘1 and

discontinued when PPF reached 400 pmol-m'Z-S'1 as measured by a LICOR

quantum sensor (model Ll-189; Ll-COR, lnc., Lincoln, NE). Due to a lighting

problem in the greenhouse, data for the 12-week-old plugs grown under SD are

not presented for the cultivars studied.

Counts of new, fully-expanded leaves were conducted for the potted plants at 15 and 20 weeks of age for group 4 , weeks 20 and 24 for group 5, and weeks

24 and 28 for group 6 in order to establish leaf number before cold treatment.

The procedure for leaf-counting is as described in experiment 1. Air temperatures on each bench were monitored with 36-gauge (0.013-mm-diameter) type E thermocouples connected to a CR10 datalogger (Campbell Scientific, Logan, UT).

The datalogger collected temperature data every 10 seconds and recorded the

94 hourly average. Actual average daily air temperatures from the beginning of forcing to the average date of flowering were calculated for each species. 480 plugs per cultivar were used to provide 10 plants per treatment.

Data collection and analysis

The initial leaf number was recorded for each plant before the cold treatment. Date of the first visible bud and date of opening of the first flower were recorded for each plant. At flowering, the number of visible flower buds, and total -1 plant height were determined. Days to visible bud, days from visible bud to

flower, and days to flower were calculated.

Results and Discussion

Leaf unfolding rates (Expt 1). All cultivars studied had similar rates of leaf unfolding. Increasing temperature from 17 to 26 °C slightly increased total leaf number (Table 3, Figure 1). A peak leaf unfolding value was observed for

‘Bluejay’ and ‘Crimson Star’ at 23 °C (Table 3, Figure 1). For ‘Cameo’.

‘McKana’s Giant”, and ‘Musik—White’, the rate of leaf unfolding rate continued to increase linearly as temperature increased from 17 to 26 °C (Table 3). Maximum rates for all cultivars were between 0.14 and 0.13 leaves/day (Table 3). Thus, 7.3 days and 7.9 days are expected to be required to unfold one leaf for each cultivar.

The overall effect of temperature on the rate of leaf unfolding was small for all cultivars studied. The difference in rate of leaf unfolding per day from the lowest temperature to the temperature that promoted maximum leaf unfolding

95 (M .Li‘. ‘9! ..

9'

4.1:- ‘r rate was 0.004 leavesl day for ‘Crimson Star‘ , 0.025 leaves/ day for ‘McKana's

Giant’, and 0.03 leaves/ day for ‘Bluejay’ and ‘Musik-White’ (Table 3). The largest

difference observed was for plants of ‘Cameo’. which ranged from 0.06 leaves

unfolded/ day at 17 °C, to 0.14 leaves/day at 26 °C (Table 3). The temperatures

that promoted the highest rate of leaf development in Aquilegia cultivars, 23 to 26

°C, were similar to other plants such as Saintpaulia ionantha (African Violet)

(Faust and Heins, 1993).

Effects of temperature, plant age, and leaf number before cold, and the effects of photoperiod before and after cold. (Expt. 2).

Initial leaf number. Increasing temperature before cold treatment

increased total leaf number (Table 9). The temperature during plug growth had a

lasting effect on seedlings from groups 4, 5, and 6 that were transplanted and

grown for eight additional weeks at 20 °C prior to cold treatment. The plants that

were previously grown at higher temperatures consistently maintained a slightly

higher leaf number than those previously grown at lower temperatures (Tables 4-

8, Figure 2). As expected, older seedlings had more leaves than younger

seedlings (Tables 4-10, Figure 2).

There were some statistically significant effects of photoperiod on leaf

number for the pot-cooled plants of all cultivars (Tables 4-8). Plants grown under

NI lighting before cold treatment almost always had a higher leaf number than those grown under SD lighting (Tables 4-8, Figure 2). The greatest differences in

leaf number in relation to photoperiod were observed for plants of ‘Bluejay’ and

‘Cameo’. For example, the 20 week-old potted plants of ‘Bluejay’ averaged 13.2

96 to 14.7 under SD, and 15.1 to 18.8 leaves under NI before cold treatment (Table

5, Figure 2). The 20 week-old potted plants of ‘Cameo’ averaged 11.9 to 12.1

leaves under SD, and 12.5 to 17.6 leaves under Nl lighting (Table 5, Figure 2).

Differences were less significant for plants of “Crimson Star’ , ‘McKana’s Giant’,

and ‘Musik-White’ (Tables 6-8).

There was no effect of photoperiod on the initial leaf number of the plug-

cooled plants because different photoperiods were not delivered to these plants

r“. l . until after cold treatment (Tables 6-8, Figure 2). '7,

Flowering percentage. Initial leaf number before cold treatment had a

significant effect on percentage flowering for all cultivars studied. All pot-cooled

plants of ‘Cameo’. ‘Cn'mson Star‘, and ‘McKana’s Giant’, had _>_ 10 leaves before

cold treatment. One hundred percent of these plants flowered with the exception

of the 10-Ieaf ‘McKana’s Giant’ plants grown under Nl lighting before cold (Tables

4,5, and 7, Figure 3).

All pot-cooled plants of ‘Bluebird’ or ‘Musik-White had 3 11 or 14 leaves,

respectively, before cold treatment (Tables 4 and 8, Figure 3). No pot-cooled plants of ‘Bluejay’ or ‘Crimson Star’ flowered with an initial leaf number 5 12

(Tables 4 and 6, Figure 3). All plants of both cultivars with an initial leaf number

>12 flowered, with the exception of the 13-leaf plants of ‘Bluejay’ grown under SD before cold (Tables 4 and 6, Figure 3).

Flowering percentage for the plug-cooled plants of all cultivars was far

lower than for pot-cooled plants (Tables 4-8, Figure 3). According to the results

97 of the present study, leaf number before cold is not the only factor determining

ripeness to flower. In most cases, pot-cooled plants reached a much higher

flowering percentage than plug-cooled plants with the same initial leaf number

(Tables 4-8, Figure 3). For example, no plug-cooled plants of ’Musik-White’ with

14 initial leaves flowered, whereas all 14-leaf pot-cooled plants flowered (Table 8,

Figure 3). In fact, 100% flowering was never reached in any treatment for plug-

cooled plants of ‘Musik—White’ (Table 8, Figure 3). Only several treatments of

plug-cooled plants of ‘Bluejay’, ’Crimson Star', and ‘McKana’s Giant’ reached I.

100% flowering (Tables 4-7, Figure 3). ‘Cameo’ was the only cultivar in which

the plug-cooled plants flowered similarly to the pot-cooled plants (Table 5, Figure

7). All plug-cooled plants of ‘Cameo’ with an initial leaf number _>_ 5 flowered

(Table 5, Figure 7). Only plug-cooled plants of ‘Cameo’ with an initial leaf number

< 5 did not flower or did not reach 100% flowering (Table 5, Figure 3).

Temperature before cold treatment greatly affected flowering percentage for plug-cooled plants (Tables 4-8). Flowering percentages for ‘McKana’s Giant’,

‘Crimson Star’ , ‘Bluejay’, and ‘Musik—White’ were higher for plugs that were grown at 17 and 20 °C as opposed to 23 and 26 °C (Tables 4-8).

Photoperiod also affected flowering percentage for plug-cooled plants, overall, more plants flowered when grown under SD after cold than under NI lighting

(Table 4-8, Figure 3)

Flower timing. Overall, the temperature at which the plugs were grown before cold treatment, and the photoperiod plants were grown under before and

98 after cold, both had an effect on days to visible bud after cold treatment for all

cultivars studied (Tables 4-8, Figure 8). Plants grown under SD before cold

flowered slightly faster than plants grown under NI before cold. Conversely,

plants forced under SD after cold flowered slightly slower than plants forced

under NI, although, the actual differences were small and no trends were

observed (Tables 4-8).

Total plant age had a significant effect on days to visible bud for all

cultivars tested (Tables 4-10). For example, days to visible bud for pot-cooled

and plug-cooled plants of ‘Bluejay’ decreased by and average of 6 and 14 days

as age increased from 20 to 28 and 12 to 20 weeks, respectively (Table 4, Figure

5). Days to visible bud for pot-cooled plants of ‘Cameo’ decreased by an average

of 6 days, and 9 days for plug-cooled plants of ‘Cameo’ (Table 5, Figure 5). Days

to visible bud for pot-cooled plants of ‘Crimson Star’ decreased by an average of

14 days, but decreased by only four days for plug-cooled plants (Table 6, Figure

5). Highly significant linear and quadratic trends were observed for ‘Bluejay’,

“Crimson Star’, and ‘Cameo’ (Tables 4-6). Significant quadratic trends were

observed for plants of ‘McKana’s Giant’ and ‘Musik-White’ (Tables 7 and 5).

Days to visible bud for pot-cooled plants decreased by an average of 14 days for

both ‘McKana’s Giant’ and ‘Musik-White’ as age increased from 20 to 28 weeks,

but varied with age for the plug-cooled plants (Tables 7 and 8, Figure 5).

As leaf number increased with age, there were statistically significant decreases in days to flower for all cultivars studied (Tables 4-8). However, no linear or quadratic trends were observed for plants of ‘McKana’s Giant’ or ‘Musik—

99 White‘ (Tables 7 and 8). Significant linear and quadratic trends were observed for

plants of ‘Bluejay’, ‘Cameo’. and ‘Crimson Star’ and days to flower decreased with

increased leaf number (Tables 4, 5, and 6, Figure 6). For plants of ‘Bluejay’ and

‘McKana’s Giant’, the time from visible bud was affected by total plant age as

initial leaf number increased (Table 4 and 6, Figure 6). As leaf number increased,

the days from visible bud to flower decreased slightly and significant linear and

quadratic trends were observed (Tables 4 and 6). There were no significant I.“ trends for days from visible bud to flower for plants of ‘Cameo'. ‘Crimson Star’, or '

‘Musik-White’ (Tables 5, 6, and 8). Overall, days to visible bud and flower were SJHK"

significantly lower for pot-cooled plants than plug-cooled plants (Tables 5-9).

