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A THESIS FOR THE DEGREE OF MASTER OF SCIENCE

Growth and Flowering Responses of

Lysimachia mauritiana Lam. to

Cold Temperature, Photoperiod, and Juvenility

저온과 일장 및 유년성에 대한

갯까치수염의 생장 및 개화 반응

BY

NAM HYUN LIM

FEBRUARY, 2019

MAJOR IN HORTICULTURAL SCIENCE AND BIOTECHNOLOGY

DEPARTMENT OF PLANT SCIENCE

THE GRADUATE SCHOOL OF SEOUL NATIONAL UNIVERSITY

i

Growth and Flowering Responses of

Lysimachia mauritiana Lam. to

Cold Temperature, Photoperiod, and Juvenility

NAM HYUN LIM

DEPART OF PLANT SCIENCE

THE GRADUATE SCHOOL OF SEOUL NATIONAL

UNIVERSITY

ABSTRACT

These studies were conducted to investigate the growth and flowering responses

of Lysimachia mauritiana Lam. (갯까치수염, spoonleaf yellow loosestrife) to

photoperiod and cold treatment (Expt. 1), and juvenility (Expt. 2). In Exp. 1, seed

propagated 8-week-old seedlings with 4–6 leaves were delivered to cold treatment

at 5oC and 9/15 h photoperiod for 0, 3, 6, 9, and 12 weeks (wk). After cold treatments,

plants were forced under five photoperiod treatments (9/15, 12/12, 14/10, 16/8, and

ii

24/0 h) in a greenhouse. Vegetative growth such as the number of leaves, plant width

and height, and biomass increased with increasing photoperiod, regardless of cold

treatments. In the non-cold treatment group, flowering did not occur during the

experimental period, even under ≥ 16/8 h. Under 16/8 and 24/0 h, flowering

percentages were 83% and 82% at 6-wk, and 100% at both 9- and 12-wk cold

treatments, respectively. However, flowering was also induced under 14/10 h with ≥

6 wk cold treatments with flowering percentages of ≤ 20%. In Expt. 2, growth stages

were divided into two groups according to the number of leaves: growth stage 1 (4–

6 leaves) and 2 (10–12 leaves). The seedling period were 6 and 10 weeks in the

growth stage 1 and 2, respectively. Cold treatments were forced for 0, 5, and 10 wks

and photoperiod treatments were the same as Exp. 1, but the experimental location

was changed to a closed system to examine accurate responses under controlled

temperature conditions. Similar to the former results, vegetative growth was

significantly promoted under longer photoperiod, while decreased with increasing

duration of cold treatment. Furthermore, although the plants were exposed to a cold

temperature at 5oC, flowering was not induced in the growth stage 1. Only in the

growth stage 2, flowering was observed with flowering percentages of 40% and 47%

at 5-wk, and 71% and 91% at 10-wk cold treatments under 16/8 and 24/0 h,

respectively. Therefore, both cold temperature at 5oC and long-day photoperiod were

absolutely required for the flowering, implying that these plants require vernalization

and long-day. In addition, L. mauritiana requires an appropriate level of vegetative

iii

growth to respond to cold temperature, but more detailed studies are needed for

juvenility.

Additional key words: daylength, floral induction, photoperiodism, qualitative long-

day plant, vernalization

Student number: 2017-22672

iv

CONTENTS

ABSTRACT ·················································································· i

CONTENTS ················································································ iv

LIST OF TABLES ·········································································· v

LIST OF FIGURES ········································································ vi

INTRODUCTION ·········································································· 1

LITERATURE REVIEW··································································· 4

Flowering Responses to Photoperiod ·················································· 4

Flowering Responses to Cold Temperature ··········································· 4

Flowering Responses to Juvenility ····················································· 5

MATERIALS AND METHODS ·························································· 7

RESULTS ··················································································· 11

Flowering Responses to Cold Temperature and Photoperiod (Expt. 1) ··········· 11

Flowering Responses to Juvenility (Expt. 2) ········································ 17

DISCUSSION ············································································· 22

LITERATURE CITED ··································································· 27

ABSTRACT IN KOREAN ······························································ 33

v

LIST OF TABLES

Table 1. Effects of cold temperature and photoperiod on vegetative growth of

Lysimachia mauritiama after 15 weeks of photoperiod treatment in Expt. 1.