Flower number. Cooling plants in pots had a significant effect on flower number for all cultivars (Tables 4-8). Pot-cooled plants of all cultivars tested consistently had a higher flower number than plug-cooled plants of the same age

(Tables 4-8, Figure 7). For example, 20 week-old pot-cooled plants of ‘Bluejay’ had an average of 22 flowers at the time of first flower, whereas 20 week-old plug-cooled plants had an average of only 9 flowers (Table 4, Figure 7). This trend was similar for all other cultivars tested (Tables 4-8).

Plant age in weeks had a significant effect on flower number and significant linear and quadratic trends were observed for all cultivars tested

(Tables 4-8). Flower number of plug-cooled plants increased as age increased from 12 to 20 weeks, however, flower number for the pot-cooled plants actually

decreased as age increased from 20 to 28 weeks (Tables 4-8, Figure 7). For

100 example, flower number for the plug-cooled plants of ‘Musik-White’ increased by

9 as age increased, but decreased by 7 for the pot-cooled plants as age increased from 20 to 28 weeks (Table 8, Figure 7). Flower number or the pot- cooled plants may have decreased because they flowered earlier than plug- cooled plants. Some flower buds may develop after the first flower opens, and some may not yet be visible, therefore, the number of visible flower buds at first

flower may not be an accurate estimation of total flower number.

Plant height. Plant size (pot-cooled versus plug-cooled), age, photoperiod, 2.- and temperature before cold, all affected plant height (Tables 4-8). Plant height was generally greater for older plants cooled in pots, and there were significant linear and quadratic trends for ‘Bluejay’, ‘Cameo’. Crimson Star‘, and ‘McKana’s

Giant’ (Tables 4—7). There was no significant difference in plant height at flower between the pot-and plug-cooled plants of ‘Musik—White’ (Table 8, Figure 8).

Overall, plants grown under NI lighting before or after cold were significantly taller than those grown under SD (Tables 4-8, Figure 8). Temperature before cold treatment significantly affected plant height for plants of ‘Bluejay’ and ‘Crimson

Star’ with linear and quadratic trends, respectively (Tables 4 and 6). Plant height tended to decrease as the pre-cold temperature increased. There were no significant effects of temperature before cold for ‘Cameo’. ‘McKana’s Giant’, or

‘Musik-White' (Tables 5, 7, and 8, Figure 8).

101 Conclusion

The results of the present study indicate that faster leaf unfolding and vegetative development may be achieved at high temperatures for the Aquilegia cultivars studied. In the temperature range of 17 to 26 °C , the leaf unfolding rate showed minor variations and remained above 0.13 leaves/day for all cultivars tested as the predicted peak leaf unfolding rate was approached at 23 °C for two cultivars and 26 °C for three cultivars. Overall, the differences in leaf unfolding rates between temperature treatments were small. Because vapor pressure .1 Aim-h deficit (VPD) was not monitored in the greenhouse, it is possible that the . Fl difference in shoot tip temperature was smaller than the range of air temperatures. VPD increases at higher temperatures and causes evaporation from the soil surface, which may result in a lower shoot tip temperature.

Aquilegia remain in a rosette until bolting and leaves unfold close to the soil surface. Combined, these factors may account for the small difference in leaf unfolding rates in relation to air temperature. In most practical applications, this small difference in leaf unfolding rate is unimportant, but the rates of leaf unfolding can be used to monitor the vegetative plant development in the cultivars of Aquilegia studied. The rates can be used to predict how many weeks are necessary at specific temperatures for these cultivars of Aquilegia to develop enough leaves for maturity and subsequent flowering.

The higher temperatures may have promoted a higher rate of leaf development, however, flowering percentage for plants grown at higher temperatures before cold treatment decreased. It was expected that the

102 seedlings grown at the higher temperatures would have a higher flowering

percentage because they had more leaves before cold treatment. However, it

seems that the higher temperatures caused deleterious effects on subsequent

flowering. It is possible that soil temperature increased sufficiently to impede root growth. Because the differences in leaf unfolding rates/day are so small in

relation to temperature for each cultivars studied, it is advantageous to grow

plugs at 20 °C as opposed to temperatures _>_ 23 °C to prevent deleterious effects on flowering. "I

The purpose of the second phase of the experiment was to accurately ll.‘

determine the specific leaf number required by each cultivar for flowering.

However, leaf number alone could not be used to predict maturity. Plant size affected flowering percentage as well as leaf number. Chilling plants that were allowed to grow and establish in pots before cooling increased flowering percentage in comparison to plug-cooled plants, regardless of leaf number.

Results from the present study suggest that root growth, in addition to leaf development, may be an important factor in reaching maturity for Aquilegia

plants. When root growth is restricted in small containers, it is possible that

plants are unable to store enough nutrients for subsequent flowering even if the vemalization requirement has been met. The results correspond to those

reported in chapter 2. No studies were found in the literature which provided

similar information on Aquilegia.

The Aquilegia cultivars studied flowered as day-neutral plants, but flowered

slightly faster under long days, Leaf petioles elongated excessively under NI

103 compared to those of plants grown under SD, and total plant height at flowering was also greater for plants grown under long days. Because the difference in time-to-flower in relation to photoperiod is small, growing plants under natural days or SD after cold treatment is recommended to avoid undesirable elongation and height.

The ability to cool small plugs of Aquilegia, as opposed to overwintering plants in large pots, is an attractive alternative for many commercial producers.

For crops with a long juvenile phase, such as Aquilegia, commercial producers increase vegetative growth (bulking) of the plants before they administer cold treatments. The ability to cool seedlings in a controlled-environment chamber would allow the producer to provide more accurate and constant cooling and to cool plants at different times of year for staggered production schedules.

Production time and labor could be reduced, and space utilization maximized if plants could be cooled in small plugs instead of large containers. However, based on the results of the present study and those presented in chapter 2, it is difficult to achieve proper plant size when seedlings are grown and cooled in the plug tray. Container volume in addition to container depth must be considered when forcing Aquilegia. Aquilegia plants need to be grown in containers that do not restrict root growth prior to cold treatment. Growing and forcing recommendations for ‘Cameo’. ‘Bluejay’, ‘Crimson Star’, ‘McKana’s Giant’, and

‘Musik-White’ are listed in Table 11.

104 Literature cited

Bailey Hortorium Staff, Hortus Third. Macmillan. New York. 1976.

Faust, J.E. and RD. Heins. 1993. Modeling leaf development of the African violet (Saintpaulia ionantha Wendl.). J. Amer. Soc. Hort. Sci. 118:747-751.

Friend, D.J.C., V.A. Helson, and J.E. Fisher. 1962. Leaf growth in marquis wheat as regulated by temperature, light intensity, and daylength. Can. J. Bot. 40:1299-1311.

Karlsson, MG. 1992. Leaf unfolding in Begonia xhiemalis. HortScience. 27:109- 1 10. :-

J

.r Shedron, K. G., and T. C. Weiler. 1982. Regulation of growth and flowering in Aquilegia xhybn'da Sims. J. Amer. Soc. Hort. Sci. 107: 878-882.

Eff—FTOLLA

Thomas, B. and D. Vince-Prue. 1984. Juvenility, photoperiodism and vemalization, p. 408-439. In: Advanced Plant Physiology. Pitman Publ., London.

Tollenaar, M., T.B. Daynard, and RB. Hunter. 1979. Effect of temperature on rate of leaf appearance and flowering date in maize. Crop Sci. 19:363-366.

Vince-Prue, D. 1975. Vemalization, p. 262-291. In: Photoperiodism in Plants. Maidenhead, McGraw-Hill.

Weiler, T. C., and K.G. Shedron. 1986. Aquilegia xhybrida, p. 18-21. In: A.H. Halevy (ed.). CRC Handbook of Flowering. Vol. V. CRC Press, Boca Raton, Florida.

White, J W., D.J. Beattie, and E. J. Holcomb. 1989. Flowering studies with aquilegia cultivars. Acta Hort. 252:219-226.

White, J.W., J. Chen, and D.J. Beattie. 1990a. Gibberellin, light, and low- temperature effects on flowering of Aquilegia. HortScience.25:1422-1424.

White, J.W., H. Chen, X. Zhang, D.J. Beattie, and H. Grossman. 1990b. Floral initiation and development in Aquilegia. HortScience 25: 294-296.

Zhang, X., J.W. White, and D.J. Beattie. 1991. Regulation of flowering in Aquilegia. J. Amer. Soc. Hort. Sci. 1 162792-79

105 Table 1. All seedlings were four weeks of age when they were received at MSU on October 13, 1997. 128 Seedlings of each cultivar were placed at 17, 20, 23, and 26 °C under 16-h photoperiods with HPS lamps. Half of the seedlings from each cultivar were moved directly into a 5 °C cooler on the dates listed below and cooled in the plug tray. The treatments for the remaining seedlings are listed in Table 2.

All plugs were received on 10/13l97 at four weeks of age. Plugs were placed under each Average greenhouse air temperature temperature on 10/15/97. during plug growth (°C)

Weeks grown Temperature

under each Total age when Date moved to o o o 0 temperature moved to cooler cooler 17 C 20 C 23 C 26 C

8 12 12/8l98 17.3 20.3 23.2 26.4 g 12 16 1/5/98 17.5 20.4 22.9 26.5 I" 16 20 2/2/98 17.7 20.8 23.4 26.1

Table 2. Average greenhouse temperatures for all cultivars during forcing, after cold treatment.

Average greenhouse temperature Total age when moved to (°C)

greenhouse after cold under Date moved to o 0 SD or NI at SD or Ni 20 C 20 C

12 3/8/98 20.5 20.6 16 4/6/98 21.1 21.2 20 5/3/98 21.3 21 ,3

Total age when moved Average greenhouse temperature greenhouse after cold to D t edt (°C) 16-hour photoperiod at a 8:23: ° 20 cc 20 CC photoperiod SD

12 5/7/98 21.1 16 6/6/98 22.2 20 7l4/98 23.4

106 Table 3. Leaf unfolding rates/day at each growing temperature before cold treatment.

Temperature (°C) Leaf unfolding rate lday r2

A. xhybrida ‘Bluejay’ 17 0.108 0.99 20 0.122 0.99 23 0.138 0.99 26 0.135 0.99 A. flabellata ‘Cameo’ 1 7 0.068 0.98 20 0.100 0.97 23 0.122 0.98 t" 26 0.135 0.99 5 A. . xhybn'da ‘Crimson Star’ g 17 0.121 0.99 13;, 20 0.125 0.99 23 0.125 0.99 26 0.105 0.97 A. xhybrida ‘McKana’s Giant’ 17 0.100 0.98 20 0.104 0.99 23 0.114 0.99 26 0.125 1.00 A. xhybrida ‘Musik-White’ 17 0.108 0.98 20 0.127 0.99 23 0.124 0.98 26 0.138 0.99

107 FIIIIE.I.IIL...

Table 4. Plants of Aquilegia xhybn'da ‘Bluejay’ were cooled in either pots (20, 24, and 28 weeks old) or in plug trays (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower.