······················································································ 13

Table 2. Effects of cold temperature and photoperiod on reproductive growth

of Lysimachia mauritiana in Expt. 1. ········································· 16

Table 3. Effects of growth stage, cold temperature and photoperiod on

vegetative growth of Lysimachia mauritiama after 15 weeks of

photoperiod treatment in Expt. 2. ············································· 18

Table 4. Effects of growth stage, cold temperature and photoperiod on

reproductive growth of Lysimachia mauritiana in Expt. 2. ················· 21

vi

LIST OF FIGURES

Fig. 1. Schematic diagram of experimental schedule and treatment timing in

this study. ··········································································· 9

Fig. 2. Effects of cold temperature and photoperiod on vegetative and

reproductive growth of Lysimachia mauritiana in Expt. 1. ··············· 14

Fig. 3. Changes in indoor temperature of a greenhouse during Expt. 1. ···· 15

Fig. 4. Effects of cold temperature and photoperiod treatments during the

growth stage 1 on growth of Lysimachia mauritiana in Expt. 2. ········· 19

Fig. 5. Effects of cold temperature and photoperiod treatments during the

growth stage 2 on growth of Lysimachia mauritiana in Expt. 2. ········· 20

1

INTRODUCTION

Interest in Korean native plants continues to increase and make a new demand in

the flower market, and the scale of wild flower markets is consistently increasing

(Korean Forest Service, 2018). However, the flowering responses are still unknown

in many wild species and many greenhouse growers have difficulties in scheduling

native plants and developing new flowering crops from wild flower (Davis and

Andersen, 1989; Fausey and Cameron, 2005).

Lysimachia mauritiana Lam. (갯까치수염, spoonleaf yellow loosestrife; family

Primulaceae) is regarded as a potential pot plant for succulent leaves and attractive

white flowers. This species is a monocarpic herbaceous plant, biennial, and

flowering season is from July to August in Korea. Many of these plants grow

naturally in exposed rock crevices and seepages on the seacoast and are distributed

in southern coastal region and island, Jeju-do and Ullung-do in Korea (Korea

Biodiversity Information System). Most of the coastal species have high tolerances

to environmental stresses, so their hardiness can be one of the advantages in

introduction and development for new ornamental flowering plants (Kodela et al.,

2014).

In many herbaceous perennials, growers control flowering by temperature,

photoperiod, or both. Ornamental herbaceous species in a temperate climates require

optimum photoperiod and exposure to cold temperature for flowering (Mattson and

2

Erwin, 2005). Photoperiodic flowering responses are generally divided into five

groups: short-day plants (SDP), long-day plants (LDP), day-neutral plants (DNP),

intermediate day plants, and ambiphotoperiodic-day plants. SDP and LDP groups are

further classified into two categories, obligate (qualitative) and facultative

(quantitative) (Tomas and Vince-Prue, 1997). Many herbaceous perennials in

temperate climate are commonly LDPs, so flowering can be promoted by artificial

long-day (LD) lighting (Heins et al., 1997). Furthermore, many species classified as

LDPs need several weeks of cold temperature (1 to 7 oC) that is exposed to seeds or

young plants and promotes flowering. This exposure to cold temperature for floral

induction is defined as vernalization and the duration of exposure usually ranged

from several days to months (Lang, 1965; Thomas and Vince-Prue, 1997).

Many researchers reported that the effects of photoperiod and cold treatment on

flowering of herbaceous ornamental crops. For example, Lobelia ×speciosa Sweet

‘Compliment Scarlet’ is obligate LDP and cold treatment promoted the flowering

even under SD photoperiod (Runkle et al., 1999). In Leucanthemum ×superbum

‘Snowcap’, ≥ 6 weeks at 5oC and ≥ 16/8 h photoperiod were recommended for

complete and rapid flowering. In Phlox paniculata, LD photoperiod and cold

treatment considerably decreased the time to flower and enabled to have at least three

times more flowers than at SD photoperiod and non-cooled treatment (Runkle et al.,

1998).

However, although plants are forced at cold temperature and LD condition, there

3

may not be flowering if plants are not mature enough to respond to environmental

signals. This is defined as juvenility that is insensitive to environmental signal for

flowering. The juvenile phase of plants is usually related to plant longevity and long-

lived species such as trees require long-term vegetative growth for flowering (Erwin,

2006). The research of juvenile phase must be accompanied with researches of

photoperiod and cold treatment for floral induction.

Thus, the objectives of this study were to investigate the flowering responses of L.

mauritiana to photoperiod and cold treatment (Expt. 1) and to examine the effect of

juvenility on sensing environmental stimuli for floral induction (Expt. 2). The results

of this study will provide a foundation for flowering manipulation of floricultural

researches and ornamental crop production.

4

LITERATURE REVIEW

Flowering Responses to Photoperiod

Photoperiod, which is decided by night length, are closely related to flowering,

floral induction and initiation, and development of many plant species. At a given

latitude, plants detect the seasonal information from changing photoperiod, and

flowering is induced under optimum season that is the proper environment for

reproduction of the next generation (Mattson and Erwin, 2005). Thomas and Vince-

Prue (1997) divided photoperiodic responses into five groups: SDPs, LDPs, DNPs,

intermediate day plants, and ambiphotoperiodic-day plants, and this classification

have been generalized in flowering plants. Most photoperiodic ornamental

floricultural crops that flower in spring and summer usually require long-days

(facultative LDP), thus growers have applied artificial LD condition or night

interruption (NI) to facilitate growth and control crop production (Heins et al., 1997;

Runkle et al. 1998). However, because photoperiodic responses about all plant

species are not identified, it is important to investigate how photoperiod affects

flowering of herbaceous ornamentals (Runkle et al., 1999).

Flowering Responses to Cold Temperature

In addition to photoperiod, cold temperature is required for flowering of many

herbaceous plants and one of the important signals to notice seasonal changes. The

5

cold temperature treatment for flowering is defined as vernalization and cold

temperature requiring plants in temperate climate experience cold at 1 to 7oC for

several weeks in winter. The duration and effective temperature range are very

different and diverse depending on species (Lang, 1965). While warm temperature

and LD condition generally hasten flowering, flowering time or floral induction may

be delayed in non-cooled plants. Thus, vernalization is often considered as a

prerequisite before optimum photoperiod (Roberts and Summerfield, 1987). Merit et

al. (1997) reported that vernalization of dry seed promoted earlier flowering, and

Weiler and Shedron (1986) also showed that flowering of vernalized plants was

hastened in LD. In Calandrinia and Brunonia, vernalization for 6 weeks at 4.8oC

was effective for promoting flowering and qualities such as the number of

inflorescences or flowers. The combination of vernalization and LD condition

commonly have a positive effect on the floral induction of ornamental species that

flower in the spring and summer season (Cave and Johnston, 2010).

Flowering Responses to Juvenility

Plants are insensitive to environmental stimuli during the juvenile stage and need

a certain period for enough growth or adult stage to initiate flowering (Bernier et al.,

1981). Generally, long-lived species such as trees have a long juvenile period and

juvenile phases of Citrus sinensis and Fagus sylvatica are 5–8 and 30–40 years,

respectively. On the other hand, most floriculture crops have relatively short juvenile

6

phase and Athirrhinum majus needs about one month (Cockshull, 1985; Dole and

Wilkins, 2004; McMahon et al., 2001). In Cinenaria cultivars, when the number of

leaves was 6–8, plants were sensitive to floral stimuli such as cold temperature and

photoperiod (Yeh and Atherton, 1997).

7

MATERIALS AND METHODS

Plant Materials and Growth Conditions

Seeds of Lysimachia mauritiana were sown in 105-cell plug trays filled with a

commercial soil (Baroker, Seoul Bio Co., Ltd., Eumseong, Korea) on May 19, 2017,

in Expt. 1 and on March 23, 2018, in Expt. 2 (Fig. 1). Seedlings were proceeded in

a closed plant production system of experimental farm (Seoul National University,

Suwon, Korea). The temperature and relative humidity (RH) were controlled at 22oC

and 60%, and light intensity and photoperiod were 110 μmol·m-2·s-1 and 9/15 h,

respectively.

In Expt. 2, growth stages were divided into two groups: stage 1 (4–6 leaves) and

stage 2 (10–12 leaves). Growth stage 2 required four more weeks than stage 1.

Transplanted plants in photoperiod treatments were irrigated at least twice a week.

The plants of Exp. 1 were only irrigated with water, whereas plants in Exp. 2 were

additionally fertigated once every two weeks with water soluble fertilizer (EC 0.8

mS·cm-1; HYPONeX professional 20N-20P-20K, HYPONeX Japan Co., Ltd.,

Osaka, Japan).

Cold Treatments

When the number of leaves of seedlings was 4–6 (Expt. 1 and 2) or 10–12 (Expt.