108 Table 4. Photo- period 'before Plug growing Final cold temperature Initial Days to Days from Days plant “after Age before cold Percentage leaf visible visible bud to Flower height cold wks ( ° C) flowering number bud to flower flower number (cm)

Pots *9 20 17 100 13.2 15 15 30 22 41 '9 20 20 100 14.1 20 16 36 22 40 *9 20 23 100 14.9 21 17 38 18 36 ‘9 20 26 90 14.7 27 1 5 42 23 39 Ni 20 17 70 15.1 18 15 33 21 42 *Nl 20 20 100 16.1 17 15 32 25 47 *Nl 20 23 100 17.2 23 17 40 23 41 *Nl 20 26 100 18.8 21 16 37 26 47 ‘9 24 17 100 16.3 20 18 38 27 39 *9 24 20 100 16.5 20 18 38 26 43 ‘9 24 23 100 16.9 19 17 36 21 36 0 24 26 100 16.8 15 18 33 26 40 'Nl 24 17 100 18.3 20 16 36 24 45 ‘Nl 24 20 100 18.6 19 17 36 25 44 Ni 24 23 100 20.6 20 15 35 24 45 "Nl 24 26 100 21.2 19 17 36 28 38 *9 28 17 100 19.7 14 11 25 13 34 *9 28 20 100 20.9 14 14 28 19 36 '9 28 23 100 22 12 16 28 16 41 *9 28 26 100 23.8 1 1 15 26 20 43 ‘Nl 28 17 100 22.9 17 15 32 16 35 'Nl 28 20 100 23.1 18 14 32 18 41 ‘Nl 28 23 100 24 19 17 36 18 39 Ni 28 26 100 25.1 17 15 32 14 44 Plugs “NI 12 17 0 7.1 “NI 12 20 0 7.3 “NI 12 23 0 7.3 ”NI 1 2 26 0 8.2 . . . . . “9 16 1 7 30 10 32 17 49 8 29 “9 16 20 50 1 1.7 33 16 49 7 31 **9 16 23 50 12.2 35 16 51 8 28 **9 16 26 40 12.6 33 14 47 1 1 30 "Nl 16 17 60 10.1 31 18 49 9 32 "Nl 16 20 40 10.8 29 16 45 10 31 "Nl 16 23 80 12.6 30 16 46 14 31 “NI 16 26 60 12.9 31 19 49 9 24 “9 20 1 7 20 12.8 33 13 45 9 29 "9 20 20 60 1 1.6 31 13 44 9 30 “9 20 23 50 16.1 28 16 44 8 31 “9 20 26 40 16.1 30 14 44 9 27 “NI 20 17 50 13.2 32 14 46 10 27 “N1 20 20 20 13.4 50 14 64 1 1 30 “Ni 20 23 60 16.6 30 15 45 1 1 27 \ “3L 20 26 50 16.6 34 16 50 12 37

109 Table 4 (con’t) Days from Final Initial Days to visible plant leaf visible bud to Days to Flower height number bud flower flower number (cm)

Signlflcance Pot or Plug (Potplug) '“ “' NS “' *** "" Age m m m m m ..

Potplug x Age ------Photoperiod *** NS NS NS NS “' Potplug x Photoperiod *** “ NS NS NS ' Age x Photoperiod *** NS ' *“ NS NS Potplug x Age x Photoperiod ------Temperature ““ NS NS ** * * Potplug x Temperature ' ' *“ NS NS * Age x Temperature m m m m NS m

Potplug x Age x Temperature ------Photoperiod x Temperature “‘ NS NS NS NS NS Potplug x Photoperiod x Temperature ‘ " ‘“ NS NS " Age x Photoperiod x Temperature ’“ ' NS NS NS ‘ Potplug x Age x Photoperiod x Temperature ------Contrasts 9 hours vs. 4-hr Night Interruption (NI) NS NS NS " NS " Age: 12, 16, 20, ‘20, 24, 28 wks P mm m m NS m m m P (“mm . .. NS m m m

Temperature: 17C, 20C, 230, and 26C P Um, NS NS NS NS NS “ P quadm NS NS " NS NS NS

W8, ', “, '" Nonsignificant or significant at P5005, 0.01, or 0.001, respectively

110

Table 5. Plants of Aquilegia flabellata ‘Cameo’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower.

111 Table 5.

Photo- period Plug growing Days 'before temperature from Final cold before cold Initial Days to visible plant **after Age (° C) Percentage leaf visible bud to Days to Flower height cold (wks) flowering number bud flower flower number (cm)

Pots *9 20 17 100 11.9 16 13 29 9 12 *9 20 20 100 12.4 16 6 29 8 12 *9 20 23 100 13.3 18 8 32 8 11 *9 20 26 100 12.1 9 9 23 11 13 WI 20 17 100 12.5 18 10 28 10 10 *Nl 20 20 100 15.0 13 12 25 13 11 *Nl 20 23 100 16.5 18 9 27 10 11 WI 20 26 100 17.6 18 8 26 10 11 *9 24 17 100 14.7 13 5 18 6 11 *9 24 20 100 16.4 14 5 19 6 11 *9 24 23 100 19.0 13 9 23 6 12 *9 24 26 100 22.2 12 8 20 8 14 *Nl 24 17 100 12.5 21 5 30 8 13 *Nl 24 20 100 13.6 17 9 26 8 12 e. Kw-__ *Nl 24 23 100 12.9 17 9 26 8 12 ‘4 *Nl 24 26 100 11.8 19 10 29 10 14 ‘9 28 17 100 14.1 19 5 24 14 12 *9 28 20 100 15.0 20 7 27 14 15 *9 28 23 100 17.6 22 9 31 12 13 '9 28 26 100 19.2 23 11 34 12 12 *Nl 28 17 100 16.3 22 8 30 8 13 *Nl 28 20 100 18.2 21 8 29 9 10 *Nl 28 23 100 20.3 23 10 33 7 13 Ni 28 26 100 23.7 22 8 30 8 14 Plugs “M 12 17 100 4.7 28 10 37 6 9 “M 12 20 100 6.9 22 10 32 7 9 “MI 12 23 100 8.0 27 10 37 7 7 “MI 12 26 100 6.5 29 16 45 5 7 **9 16 17 100 7.2 23 14 37 5 8 **9 16 20 100 9.5 23 14 36 6 10 “9 16 23 100 11.1 22 14 36 4 8 “9 16 26 100 14.6 22 14 36 6 8 “MI 16 17 100 7.7 19 14 33 7 9 “Ni 16 20 100 8.7 22 11 32 7 10 “Ni 16 23 100 11.9 17 13 30 6 8 “M 16 26 100 12.6 18 14 32 6 8 **9 20 17 100 10.1 18 15 33 6 8 **9 20 20 100 12.3 18 14 24 7 9 **9 20 23 100 14.6 19 15 34 8 9 **9 20 26 100 15.8 18 15 35 7 8 “M 20 17 100 9.4 15 16 31 7 10 "MI 20 20 100 11.0 15 16 31 7 9 **NI 20 23 100 15 16 14 30 7 8 __ “NI 20 26 100 16.2 17 48 35 8 9—

112 Table 5 (con’t)

Days from Final Initial Days to visible plant leaf visible bud to Days to Flower height number bud flower flower number (cm)

Significance Pot or Plug (Potplug) **‘ NS NS *** *** *** Age m NS NS m m . Potplug x Age ------Photoperiod **“ NS NS * *** *** Potplug x Photoperiod *** NS NS NS NS * Age x Photoperiod ** NS NS * * * Potplug x Age x Photoperiod ------"‘7' Temperature *** NS NS *** NS NS E Potplug x Temperature *** NS NS NS *** *** 5:? Age x Temperature **' NS NS *** ** *** L — Potplug x Age x Temperature ------Photoperiod x Temperature NS NS NS NS NS * Potplug x Photoperiod x Temperature * NS NS NS NS NS Age x Photoperiod x Temperature ** NS NS NS NS NS Potplug x Age x Photoperiod x Temperature ------Contrasts 9 hours vs. 4-hr Night Interruption (NI) * NS NS NS NS * Age: 12, 16, 20, '20, 24, 28 wks P mm m NS NS m m m

P mm,” NS NS NS *** *** NS Temperature: 17C, 200, 23C, and 26C P L5,“, *** NS NS NS NS NS P Mm“ NS NS NS NS NS NS

NS, ', **, *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively

113

Table 6. Plants of Aquilegia xhybrida ‘Crimson Star’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower.