2), seedlings were moved to the cold storage room (Seoul National University Farm,

8

Suwon, Korea) for vernalization. The temperature of the cold storage room was 5oC,

which lasted for 0, 3, 6, 9, or 12 weeks (wk) in Expt. 1 and 0, 5, or 10 wk in Expt. 2

(Fig. 1). During the cold treatment, light intensity and photoperiod were 5 μmol·m-

2·s-1 and 9/15 h, respectively. After cold treatments, seedlings were transplanted in

7-cm (Expt. 1) and 10-cm (Expt. 2) plastic pots filled with commercial soil (Baroker,

Seoul Bio Co., Ltd., Eumseong, Korea) for photoperiod treatments.

Photoperiod Treatments

In Expt. 1, plants were placed in a greenhouse (Seoul National University Farm,

Suwon, Korea) after cold treatments, and then were grown under various

photoperiods. The photoperiods were controlled by supplemental lighting with white

LED (12V SMD 5050 LED, CamFree Co., Ltd., Seoul, Korea) at 9 h short-day to

make 9/15, 12/12, 14/10, 16/8, or 24/0 h photoperiods. The light intensity of white

LED for day extension was only 3 μmol·m-2·s-1 to avoid effects of daily light integral

(DLI) Expt. 2 was proceeded in a closed plant production system for accurate

environmental control. The temperature, RH, and light intensity was maintained at

22oC, 60%, and 300 μmol·m-2·s-1 [110 μmol·m-2·s-1 fluorescent lamp (TL-D 32W RS

865, Philips Lighting Co., Ltd., Eindhoven, Netherlands) + 190 μmol·m-2·s-1 white

LED (LEDT5-9015-DHE, FOCUS lighting Co., Ltd., Bucheon, Korea)],

respectively. Photoperiod treatments were the same as in Expt. 1 and the light

intensity for day extension was only 4 μmol·m-2·s-1.

9

Fig. 1. Schematic diagram of experimental schedule and treatment timing in this study.

10

Data Collection and Statistical Analysis

All vegetative growth parameters, such as the number of leaves, plant width, plant

height, and dry weight, were recorded after 15 weeks of photoperiod treatment. Plant

height was measured from the base of the stem to the shoot apex or the top of the

inflorescence. Dry weight was measured after drying at 80oC for 72 hours. Days to

visible bud and the first open flower, height at floral induction and the first open

flower, and the number of inflorescences were recorded as reproductive growth

parameters. Days to visible bud and the first open flower was counted from the start

date of photoperiod treatment. Plants not having an inflorescence after 15 weeks of

photoperiod treatment were considered as vegetative or non-flowering. All photos

were taken after 15 weeks of photoperiod treatment. In Expt. 1, the greenhouse air

temperature was recorded using a data logger (WatchDog 1000 series, Spectrum

Technologies, Inc., Plainfield, IL, USA). Statistical and graph module analyses were

performed using the SAS version 9.4 (SAS Institute, Inc., Cary, NC, USA) and

SigmaPlot version 10.0 (Systat Software, Inc., Chicago, IL, USA).

11

Results

Flowering Responses to Cold Temperature and Photoperiod (Expt. 1)

In all treatments, vegetative growth such as the number of leaves, plant width and

height, and shoot dry weight was significantly enhanced under longer photoperiod,

whereas plant width and height in non-flowering plants decreased with increasing

duration of cold treatment. Root dry weight of flowering plants was lower than those

of non-flowering plants (Table 1, Fig. 2). Plant height of flowering plants was

promoted as increasing duration of cold treatment by bolting. However, this response

assumed to be overgrowth that was caused by the difference from the starting time

of photoperiod treatments and the low light intensity by a thermal screen in the

winter season (Figs. 1, 3).

Without cold treatment, plants could not induce flowering in all photoperiod

treatments. Flowering occurred when the plants were grown under 14/10, 16/8 and

24/0 h LD photoperiod after cold treatment (Fig. 2). The flowering plants treated

under 14/10 h photoperiod showed very low flowering percentages, 18, 10, and 15%

under 6, 9, and 12 wk cold treatment, respectively, and floral organ degeneration and

abnormal late flowering were observed. The 3 wk cold treatment was not sufficient

for flowering because of low percent flowering, 38 and 33% at 16/8 and 24/0 h,

respectively (Table 2).