114 Table 6.

Photo- Plug growing Days period temperature from Final *before cold before cold Initial Days to visible Days plant **after cold Age (° C) Percentage leaf visible bud to to Flower height wks flowering number bud flower flower number (cm)

Pots *9 20 17 60 12.8 31 16 47 26 39 *9 20 20 90 14.6 19 25 44 32 35 *9 20 23 70 14 31 14 45 20 37 *9 20 26 90 15 30 16 46 22 40 *Nl 20 17 50 13.1 35 15 50 17 41 “Ni 20 20 80 14.9 38 15 53 22 42 *Nl 20 23 80 14.8 37 15 53 24 39 *Nl 20 26 100 15.1 36 15 51 23 41 *9 24 17 100 16.9 27 13 39 30 31 *9 24 20 100 17.7 26 17 43 28 39 *9 24 23 100 17.8 24 17 46 23 35 *9 24 26 100 18.2 28 20 41 26 36 *Nl 24 17 100 18.2 30 18 48 26 46 *Nl 24 20 100 19.1 31 17 48 27 41 *Nl 24 23 100 18.5 29 18 46 24 43 *Nl 24 26 100 20.8 31 16 46 29 42 *9 28 17 100 20.9 19 16 35 22 40 *9 28 20 100 20.5 20 15 35 23 40 *9 28 23 100 20.8 19 13 32 24 45 *9 28 26 100 21.6 19 16 35 20 44 *Nl 28 17 100 22 20 15 34 19 43 *Nl 28 20 100 22.6 18 16 34 23 44 Ni 28 23 100 22.9 19 15 34 22 46 *Nl 28 26 100 23.9 19 16 36 21 43 Plugs “NI 12 17 50 8.1 40 18 58 8 17 "Ni 12 20 40 8.6 30 16 45 24 31 “M 12 23 0 8.9 - - - - - ”Ni 12 26 0 9.3 - - - - - **9 16 17 30 12.1 43 16 59 13 27 **9 16 20 40 12.4 41 16 57 17 29 **9 16 23 30 12.7 42 16 58 16 30 ”9 16 26 20 14.4 40 16 56 18 31 "Ni 16 17 40 13.2 33 16 49 17 39 “Ni 16 20 40 12.4 34 16 50 17 43 “M 16 23 40 12.4 33 16 49 17 42 ”Ni 16 26 40 12.8 34 17 51 17 40 **9 20 1 7 40 14.5 34 18 42 20 29 **9 20 20 40 15.4 37 18 54 22 32 **9 20 23 30 15.7 36 15 51 23 30 **9 20 26 0 16 - - - - - "M 20 17 30 14.3 34 18 52 20 39 “Ni 20 20 20 15.5 34 17 51 20 37 "N1 20 23 30 15.3 32 16 48 26 44 ‘ ml] 20 26 0 16.1 - - - - -

115 Table 6 (con’t).

Days from Final Initial Days to visible plant leaf visible bud to Days to Flower height number bud flower flower number (cm)

Signlflcance Pot or Plug (Potplug) m m . u m m Age m m NS m m m

Potplug x Age ------Photoperiod ‘* NS NS NS NS *** Potplug x Photoperiod *** NS * NS NS *** Age x Photoperiod *** NS NS NS NS ***

Potplug x Age x Photoperiod ------j Temperature *** NS NS NS * ***

Potplug x Temperature * ** *** ** NS NS 3‘“ l Age x Temperature NS *** NS *** NS NS IL'J' Potplug x Age x Temperature ------Photoperiod x Temperature NS NS NS NS NS NS Potplug x Photoperiod x Temperature ** NS *** NS NS NS Age x Photoperiod x Temperature NS NS NS NS NS NS Potplug x Age x Photoperiod x Temperature ------Contrasts 9 hours vs. 4-hr Night Interruption (NI) * NS NS NS NS ** Age: 12, 16,20, 20‘, 24, 28 wks P Um! m m NS m m m P Wm N S m NS m .. .4

Temperature: 17C, 20C, 23C, and 26C P Lm, NS NS NS * NS NS P Mam NS NS NS NS NS **

NS, *, **, *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively ' 20 week old pots

116

Table 7. Plants of Aquilegia xhybrida ‘McKana’s Giant' were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower.

117 ‘ Table 7.

Photo- Plug growing Days period temperature from Final *before before cold Initial Days to visible Days plant cold Age (° C) Percentage leaf visible bud to to Flower height “after cold wks flowering number bud flower flower number (cm)

Pots *9 20 17 90 14.3 31 16 47 26 38 *9 20 20 100 14.2 29 15 44 24 44 *9 20 23 100 13.9 30 16 46 22 41 *9 20 26 100 14.7 34 15 49 24 39 *Nl 20 17 100 15.0 37 15 52 26 45 *Nl 20 20 100 14.9 36 15 51 30 39 *Nl 20 23 100 14.9 26 14 50 24 39 *Nl 20 26 100 14.3 33 18 51 26 40 *9 24 17 100 18.8 29 17 46 28 39 '9 24 20 100 17.4 32 13 45 20 34 "9 24 23 100 17.8 31 14 45 25 41 *9 24 26 100 18.8 33 16 49 21 42 'Nl 24 17 100 19.8 34 14 48 19 40 *Nl 24 20 100 19.0 30 20 50 20 43 *Nl 24 23 100 18.4 27 19 46 19 42 *Nl 24 26 100 19.8 30 19 49 24 35 *9 28 17 100 21 .4 22 12 34 24 39 *9 28 20 100 19.5 20 1 5 35 22 38 *9 28 23 100 20.7 22 15 37 21 43 *9 28 26 100 21.4 20 15 35 19 35 *Nl 28 17 100 22.1 23 16 39 20 39 *Nl 28 20 100 19.9 22 17 39 20 45 ‘Nl 28 23 100 21.3 23 17 40 21 47 *Nl 28 26 100 22.1 23 16 39 18 44 Plugs “MI 12 17 50 9.6 26 24 50 12 30 "M 12 20 20 7.2 33 18 51 11 27 “M 12 23 O 7.3 "MI 12 26 0 7.9 . . . . . **9 16 17 30 10.9 45 17 62 19 32 **9 16 20 30 10.3 44 17 61 26 30 **9 16 23 40 10.9 45 18 62 12 22 **9 16 26 30 1 1.3 45 16 61 18 26 “MI 16 17 30 10.4 46 19 65 16 26 “M 16 20 40 10.2 50 16 65 17 40 ”M 16 23 10 11.1 51 16 67 16 45 “Ni 16 26 0 1 1 .3 . . . . . **9 20 17 60 14.1 36 17 52 13 34 **9 20 20 40 13.3 35 17 52 13 32 **9 20 23 30 12.1 36 16 52 12 31 **9 20 26 30 14.7 36 17 43 13 29 **Nl 20 17 50 13.5 31 17 48 15 49 “Ni 20 20 30 13.3 32 16 48 17 47 "MI 20 23 20 13.4 31 16 47 16 50 _ “MI 20 26 40 15-8 30 1 6 46 1 9 59

118 Table 7 (can’t)

Days from Final Initial Days to visible plant leaf visible bud to Days to Flower height number bud flower flower number (cm)

Significance Pot or Plug (Potplug) *** * NS * *** *** Age 1... . m . m m

Potplug x Age - - - - - Photoperiod *** NS ** NS NS '** Potplug x Photoperiod * NS NS NS NS *** Age x Photoperiod NS NS NS NS * * ad: Potplug x Age x Photoperiod ------.' Temperature *** NS NS NS NS NS _“L‘

Potplug x Temperature NS *** ** *** NS NS 81.11.“ K’v Age x Temperature *** ** NS ** NS NS I Potplug x Age x Temperature ------Photoperiod x Temperature NS NS NS NS NS NS Potplug x Photoperiod x Temperature NS NS * NS NS NS Age x Photoperiod x Temperature NS NS NS NS NS ** Potplug x Age x Photoperiod x Temperature ------Contrasts 9 hours vs. 4-hr Night Interruption (NI) * NS NS NS NS *** Age: 12, 16, 20, *20, 24, 28 wks P mm 1... NS .1... NS 1... m P Mam N 8 NS 1... NS 1.. ..

Temperature: 17C, 200, 23C, and 260 P Lm, NS NS NS NS NS NS P mam * NS NS NS NS NS

NS, ', **, *‘* Nonsignificant or significant at P5005, 0.01, or 0.001, respectively

119

‘ Table 8. Plants of Aquilegia xhybrida ‘Musik White’ were cooled as either pots (20, 24, and 28 weeks old) or as plugs (12, 16, and 20 weeks old). Initial leaf counts for each plug were taken before they were placed into the 5°C cooler. Flower number and final plant height were measured at the time of first open flower.

120 Table 8.

Photo- period Plug growing Days *before temperature from Final cold before cold Initial Days to visible plant **after Age (° C) Percentage leaf visible bud to Days to Flower height cold wks flowering number bud flower flower number (cm)

Pots *9 20 17 90 14.1 34 15 49 29 36 *9 20 20 100 15.8 29 15 44 33 35 *9 20 23 100 16.2 29 16 45 24 35 *9 20 26 100 16.6 31 16 47 31 39 *Nl 20 17 100 15.9 37 17 54 28 36 *Nl 20 20 100 16.4 37 17 53 26 34 *Nl 20 23 100 17.1 37 17 54 21 34 *Nl 20 26 100 17.9 36 18 52 26 38 *9 24 1 7 100 16.9 29 15 44 20 29 *9 24 20 100 18.5 18 17 35 22 31 ~41 *9 24 23 100 19.6 28 18 46 22 32 *9 24 26 100 20.5 22 19 41 28 35 *Nl 24 17 100 18.6 30 19 49 19 36

*Nl 24 20 100 19.3 30 17 47 22 38 ['3' *Nl 24 23 100 20.6 30 16 46 26 42 *Nl 24 26 100 22.3 28 14 42 23 37 *9 28 1 7 100 20.2 20 18 38 22 32 *9 28 20 100 21.5 20 18 38 21 34 *9 28 23 100 21.5 20 17 37 24 37 *9 28 26 100 23 21 18 39 20 39 *Nl 28 17 100 21 21 16 37 26 45 *Nl 28 20 100 22.5 20 17 37 23 40 *Nl 28 23 100 22.5 20 14 34 21 43 *Nl 28 26 100 24.4 20 17 37 19 40 Plugs "M 12 17 0 7.3 . . . . . "M 12 20 20 7.2 32 14 46 13 31 “Ni 12 23 0 8.9 “MI 12 26 0 9.7 . . . . . **9 16 17 20 11.8 36 17 52 10 28 **9 16 20 20 12.4 35 16 51 12 30 **9 16 23 0 12.5 **9 16 26 0 13.1 “MI 16 17 0 1 1 .9 “M 16 20 0 12.3 "MI 16 23 0 12.7 “M 16 26 0 13.1 . . . . . **9 20 17 30 13.9 35 20 55 25 34 **9 20 20 30 14.5 36 19 55 22 29 **9 20 23 0 15.2 **9 20 26 0 15.7 . . . . . “M 20 17 50 14.4 37 13 51 22 43 "M 20 20 40 16.3 36 14 40 22 42 “Ni 20 23 0 15.4 “MI 20 26 0 1 7-4 - -

121 Table 8 (con’t).

Days from Final Initial Days to visible plant leaf visible bud to Days to Flower height number bud flower flower number (cm)

Significance Pot or Plug (Potplug) *** *** ** *** ** NS Age .1... m . m . m

Potplug x Age ------Photoperiod m m m .. NS m

Potplug x Photoperiod NS NS *** ** NS * Age x Photoperiod m m . 1... NS m

Potplug x Age x Photoperiod ------Temperature *** * * * NS NS

Potplug x Temperature *** NS NS NS NS NS _..'.lL....“-.a‘ IL: Age x Temperature *** ** * NS ** NS Potplug x Age x Temperature ------Photoperiod x Temperature NS *** NS * NS NS Potplug x Photoperiod x Temperature NS NS NS * NS NS Age x Photoperiod x Temperature ** *** NS NS NS NS Potplug x Age x Photoperiod x Temperature ------Contrasts 9 hours vs. 4-hr Night Interruption (NI) NS *** *** NS *** ** Age: 12, 16, 20, *20, 24, 28 wks P Um, *** NS NS NS * * P mm.” NS ** NS NS NS NS Temperature: 170, 200, 230, and 26C P W, ** NS NS NS NS NS P M6,,“ NS NS NS NS NS NS

NS, *, **, *** Nonsignificant or significant at P5005, 0.01, or 0.001, respectively

122 Table 9. The overall average leaf number of both the short-day and NI treatments combined at each growing temperature before cold of each species studied. Also, the overall average leaf number of pot- and plug-cooled plants combined for each growing temperature.

Temperature (° C) Pot-cooled plants Plug-cooled plants Total average leaf number

A. xhybrida ’Bluejay’

17 17.6 10.8 14.2 20 18.2 10.6 14.4 23 19.3 13.2 16.2 26 20.1 13.5 16.8 A. flabellata ‘Cameo’ 17 13.7 8.0 10.8 20 15.1 9.9 12.5 23 16.6 12.2 14.4 “*1 26 17.8 13.5 15.6 A. . xhybrida ’Crimson Star' __.

I?" 17 17.5 12.6 15.0 20 18.1 13.0 15.6 23 18.3 13.2 15.8 26 19.3 14.0 16.6 A. xhybrida ‘McKana's Giant' 17 17.2 10.6 13.9 20 17.9 11.2 14.5 23 18.4 11.7 15.1 26 19.4 12.8 16.1 A. xhybn'da ‘Musik-White’ 17 17.8 12.0 14.9 20 19.0 12.7 15.8 23 19.6 13.1 16.3 26 20.8 13.9 17.3

123 Table 10. The overall combined average leaf numbers at the ages each species studied was placed into the cooler.

Age (weeks) Pot-cooled plants Plug-cooled plants Age (weeks)

A. xhybrida ‘Bluejay' 20 15.5 7.5 12 24 18.2 11.6 16 28 22.7 14.3 20 A. flabellata ‘Cameo’ 20 13.9 6.5 12 24 15.4 10.4 16 28 18.1 13.1 20 A. . xhybrida ‘Crimson Star' 20 14.4 8.7 12 24 18.5 12.8 16 y 28 21.9 15.4 20 A. xhybrida ‘McKana’s Giant’ l’ 20 14.3 7.8 12 L 24 18.6 10.8 16 28 21.7 14.0 20 A. xhybrida ‘Musik-White’ 20 16.3 8.7 12 24 19.5 12.5 16 28 22.1 15.4 20

124 Table 11. The requirements for flowering of five cultivars of Aquilegia. The following recommendations apply to plants grown in 1.1-L or larger containers prior to cold treatment to achieve 100% flowering. Cold duration recommendations are taken from chapter 2. Force plants at temperatures between 14 and 20 °C (chapter 3).

Species Leaf # Weeks of Weeks required growing Photoperiod of cold Photoperiod for before before before cold at forcing cold cold 5 °C

Aquilegia z 5 5-6 Natural or SD 9 weeks Natural or SD flabellata at 17-20 with supple- with supple- ‘Cameo’ °C mental light mental light F Aquilegia 313 13-14 Natural or SD 9 weeks Natural or SD , xhybrida at 17-20 with supple- with supple- ,‘ ‘Bluejay’ °C mental light mental light L A. xhybrida 210 10-11 Natural or SD 3 to 6 Natural or SD ‘Crimson at 17-20 with supple- weeks with supple- Star’ °C mental light mental light A. xhybrida 310 10-11 Natural or SD > 12 Natural or SD ‘McKana’s at 17-20 with supple— weeks with supple- Giant’ °C mental light mental light

A. xhybrida 214 14-15 Natural or SD > 12 Natural or SD ‘Musik- at 17-20 with supple- weeks with supple- White’ °C mental light mental light

125 20 'Bluejay’ 'Cameo'

15-

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number Leaf

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10- =17°C number l=20°C Leaf A=23°C v=26°C

468101214161820 Plant Age (weeks)

Figure 1. The effects of temperature on leaf development of Aquilegia xhybrida 'Bluejay', A. flabellata ‘Cameo', A. xhybrida 'Crimson Star', A. xhybrida 'McKana's Giant’, and A. xhybrida 'Musik-White'. The points indicate the actual number of leaves unfolded at each age. The straight line was calculated from linear regression analysis.

126

Figure 2. The effects of initial leaf number, temperature before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on plants of ‘Bluejay’, ‘Cameo’. ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik—White’. Seedlings were grown at 17, 20, 23, and 26 °C before cold. (0) represents plants grown under SD, (a) represents plants grown under NI.

127

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128

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[Figure 2, (con't.)

129

Figure 3. The effects of initial leaf number before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on flowering percentage of ‘Bluejay’, ‘Cameo’. ‘Crimson Star', ‘McKana’s Giant’, and ‘Musik—White’. (0) represents plants grown under SD, (41) represents plants grown under NI.

130

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131

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mi Figure 4. The effects of temperature before cold treatment, and photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on days to visible bud for plants of ‘Bluejay’, ‘Cameo'. ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik- White’. Seedlings were grown at 17, 20, 23, and 26 °C before cold. (0) represents plants grown under SD, (A) represents plants grown under NI.

132

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134

Figure 5. The effects of temperature before cold treatment, and overall age of pot— and plug-cooled plants, on the days to visible bud for ‘Bluejay’, ‘Cameo’, ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik—White'. (A) = 17 °C, (0) = 20 °C, (X) = 23 °C, and (D) = 26 °C.

135

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_er-o Figure 6. The effects of average leaf number before cold treatment, and I E photoperiod before cold for “Pots”, photoperiod after cold for “Plugs”, on days to visible bud, days from visible bud to flower, and days to flower for plants of ‘Bluejay’, ‘Cameo’, ‘Crimson Star', ‘McKana’s Giant’, and ‘Musik—White’. (0) represents days to visible bud, (a) represents days to flower.

137

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r———__ Figure 6.

138

Figure 7. The effects of plant age in weeks on the flower number of “Pots” and “Plugs” of ‘Bluejay', ‘Cameo’, ‘Crimson Star’, ‘McKana’s Giant’, and ‘Musik- White’. Data points represent total average of plants grown under both SD and NI lighting at each age.

139 Flower Number- Pots Flower Number- Plugs

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141

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142

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143

CHAPTER V. THE EFFECTS OF PHOTOPERIOD, COLD TREATMENT, AND FORCING TEMPERATURE ON THE GROWTH AND FLOWERING OF GAURA LINDHEIMERI ‘WHIRLING BUTTERFLIES’.

144

fill. .. . Introduction

The genus Gaura, a member of the Onagraceae (Evening Primrose) family, includes about eighteen native American species of annual, biennial and perennial plants (Bailey, 1976). All eighteen species are native to Texas,

Louisiana, and Mexico. G. Iindheimeri Engelm & A. Gray ‘Whirling Butterflies’ is found growing in climates ranging from USDA zones 5 to 10, and is versatile, and resilient. Gaura usually performs best in hot, dry, full sun locations, and interest __l in the species has increased along with the popularity of low-maintenance "T

-

. gardening and water conservation.

I

1."?-

Most Gaura species are tall and bushy and range in height from 3 to 4 feet. A

They are free-flowering plants that bloom for most of the growing season. The foliage may turn a deep purple color as temperatures decrease in the fall and winter. Although individual plants do not spread rapidly, Gaura self-sows seed freely. There is also a pink-flowered cultivar of Gaura Iindheimeri called “Siskiyou

Pink”. The foliage is dark green with pink veins and has a mottled appearance.

This cultivar originated at the Siskiyou Rare Plant Nursery in Oregon as a chance seedling.

Plant response to environmental stimuli, such as temperature and photoperiod, depends upon the species and genotype of the plant. Many perennial plants require a period of cold, or vemalization, for optimal flower development (Jones, 1992). Vemalization has been defined as a cold treatment that promotes subsequent flowering when it is given to imbibed seeds, bulbs, or whole plants (Vince-Prue, 1975). For plants with an obligate requirement for

145 vemalization, flower buds will not form until the requirement for cold is fulfilled and the plants are returned to higher temperatures.

Cold temperatures promote flowering in many plants by breaking dormancy of existing buds when reproductive structures are present prior to the onset of low temperatures. Many woody perennials and flowering bulbs such as azaleas, hydrangeas, and tulips for example, form their flower buds in the summer or fall and remain dormant throughout the winter (Thomas and Vince- kw Prue, 1984). The floral primordia of Paeonia are initiated soon after anthesis of '1. the current year's flowers and require a minimum of four weeks at 5.6 °C to break r J dormancy (Byme and Hadley, 1986). Full reproductive development, or anthesis, will not occur without a subsequent cold treatment. This would be considered a breaking of dormancy, not vemalization (Thomas and Vince-Prue, 1984).

in some plants, such as Brassica oleracea (Brussels sprouts) and

Dicentra spectabilis, flower differentiation occurs during the cold, or cool treatment (Thomas, Vince-Prue, 1984). In Allium cepa (onion), and his cv.