However, 16/8 and 24/0 h significantly promoted flowering. The flowering

12

percentages were 83 and 82% after 6 wk, 100% after both 9 and 12 wk cold

treatments, respectively. Days to visible bud and the first open flower did not always

show significant differences among all treatments, but slightly decreased as

photoperiod increased. Likewise, the number of inflorescences had a little increasing

tendency from 16/8 to 24/0 h (Table 2).

13

Table 1. Effects of cold temperature and photoperiod on vegetative growth of

Lysimachia mauritiama after 15 weeks of photoperiod treatment in Expt. 1.

Cold treatment (weeks)

Photoperiod (h)

Number of leaves

Plant width (cm)

Plant height (cm)

Dry weight (g)

Shoot Root 0 9/15 25.9 cz 12.8 d 3.4 e 1.01 c 0.66 b 12/12 32.3 b 15.0 d 5.3 d 1.46 b 0.96 a 14/10 29.3 bc 17.7 c 7.9 c 1.44 b 0.91 a 16/8 26.6 bc 20.6 b 11.1 b 1.63 b 0.87 a 24/0 39.8 a 24.5 a 13.0 a 2.21 a 0.97 a Significance ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ 3 9/15 26.3 bc 15.2 c 4.1 c 1.08 c 0.90 b 12/12 28.6 b 15.0 c 5.2 c 1.24 c 0.85 b 14/10 21.9 c 19.9 ab 11.3 ab 1.57 b 1.08 a 16/8 29.7 ab 17.7 bc 22.5 bc 1.84 b 0.93 ab 24/0 35.3 a 21.8 a 29.0 a 2.42 a 1.00 ab Significance ∗∗ ∗∗∗ ∗∗∗ ∗∗∗ ∗ 6 9/15 25.8 b 12.1 c 4.2 c 0.83 d 0.54 c 12/12 25.5 b 15.2 b 7.1 b 1.06 c 0.69 b 14/10 21.7 b 17.3 a 13.4 a 1.32 b 0.84 a 16/8 33.9 a 13.7 bc 45.3 bc 1.51 a 0.49 c 24/0 35.4 a 14.6 b 41.3 b 1.59 a 0.59 bc Significance ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ 9 9/15 21.9 b 9.9 c 4.4 c 0.67 c 0.51 d 12/12 22.4 b 16.6 ab 8.8 ab 0.93 b 0.67 b 14/10 23.1 b 17.2 a 15.2 a 1.15 b 0.82 a 16/8 34.4 a 14.8 b 48.8 b 1.70 a 0.63 bc 24/0 36.7 a 17.0 a 49.0 a 1.67 a 0.53 cd Significance ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ 12 9/15 29.1 c 11.2 c 5.3 c 0.75 e 0.58 b 12/12 26.8 c 19.2 a 11.1 a 1.11 d 0.75 a 14/10 28.0 c 19.4 a 19.5 a 1.41 c 0.76 a 16/8 35.9 b 15.1 b 47.3 b 1.63 b 0.51 b 24/0 41.4 a 18.0 a 56.9 a 1.88 a 0.53 b Significance ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ ∗∗∗ zMeans within columns followed by different letters are significantly different by Duncan’s multiple range test at p ≤ 0.05. ∗, ∗∗, ∗∗∗ Significant at p ≤ 0.05, 0.01, 0.001, respectively.

14

Fig. 2. Effects of cold temperature and photoperiod on vegetative and reproductive

growth of Lysimachia mauritiana in Expt. 1.

15

Date (mm/dd/yy)

08/01/17 09/01/17 10/01/17 11/01/17 12/01/17 01/01/18

Tem

pera

ture

(o C)

0

10

20

30

40 AvgMaxMin

Fig. 3. Changes in indoor temperature of a greenhouse during Expt. 1.

16

Table 2. Effects of cold temperature and photoperiod on reproductive growth of

Lysimachia mauritiana in Expt. 1.

Cold treatment (weeks)

Photoperiod (h)

Flowering percentage

Days to visible bud

Days to first open

flower

Height at floral

induction

Height at first open

flower

Number of inflorescences

0 9/15 0 – – – – – 12/12 0 – – – – – 14/10 0 – – – – – 16/8 0 – – – – – 24/0 0 – – – – – Significance – – – – – – 3 9/15 0 – – – – – 12/12 0 – – – – – 14/10 0 – – – – – 16/8 38 57.7 70.3 18.2 az 26.3 1.0 24/0 33 50.7 64.0 12.9 b 23.6 1.3 Significance – NS NS ∗ NS NS 6 9/15 0 – – – – – 12/12 0 – – – – – 14/10 18 83.5 a 99.0 a 15.9 21.7 1.0 16/8 83 46.2 b 70.1 b 13.0 26.0 2.0 24/0 82 51.2 b 69.3 b 15.0 24.0 2.6 Significance – ∗∗∗ ∗∗∗ NS NS NS 9 9/15 0 – – – – – 12/12 0 – – – – – 14/10 10 56.0 77.0 10.5 17.9 b 1.0 16/8 100 54.4 79.5 11.8 31.6 a 1.5 24/0 100 51.3 76.2 12.0 30.0 a 1.9 Significance – NS NS NS ∗∗∗ NS 12 9/15 0 – – – – – 12/12 0 – – – – – 14/10 15 73.5 a 96.0 15.4 27.7 b 2.0 16/8 100 58.1 b 82.9 15.7 36.8 a 1.9 24/0 100 51.9 b 81.4 15.3 39.1 a 3.3 Significance – ∗ NS NS ∗ NS zMeans within columns followed by different letters are significantly different by Duncan’s multiple range test at p ≤ 0.05. NS, ∗, ∗∗∗ Non-significant or significant at p ≤ 0.05, 0.001, respectively.

17

Flowering Responses to Juvenility (Expt. 2)

Similar to Expt. 1, vegetative growth remarkably increased under longer

photoperiod. By contrast, plant width and height generally showed declining

tendencies as the duration of cold treatment and growth stage increased (Table 3).

The overgrowth of plant height, which was observed in Exp. 1, did not occur because

plants received the high light intensity (300 μmol·m-2·s-1) in a closed system. In

addition, the vegetative growth was improved and the plant shape in Exp.2 was more

compact than those in Exp. 1 (Fig. 3, 4, 5)

In flowering response, flowering was not induced in growth stage 1 despite

receiving cold treatment (Fig. 4, Table 4). Flowering was only induced in growth

stage 2 and flowering percentages were 40 and 47% at 5 wk, 71 and 91% at 10 wk

cold treatments under 16 and 24 h of photoperiods, respectively. Days to visible bud

and the first open flower significantly decreased with increasing duration of cold and

photoperiod. Height at floral induction also showed the same trend with days to

visible bud. The number of inflorescences under 10 wk of cold and 24/0 h was at

least three times higher than that under other treatments (Fig. 5, Table 4).

18

Table 3. Effects of growth stage, cold temperature and photoperiod on vegetative growth

of Lysimachia mauritiama after 15 weeks of photoperiod treatment in Expt. 2.

Growth stage Cold treatment (weeks)

Photoperiod (h)

Plant width (cm)

Plant height (cm)

4–6 leaves 0 9/15 24.8 e–jz 10.3 h–l 12/12 27.2 b–f 11.2 e–i 14/10 27.5 b–e 11.9 d–h 16/8 28.9 bc 12.6 def 24/0 32.2 a 15.0 b 5 9/15 20.6 lmn 9.1 klm 12/12 21.2 klm 9.7 i–m 14/10 25.9 d–h 10.2 h–l 16/8 27.5 b–e 11.1 f–j 24/0 27.7 bcd 11.2 e–I 10 9/15 18.4 no 8.6 m 12/12 19.6 mn 9.3 klm 14/10 20.5 lmn 8.3 mn 16/8 22.9 i–l 9.7 i–m 24/0 24.4 g–j 11.2 f–j 10–12 leaves 0 9/15 24.9 e–j 11.9 d–h 12/12 24.9 e–j 11.4 e–i 14/10 26.4 c–g 12.3 d–g 16/8 25.5 d–I 12.6 def 24/0 29.2 b 15.5 b 5 9/15 16.5 o 6.9 n 12/12 20.4 lmn 9.5 j–m 14/10 24.7 f–j 10.7 g–k 16/8 26.2 d–g 13.2 cd 24/0 22.4 jkl 12.9 de 10 9/15 18.9 mno 8.8 lm 12/12 21.0 k–n 9.0 lm 14/10 23.3 h–k 10.8 g–k 16/8 25.1 d–I 14.5 bc 24/0 26.8 b–g 19.2 a Significance

Growth stage (G) ∗∗ ∗∗∗ Cold treatment (C) ∗∗∗ ∗∗∗ Photoperiod (P) ∗∗∗ ∗∗∗ G×C ∗∗∗ ∗∗∗ G×P NS ∗∗∗ C×P ∗∗∗ ∗∗∗

zMeans within columns followed by different letters are significantly different by Duncan’s multiple range test at p ≤ 0.05. NS, ∗∗, ∗∗∗ Non-significant or significant at p ≤ 0.01, 0.001, respectively.