Wedgewood, low temperatures are also required during floral initiation. in

Matthiola incana, low temperatures must continue until the buds are visible in order the development to continue upon return to higher temperatures (Kohl,

1957)

For some plants, a cold treatment will speed the process of flowering but is not necessary to initiate flowering. This is termed a facultative, or quantitative response to cold. The hastening of flowering can be measured directly by calculating time to flower and also has been correlated with a reduction of the

146 number of leaves formed before flowering in grains (Chouard, 1960). No

information was found in the literature that documented the response of Gaura

Iindheimeri ‘Whirling Butterflies’ to cold treatment.

The requirement for vemalization is commonly, but not only, found in long day plants (Thomas and Vince-Prue, 1997). Many plants that have a cold

requirement, such as Coreopsis ’Sunray’, also require subsequent treatment with

long days. In some cold-requiring plants, such as Campanula medium, short days may substitute for, enhance, or be required instead of cool temperatures in order to achieve anthesis (Roberts et al.,1988). The low temperature

requirement for flowering in Japanese winter wheat varieties Norinn 27 and 8 is

completely replaced by short-day treatments (Krekule, 1987). There was no

available information in the literature on the response of Gaura Iindheimeri

‘Whirling Butterflies’ to photoperiod.

The rates of many plant developmental processes, and the timing of

phenological stages, are strongly temperature dependent (Jones, 1992). Thus,

production time for any crop is related to temperatures supplied during forcing

(Whitman et al., 1996). Under long-day conditions, average daily temperature is

the primary factor influencing of flower development. Although increasing the

temperature to speed up the crop may seem desirable, negative consequences

may result from growing the crop too warm. Delayed flowering and reduced plant

vigor and flower size can result at high temperatures. At temperatures below a

species-specific minimum (Tb), the time to flower (f) is infinite (Roberts and

Summerfield, 1987). Temperatures above a ceiling value (Tom) delay flowering. 147 When developmental time is converted to a rate by taking the reciprocal, the relationship between mean temperature and rate of development is often linear in the range between Tb and Top, (Roberts and Summerfield, 1987). Thus, the relationship between the rate of development toward flowering (1/DTF, where

DTF is the days to flower) and temperature can be described as follows [1] 1/DTF=bO+b1*T

(Yuan et al., 1998). Using the constants b0 and b,, the base temperature, Tb. and T" degree-days (°days) can be calculated as follows: z _ T” = 'b°’b‘ ' [21 [3] [ °days = 1/b1

(Yuan et al., 1998). Base temperature is the temperature at, or below, which, the rate of progress toward flowering is zero, and degree-days represent the thermal time required for flowering (Yuan et al., 1998). The base and optimum temperatures vary within and between species and are related to climatic origin

(Roberts and Summerfield, 1987). The effects of temperature have been documented for a limited number of herbaceous perennials, such as Campanula carpatica “Blue Clips’, and Rudbeckia fulgida (Whitman et al., 1997, Yuan et al.,

1998), but no such available information was found for Gaura Iindheimeri

‘Whirling Butterflies’.

The objective of our study was to 1) measure the influence of cold treatment, photoperiod and forcing temperature on flower number, flower diameter, leaf number and plant height of Gaura Iindheimeri ‘Whirling Butterflies’

2) establish the optimum cold treatment duration, photoperiod, and forcing

148 temperature that induced the most complete and rapid flowering. Percentage of plants flowering, time to flower, flower quantity and size were evaluated.

Materials and Methods

General. Vegetative cuttings with an average of 7 to 10 nodes in 72-cell trays (SO-ml cell volume) were received from a commercial producer during the

first week of November 1996 and 1997. Plants were grown in a commercial ‘1 ' 1. soilless medium composed of composted pine bark, horticultural vermiculite, .21

Canadian sphagnum peat moss, processed bark ash, and washed sand [-1

(MetroMix 510, Scotts-Sierra Horticultural Products Company, Marysville, Ohio, year 1). Plants were top-watered with well water acidified (two parts H3PO4 plus one part H2804, which provided=2.5 mol P-m‘3) to a titratable alkalinity of approximately 130 mg calcium bicarbonate per liter and fertilized with 14N-0P-

6K20 (mol-m3) from potassium nitrate (14-OP-55K20) (Vicksburg Chemical Co.,

Vicksburg, MS) and ammonium nitrate (34N-0P-0K20) (Cargill, Lexington, KY).

Fertilization and acidification rates were adjusted in response to weekly soil test results, so regimens varied during experiments. Our target range for soil pH was

5.8 to 6.2 and, and 0.5 to 1.2 for soil EC.

Supplemental lighting from high-pressure sodium lamps provided a photosynthetic photon flux (PPF) of approximately 50 umol-m'z s‘1 at plant level when the ambient greenhouse PPF dropped to 200 umol-m'z-S'1 between 800 hr to 1700 hr daily. Supplemental lighting was terminated when PPF exceeded 400 umol-m‘2 s".

149 Plants were exposed to cold treatments in a 5 1 0.5 °C cooler illuminated for 9 h oday " with cool-white fluorescent lamps (VHOF96T12; Philips, Bloomfield,

N.J.), as measured by a LICOR quantum sensor (model Ll-189; Ll-COR, lnc.,

Lincoln, NE). While in the cooler, plants were watered two to three times a week as needed with well water acidified (H2804) to an approximate pH of 6.0.

Critical photoperiod (Expt. 1). Upon arrival, 7O seedlings were removed from the plug-trays and transplanted into 1.1-L (5" US.) square pots. Ten plants were placed under 10, 12, 13, 14, 16, and 24 hours of continuous light and 9 hours of natural light plus a 4-hour night interruption (Nl) from 2200 to 0200 HR.

Photoperiods were completed with incandescent lamps at 1 to 3 umol-m‘2 s". For the continual photoperiodic treatments, lamps provided day-extensions; they were turned on at 1700 HR and turned off after each photoperiod was completed.

Photoperiod treatments were assigned randomly to benches in the greenhouse.

The remaining 70 seedlings were placed in the cooler for 15 weeks. After the cold treatment, plants were transplanted into 1.1-L (5" US.) square pots. Ten plants were placed under each of seven previously mentioned photoperiod treatments. Dates of the first visible bud, and first open flower were recorded for each plant, and days from transplant to visible bud and flowering were calculated.

At first flower, total plant height, visible flower bud number, and node number on the main stem were recorded.

Greenhouse temperature was set at 20 °C. Air temperatures on each bench were monitored with 36-gauge (0.013-mm-diameter) type E thermocouples 150 connected to a CR10 datalogger (Campbell Scientific, Logan, UT). The datalogger collected temperature data every 10 seconds and recorded the hourly average. To provide uniform temperature conditions, the datalogger controlled at

1500-W electric heater under each bench that provided supplemental heat as

needed throughout the night. Actual average daily air temperatures from the

beginning of forcing to the average date of flowering were calculated for each species.

‘

Cold treatments (Expt 2). Ten plants were removed from the plug-tray and transplanted into 1.1-L (5" US.) pots, and were then placed in the greenhouse under 16 hours of extended day light from high-pressure sodium

lamps. The remaining fifty plants were placed in the cooler for 3, 6, 9, 12, and 15 weeks. After each cold duration treatment, ten plants were removed and transplanted into 1.1-L (5" US.) square pots, and placed in the greenhouse under

16 hours of extended day light from high-pressure sodium lamps. The

greenhouse was lit from 0700 to 0800 HR and from 1700 to 2300 HR at

approximately 50 umol-m'2 s‘1 as measured by a LICOR quantum sensor (model

Ll-189; LI-COR, lnc., Lincoln, NE) to provide the proper photoperiod. Data were recorded as described for Expt. 1.

Forcing temperature treatments (Expt. 3). Fifty plants were placed in a 5

°C cooler for 12 weeks, and then transplanted into 1.1-L (5" US.) square pots.

T9" Plants were placed in each of five greenhouses set to 17, 20, 23, 26, and 29

°C. Plants received natural day-lengths plus day-extension lighting from 0700 to

151

lam.-

0800 HR and from 1700 to 2300 HR at approximately 50 umol-m'Z-s'1 (HPS lamps) as measured by a LICOR quantum sensor (model Ll-189; Ll-COR, lnc.,

Lincoln, NE) to provide a 16-hour photoperiod.

Air temperatures on each bench were monitored with 36—gauge (0.013- mm—diameter) type E thermocouples connected to a CR10 datalogger (Campbell

Scientific, Logan, UT). The datalogger collected temperature data every 10

seconds and recorded the houdy average. The datalogger collected temperature at data every 10 seconds and recorded the hourly average. Actual average daily air temperatures from the beginning of forcing to the average date of flowering were

calculated for each species. u-‘m—w—p

Results and Discussion

Critical photoperiod (Expt. 1).

Gaura Iindheimeri responded as a facultative long-day plant both years it was studied. However, long days were nearly obligate both years Gaura was studied. For example, under the 10-hour photoperiod, without a cold treatment, only 10% of plants flowered during year 1 and no plants flowered during year 2

(Table 1, Figure 1). After 15 weeks of cold, 40-50% of plants flowered under the

10-hour photoperiod both years (Tables 1, Figure 1). Although some plants eventually flowered under the 10-hour photoperiod, and all plants flowered under the 12—hour photoperiod after cold treatment. However, the plants remained prostrate under these short photoperiods (Table 1, Figure 1). Under the 13-hour photoperiod, plant morphology was upright, as opposed to the plants grown under

152 the 10- and 12-hour photoperiods. All plants flowered under all other photoperiods with and without cold. Without a cold treatment, longer photoperiods significantly accelerated flowering, especially during year 1. For example, visible bud was accelerated by 16 days as the photoperiod increased from 10 to 24 hours of continuous light (Table 1, Figure 1). The difference in days to visible bud in relation to photoperiod was smaller during year 2, however, significant interactions were observed between photoperiod, weeks of cold, and the two separate years of data (Table 1).

Height at first flowers increased as photoperiod lengthened. Plants under the 24-hour photoperiod were about 70-cm tall, and required support, possibly due to a far-red light effect (Table 1, Figure 2). The number of flower buds per plant was not significantly different between photoperiod treatments at and above the 12-hour photoperiod (Table 1). Cold treatment, however, doubled the number of flower buds under all photoperiods (Table 1, Figure 2).

Cold treatments (Expt. 2).