19

Fig. 4. Effects of cold temperature and photoperiod treatments during the growth

stage 1 on growth of Lysimachia mauritiana in Expt. 2. No flowering was

observed.

20

Fig. 5. Effects of cold temperature and photoperiod treatments during the growth

stage 2 on growth of Lysimachia mauritiana in Expt. 2.

21

Table 4. Effects of growth stage, cold temperature and photoperiod on reproductive

growth of Lysimachia mauritiana in Expt. 2.

Growth stage

Cold treatment (weeks)

Photoperiod (h)

Flowering percentage

Days to visible

bud

Days to first open

flower

Height at floral

induction

Height at first open

flower

Number of inflorescences

4–6 0 9/15 0 – – – – – leaves 12/12 0 – – – – – 14/10 0 – – – – – 16/8 0 – – – – – 24/0 0 – – – – – 5 9/15 0 – – – – – 12/12 0 – – – – – 14/10 0 – – – – – 16/8 0 – – – – – 24/0 0 – – – – – 10 9/15 0 – – – – – 12/12 0 – – – – – 14/10 0 – – – – – 16/8 0 – – – – – 24/0 0 – – – – – 10–12 0 9/15 0 – – – – – leaves 12/12 0 – – – – – 14/10 0 – – – – – 16/8 0 – – – – – 24/0 0 – – – – – 5 9/15 0 – – – – – 12/12 0 – – – – – 14/10 0 – – – – – 16/8 40 62.7 az 73.7 a 9.0 a 11.8 1.3 b 24/0 47 59.0 a 74.8 a 7.1 b 11.6 3.2 b 10 9/15 0 – – – – – 12/12 0 – – – – – 14/10 0 – – – – – 16/8 71 50.4 b 67.6 a 6.0 b 11.2 3.4 b 24/0 89 34.9 c 51.4 b 5.8 b 11.4 10.7 a Significance

Cold treatment (C) – ∗∗∗ ∗∗∗ ∗∗∗ NS ∗∗ Photoperiod (P) – ∗∗ ∗ ∗ NS ∗∗ C×P – NS NS NS NS NS

zMeans within columns followed by different letters are significantly different by Duncan’s multiple range test at p ≤ 0.05. NS, ∗, ∗∗, ∗∗∗ Non-significant or significant at p ≤ 0.05, 0.01, 0.001, respectively.

22

DISCUSSION

Previous researches have reported that cold treatment and supplemental lighting

for LD condition could enhance vegetative growth and flowering in many

herbaceous ornamental plants (Garner and Armitage, 1998). In general, day

extension by supplementary lighting enhanced vegetative growth and flowering

quality. LD photoperiod significantly improved vegetative growth, such as stem

elongation, the number of branches, internode length, shoot dry weight in Tecoma

stans (Torres and Lopez, 2011).

In most cases, these positive effects of LD condition could be attributed to

increased daily light integral (DLI) from supplemental lighting. Adams et al. (1999)

reported that increasing DLI reduced the period of juvenile phase and promoted the

plant quality. However, in this experiment, the supplemental lighting of very low

light intensity ranging from 1 to 5 μmol·m-2·s-1 was applied for day extension in

order to minimize the effect of DLI. Actually, DLIs were similar under all treatments.

Many studies also showed that the day extension by low light intensity could

improve the crop quality (Kim et al., 2015; Padhye and Cameron, 2008; Runkle et

al., 1998; Torres and Lopez, 2011). From these result, it can be suggested that the

promoting effect by LD could be attributed not solely to DLI but LD itself.

Phlox paniculata Lyon ex Pursh ‘Eva Cullum’ plants remained vegetative under

SD photoperiod and the flowering percentage increased as increasing photoperiod.

23

Cold-treated plants had over three times more flowers than non-cold treated plants

(Runkle et al., 1998). The growth and flowering of Miltonia cultivars were hastened

by artificial chilling temperature under 16/8 h photoperiod (Matsui and Yoneda,

1997).