Vemalization was not required for flowering of G. Iindheimeri. Without a

cold treatment, all plants flowered under 16 hours of light, as in experiment 1

(Tables 1 and 2, Figures 2 and 4). Days to visible bud was variable between the two years of data (Table 2). For example, during year 1, days to visible bud

increased slightly from 47 to 53 days, but decreased from 48 to 28 days during year 2 as the duration of cold treatment increased from 0 to 15 weeks (Table 2, Figure 4). 153 Flower number was significantly affected by cold treatment, but during year

1, data varied with treatment and no trends were observed. Year 2, flower number increased by 50% as the duration of cold treatment increased from O to

15 weeks (Table 2, Figure 4). Plant node development and plant height were also variable during year 1, but decreasing trends with increased cold were observed for both node number and height during year 2 (Table 2). Cold decreased plant node development prior to flowering by about three leaves and plant height decreased by 12 cm (Table 2, Figure 4).

Forcing Temperature (Expt.3).

Plants flowered faster at temperatures up to 26 °C, but heat-delay was observed at 29 °C. The time to flower after beginning LD for Gaura Iindheimeri

‘Whiding Butterflies” was dependent upon forcing temperature: about 7 weeks at

64°F (17°C), 6 weeks at 68°F (20°), or 5 weeks at 74°F (23°C). Days to visible bud decreased from 32 to 22 days as temperature increased to 26 °C, but was delay by six days at 29 °C (Table 3, Figure 5). Days from visible bud to flower decreased linearly from 19 to 13 days as temperature increased to 29 °C with no delay observed (Table 3). All plants flowered under all temperature treatments.

There were significant linear and quadratic relationships between temperature and rate of progress toward visible bud, days from visible bud to

flowering, and days to flower for G. Iindheimeri. The rate to visible bud and

flowering followed a statistically significant quadratic pattern, whereas days from visible bud to flower followed a linear pattern (Table 4). With a 10 °C rise in 154 temperature, there was only an approximate 30% decrease in time to flower. The base temperature and degree-days for days to visible bud, days from visible bud to flower, and days to flower were determined using equations [2] and [3]. Base temperature and degree-days can be used to predict the flowering date in commercial greenhouse conditions in which temperatures fluctuate (Yuan et al.,

1998). The base temperatures calculated for ’Whirling Butterflies’ were below zero for all events, ranging from -3.4 for days to visible bud to -11.8 for days to

flower, suggesting that flower buds may have formed while plants were still in the cooler (Table 4).

There was a decrease in flower diameter, measured across the open face of the flower, as the forcing temperature increased. Flowers at the highest temperature were smaller, although no gross flower deformities or doubling were observed at 29 °C. Flower diameter decreased linearly, by about 35%, with increasing temperature (Table 3, Figure 6). Reducing forcing temperature from

29 to 17 °C promoted a six-fold increase in bud number at first flower. Bud number at first flower followed a statistically significant quadratic pattern in relation to temperature.

Plants grown at 17 °C had an average of 77 flower buds at the time of first flower as opposed to plants grown at 29 °C, which had an average of only 12 flower buds (Table 3, Figure 6).

155 Plant node development and plant height decreased linearly as temperature increased. Final leaf number decreased by five nodes, and plant height decreased by 16 cm as temperature increased from 17 to 29 °C (Table 3,

Figure 6).

Conclusion

Vince-Prue defines the critical photoperiod of a species as the daylength at which 50% of the plants flower (Vince-Prue, 1975). However, Roberts and

Summerfield (1987) define critical photoperiod for short-day plants as the photoperiod at or below which the time to flower is minimal and is not affected by variations in day-length. Conversely, they define critical photoperiod for long—day plants as the photoperiod above which time to flowering is minimal and not affected by further increased in photoperiod, and below which flowering is delayed (Roberts and Summerfield, 1987). In commercial horticulture, however, it is necessary to have the ability to keep all plants vegetative, or to promote

flowering of all plants of a certain species as desired. Therefore, we have proposed an alternative definition of critical photoperiod; critical photoperiod may be the photoperiod at which 100% of plants flower, and at which the time to flower is minimal and is not affected by variations in day-length (Whitman et al., 1996a,

1996b, 1998). Under photoperiods of 12 hours or less, 100% of Campanula carpatica “Blue Clips’ remained vegetative (Whitman et al., 1996b). Under 14 hours, 60% of plants flowered, while all plants flowered under 16 hours of continuous light or a 4-hour night-interruption from 10 pm. to 2 am. (Whitman at 156 al., 1996b). It was concluded that the critical photoperiod for C. carpatica ‘Blue

Clips’ was 16 hours.

Percentage flowering for ‘Whirling Butterflies’ reached 100% for plants that were grown under photoperiods z 12 hours, both with and without a cold treatment. Thus, 12 hours would be considered the critical photoperiod for G.

Iindheimeri. Extended exposure to 5 °C increased the percentage flowering under the short-day photoperiods, but did not entirely eliminate the requirement 5 for longer photoperiods. In addition, the plants grown under 10-hr photoperiods fl were diminutive and prostrate in their growth habit and would not be acceptable for sale. When lighting Gaura with incandescent bulbs, photoperiods longer than 3

16 hours are not recommended due to excessive stem elongation. Plants given a cold treatment, then grown under 13 hours of light were consistently of higher quality compared to those grown under longer photoperiods when incandescent lighting was used. Plants were shorter and more compact that those grown under longer photoperiods, and had a higher flower number than plants grown under both shorter and longer photoperiods. We recommend a 13-hour continuous photoperiod when using incandescent bulbs.

Plants in experiment 2 were grown using 16 hours of light from high- pressure sodium lamps as opposed to incandescent bulbs in experiment 1. Thus, plants received a higher daily light integral in experiment 2, which resulted in plants with more lateral branching, more flower buds, and less undesirable stem elongation. Plants in experiment 2 had, in most cases, 50% more flower buds

157 than those grown with incandescent lighting in experiment 1 (Tables 1 and 2,

Figures 1 and 2).

The requirement for cold treatment differs among species and can often be categorized by the relationship between exposure to cold and time of floral initiation. Our experiments show that exposure to a period of low temperatures is not required for promoting flowering in Gaura Iindheimeri ‘Whirling Butterflies’.

One hundred percent flowering was achieved without a cold treatment when grown with 16 hours of high-intensity lighting. However, a cold treatment did promote increased lateral branching, and flower number, as well as the overall _J‘ plant appearance. 1

Although all plants of G. Iindheimeri flowered under all temperature treatments, high temperatures had a negative effect on flower number and size.

Similar responses to temperature have been reported for other species, such as tomato. In two varieties of tomato, heat-sensitive Pusa Ruby and heat-tolerant

CL-1311, many floral anomalies such as stigma exertion without anthesis, empty

flowers, and persistent flowers without fruit-set were observed in when grown under a 35 I 30 °C day/night temperature regimen (Lohar and Peat, 1998).

Abnormalities such as flower doubling occurred for plants of Campanula ‘Birch

Hybrid’ when grown at constant day night temperatures of 29 °C (Frane, unpublished data).

No flower abnormalities were observed in this study for plants of ‘Whirling

Butterflies’ grown at 29 °C, but flower size and vigor were both dramatically reduced.

158 Overall, plants grown a lower temperatures had more and larger flowers and were taller, but took longer to reach flowering. Plants grown at the highest temperature were too weak and diminutive in size for 1.1L (5" US.) pots, whereas the plants grown at the coolest temperature may have been too large for them.

Settings of 17 to 20°C are recommended to force Gaura Iindheimeri ‘Whirling

Butterflies’. Plants flowered slightly faster in the temperature range of 20 to 23 °C than did those grown at lower temperatures, but those grown at 17 °C were

highly attractive. These results for Gaura Iindheimeri are similar to the responses of Coreopsis grandiflora, Gail/ardia xgrandiflora, Leucanthemum xsuperbum, and,

Rudbeckia fulgida (Yuan et al., 1998), Levendula angustifolia ‘Munstead’ “Er-m-

(Whitman et al., 1996a), and several species of Aquilegia (chapter 2).

Based on time-to-flower and flower number, and overall plant

appearance, we recommend 6 to 9 weeks of cold at 5 °C for production of Gaura

Iindheimeri ‘Whirling Butterflies’. After the cold treatment, provide long-days, which can be delivered by natural or extended photoperiods z 13 hours with a

minimum light intensity of 1 to 3 umol-m'z’s‘1 (10 footcandles). Light from high-

pressure sodium lamps is preferred over incandescent. Provide plants with

supplemental lighting from HPS lamps during dark winter climates. Supplemental

lighting at approximately 50 umol-m'z-S'1 (500 footcandles) will increased flower

number, thus plant quality. Force plants at temperatures between 17 and 20 °C (64 and 74 °F).

159 Literature Cited

Bailey Hortorium Staff. 1976. Hortus Third. Macmillan. New York.

Ball, Vic. 1991. Crop by Crop- Perennials, p. 309-338 In: V. Ball, (ed.) Ball Red Book. 15"1 ed. Ball Publishing, Chicago.

Byme, T. and H. Halevy. 1986. Forcing herbaceous peonies. J. Amer. Soc. Hort. Sci. 1 1 1 :379-383.

Chouard, P. 1960. Vemalization and its relation to dormancy. Ann. Rev. of Plant Physiol. 11: 191-238.

Finical, L.M., A.C. Cameron, R.D. Heins, W. Carlson, and K. Kern. 1998. Forcing perennials: Gaura Iindheimeri ‘Whirling Butterflies’. Greenhouse Grower. 16(7):121-124. Jones, H.G. 1992. Temperature, p. 231-263. In: Plants and Microclimate: a Quantitative Approach to Environmental Plant Physiology. Cambridge University Press, Cambridge.

Kohl, H. 1957. Flower initiation of stocks grown with several temperature regimens. Proc. Amer. Soc. Hort. Sci. 72: 481-484.

Krekule, J.1987. Vemalization in wheat, p. 159-169. In: J.G. Atherton (ed). Manipulation of Flowering. Butterworths, London.

Lohar, DP, and WE. Peat. 1998. Floral characteristics of heat-tolerant and heat- sensitive tomato (Lycopersicon esculentum Mill.) Cultivars at high temperature. Scientia Hort. 73:53-60.

Napp-Zinn, K. 1987. Vemalization- Environmental and Genetic Regulation, pp. 123-132. In: J.G. Atherton (ed). Manipulation of Flowering. Butterworths, London.

Pearson, S., Parker, A., Adams, S.R., Hadley, P., and DR. May. 1995. The effects of temperature on the flower size of pansy. J. Hort. Sci. 70:183-190.

Roberts, E. and R. Summerfield. 1987. Measurements and prediction of flowering in annual crops, p. 17-51. In: J. G. Atherton (ed.). Manipulation of Flowering. Butterworth’s, London.

160 Roberts, E. H., R. J. Summerfield, R.H Ellis, and K.A. Stewart. 1988. Photothermal time for flowering in lentils( Lens culinaris) and the analysis of potential vemalization responses. Ann. of Bot. 61 :29-39.

Thomas, B. and D. Vince-Prue. 1984. Juvenility, photoperiodism and vemalization, p 408-439. In: M. Wilkins (ed.). Advanced Plant Physiology. Pitman, London.

Vince-Prue, D. 1975. Vemalization, pp. 262-291. In: Photoperiodism in Plants. McGraw-Hill, Maidenhead.

Wellensiek, SJ. 1964. Dividing cells as a pre-requisite for vemalization. Plant Physiol. 39:832-5.

Whitman, C.M., R.D. Heins, A.C. Cameron, and CH. Carlson. 19963. Cold treatments, photoperiod, and forcing temperature influence flowering of Levendula angustifolia. HortScience. 31 :1 150-1 153. Whitman, C.M., R.D. Heins, A.C. Cameron, and CH. Carlson. 1996b. Forcing Perennials: Campanula carpatica ‘Blue Clips’. Greenhouse Grower. 14(8): 67-70.

Whitman, C.M., R.H. Heins, A.C. Cameron, and W.H. Carlson. 1997. Cold treatment and forcing temperature influence flowering of Campanula carpatica ‘Blue Clips’. HortScience. 32:861-865.

Yuan, M., W.H. Carlson, R.D. Heins, and AC. Cameron. 1998. Effect of forcing temperature on time to flower of Coreopsis grandiflora, Gail/ardia xgrandiflora, Leucanthemum xsuperbum, and Rudbeckia fulgida. HortScience. 33:663-667.

161 Table 1. The effects of photoperiod and cold treatment on flowering cooled plugs of Gaura Iindheimeri ’Whirling Butterflies’. Plants were cooled for 0 or 15 weeks in a controlled environment chamber set at 5°C. After cold treatment, plants were grown in a greenhouse set at 20 °C.

Weeks Days to Days from Photo- cold Percentage visible visible bud to Days to Flower Final leaf Final plant period SC Year flowering bud flower flower number number height (cm)

10 0 1 10 46 29 75 4 13 36 12 0 1 100 37 25 62 35 14 64 13 0 1 100 35 21 55 47 14 55 14 0 1 100 32 22 54 54 15 59 16 0 1 100 30 22 52 50 16 72 24 0 1 100 31 27 58 43 18 74 NI 0 1 100 30 27 58 45 25 63 10 15 1 40 30 19 49 34 1 1 41 12 15 1 90 30 17 47 42 13 49 13 15 1 100 28 18 46 48 13 51 14 15 1 100 29 17 46 52 15 60 16 15 1 100 29 19 48 44 15 60 24 15 1 100 30 23 53 37 16 68 N I 15 1 100 26 20 46 41 16 46 10 0 2 0 ------12 0 2 100 25 17 42 21 12 34 13 0 2 100 25 19 43 26 16 33 14 0 2 100 26 19 42 34 21 51 16 0 2 100 27 17 44 33 25 61 24 0 2 100 28 18 46 30 23 61 NI 0 2 100 25 16 41 21 21 43 10 15 2 50 28 17 45 10 23 16 12 15 2 100 31 11 42 41 25 38 13 15 2 100 26 13 39 69 26 32 14 15 2 100 25 14 39 65 27 48 16 15 2 100 24 14 38 67 21 50 24 15 2 100 23 14 37 58 14 54 Ni 15 2 100 22 13 35 57 15 60

Significance Year NS NS NS NS “' ”' Weeks of cold SC NS NS NS “" NS '“ Year x Weeks of cold 5C NS NS NS “” m " Photoperiod ... NS NS ...... Year x Photoperiod "" NS NS ‘ ... “ Week x Photoperiod '“ NS NS NS " NS Year x Weeks cold it Photoperiod “' NS NS NS " “ Contrasts for Photoperiods and Cold NI vs 16 hr 0 weeks cold NS NS NS NS " “° Nl vs 24 hr 15 weeks cold NS NS NS NS NS ' NI vs 16 hr 0 weeks cold NS ’ ' NS " ' NI vs 24 hr 15 weeks cold NS NS NS ‘ NS ' P Linear for 0 weeks of cold NS NS NS “ “ NS P Quadratic for 0 weeks of cold NS NS NS " NS NS P Linear for 15 weeks of cold NS NS NS ' ns ... P Quadratic for 15 weeks of cold NS " NS NS NS NS

NS, ', “, “' Non-significant or significant at P5005, 0.01, or 0.001, respectively.

162 Table 2. The effects of cold treatment on flowering cooled plugs of Gaura Iindheimeri “Whining Butterflies’. Plugs were cooled for 0, 3, 6, 9, 12, and 15 weeks in a controlled environment chamber set at 5 °C. After cold treatment, plants were forced in a greenhouse set at 20°C under 16 hours of light from HPS lamps.

Final Initial Days to Days from Final plant Weeks Percentage leaf New leaf visible visible bud to Days to Flower leaf height of SC Year flowering number number bud flower flower number number (cm)

0 1 100 8 11 26 21 47 85 19 48 3 1 100 10 13 32 19 50 86 23 51 6 1 100 8 1 1 28 1 8 46 67 19 37 9 1 1 00 7 14 31 21 53 86 21 45 12 1 100 8 11 35 17 52 69 19 39 15 1 100 7 10 35 18 53 81 18 51 0 2 100 8 13 26 17 44 41 21 50 3 2 100 8 13 20 17 37 49 21 51 6 2 100 8 10 19 18 38 58 18 39 9 2 100 8 19 16 35 86 17 40 12 2 100 8 19 15 34 85 17 40 15 2 100 8 12 17 28 85 17 38

Significance Year ...... NS

Weeks of cold SC ... “ NS NS '" NS '" Year x Weeks of cold SC “ "' NS '“ “' NS NS Contrasts 0, 3, 6, 9, 12, and 15 weeks 5C P w ... NS NS NS ......

P W.,, ' NS NS NS “* NS NS

NS, ’, “, ”" Non-significant or significant at P5005, 0.01, or 0.001, respectively.

163 Table 3. The effects of forcing temperature on flowering of cooled plugs of Gaura Iindheimeri ‘Whirling Butterflies’. Plugs were cooled for 12 weeks in a controlled environment chamber set at 5 °C.

Actual Final plant Set temp. Actual temp Days to Days from Flower height temp. Force to Force to visible visible bud Days to Flower Final leaf diameter (cm) (C) VB FLW bud to flower flower number number (cm)

17 17.2 17.6 32"“ 19‘“ 47 ... 77'“ 13"“ 4.4'“ 28'” 20 20.3 20.1 26 16 41 63 12 4.1 24 23 23.7 23.9 23 15 39 66 10 4.0 20 26 26.4 26.2 22 15 35 44 10 3.6 17 29 29.2 29.2 22 13 41 13 8 2.9 12

Contrasts 14, 17, 20, 23, and 26 °C p w NS ... N S ......

. P W ...... NS ..

1"“1:

-.

NS, *, **, *** Non-significant or significant at P5005, 0.01, or 0.001, respectively.

E...

Table 4. Parameters of linear regression analysis relating forcing temperature to rate of progress to visible bud (VB), to first open flower (FLW), and from VB to FLW in Gaura Iindheimeri “Whining Butterflies’. Intercept and slope were used to calculate base temperature (TD) and degree-days (°days).

Developmental stage (d) Intercept (b0) Slope (b0) Tb (°C) °days r2 1/d (1/d)/C

Forcing to VB 0.0054473 1 0.001480’ 0.001566 1 0.000215 -3.4 638 0.96 VB to FLW 0.0295019 1 0.003049 0.001559 1 0.000322 -18.9 641 0.96

Forcing to FLW 0.0088113 1 0.000758 0.000747 1 0.000114 -11.8 1337 0.88 ' Standard error.

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Figure 1. The effects of cold treatment and photoperiod on the flowering of Gaura Iindheimeri 'Whirling Butterflies'. Percent flowering, days to visible bud, and days to flower for (A) year 1, 0 weeks of cold, (B) year 1, 15 weeks of cold, (C) year 2, 0 weeks of cold, and (D) year 2, 15 weeks of cold.

165

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Figure 2. The effects of cold treatment and photoperiod on node development of Gaura Iindheimeri Whirling Butterflies' Number of nodes at first flower for (A) year 1, 0 weeks of cold, (B) year 1, 15 weeks of cold, (C) year 2, 0 weeks of cold, and (D) year 2, 15 weeks of cold.

166

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167 LE Flower Tlmlng E1 Flower Tlmlng 75 . 110096 75 u——— -—-———+—-———I——T160% I 15% n so # ,,_ . _ _.‘¥___Q_—__:_—._:.;_é15% “ i >. >‘ , —— .2 - 50% 3 25 "i- —-A~— t 50% 8 __. . *5“. "' f—w —— —— 7 25% A — ———‘_ ——‘ s——‘ {I 25% : " L

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Figure 4. The effects of different durations of cold treatment on the flowering, plant node development, flower number, and plant height of Gaura Iindheimeri Whirling Butterflies'. Flowering percentage, days to visible bud, and days to flower for (A) year 1 and (B) year 2. Number of nodes at first flower for (C) year 1 and (D) year 2). Number of flower buds at first flower for (E) year 1 and (F) year 2. Plant height at first flower for (G) year 1 and (H) year 2.

168

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170 nICHIan smTE UNIV. LIBRARIES IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 31293017721261