Besides the inductive photoperiod, many winter annuals, biennials, and perennials

require cold temperature for flowering. The exposure to cold temperature for a

certain period was called vernalization that promotes phase transition from

vegetative to reproductive state. Generally, the range of cold temperature is between

-5 and 15oC and the optimum temperature for plants in a temperate climate are 1 to

7oC. Insufficient cold treatment can cause incomplete or delayed flowering, so

vernalization has been extensively studied in many ornamental crops (Chouard, 1960;

Lang, 1965; Thomas and Vince-Prue, 1997).

In Aquilegia cultivars, which are generally considered to require vernalization,

cooling and LD photoperiod promoted growth and flowering. In addition, Aquilegia

plants without cooling at 4oC showed substantially late flowering responses

(Shedron and Weiler, 1982; White et al., 1989). Phlox paniculata ‘Fairy’s Petticoat’

did not require cold temperature for flowering, but more uniform, robust, and early

flowering was observed in plants treated for 12 weeks at 4.5oC (Iversen and Weiler,

1994).

Winter annuals and perennials show facultative vernalization response and cold

temperature is required for rapid or early flowering, but is not indispensable. By

24

contrast, biennials have an obligate vernalization requirement, so cold treatment is

essentially required for flowering (Michaels and Amasino, 2000). Lysimachia

mauritiana also showed typical biennial responses and obligatory cold temperature

requirement before LD photoperiod for flowering and the time to flower and the

flowering quality, such as the number of inflorescences, were promoted with

increasing duration of cold and photoperiod (Tables 2, 4). These results were in

agreements with Ha (2014) who stated that species that demand vernalization for

flowering are usually LD plants and the shorter time to flowering and the higher

number of flowers and/or inflorescences could be observed with increasing duration

of vernalization.

The cold requirement can also vary among species and the response is usually

quantitative. As the duration of cold increased, flowering was promoted until the

response was saturated in facultative vernalization requiring plants (Lang, 1965). For

example, for the saturated vernalization responses (100% of flowering), Campanula

‘Birch Hybrid’ needed at least 5 weeks at 0 to 12.5oC (Padhye, 2009). 100%

flowering of Dianthus grantianoplitanus ‘Bath’s Pink’ was achieved after cold

treatment at 0oC for more than 4 wk, 5oC for more than 3 wk, and 10oC for 8 wk

(Padhye and Cameron, 2008). In this study, it is suggested that, for high percent and

uniform flowering of Lysimachia mauritiana, ≥ 9 wk of cold treatment at 5oC was

required prior to ≥ 16/8 h photoperiod treatment (Tables 2, 4).

25

In addition, an optimum temperature is important for flowering and the

temperature above the optimum temperatures can inhibit development. The

temperature conditions for vernalization were 14–16oC for cauliflower and 11–17oC

for Miltoniopsis ‘Trinity’ (Lopez and Runkle, 2006; Pearson et al., 1994). In

herbaceous ornamental crops, Veronica spicata ‘Red Fox’ (Fausey and Cameron,

2007) and Dianthus grantianoplitanus ‘Bath’s Pink’ (Padhye and Cameron, 2008)

needed -2.5–0oC and 5oC, respectively, for uniform flowering. Thus, detailed

investigation for the optimum cold temperature treatment for the flowering of

Lysimachia mauritiana is needed.

Plants also required a certain age or stage to perceive cold temperature for

flowering. A juvenile stage is generally short in herbaceous plants and the short

juvenility is commercially desirable to decrease production time (Ramin and

Atherton, 1991). In some species, the meristems are able to perceive cold

temperature when seedling reach a particular stage. The number of leaves and nodes

are commonly used to estimate the juvenility in herbaceous plants (McDonald and

Kwong, 2005; Yuan et al., 1998). The length of juvenile phase was also measured

with a seedling period to inducible phase for flowering. Coreopsis lanceolate ‘Early

Sunrise’ and Anthirrhinum majus initiated flowering when they had 16 and 18–22

leaves, respectively. (Cockshull, 1985; Damann and Lyons, 1993; Yeh and Atherton,

1997). If plants are prematurely forced to inductive cold temperature in juvenile

phase, the plants cannot flower or support quality flowers. Inappropriate timing of

26

cold treatment can decrease the uniformity of flowering and lower the plants quality

(Cavins and Dole, 2001).

In Exp. 2, the floral transition did not occur when cold and photoperiod treatments

were applied during growth stage 1 with 4–6 leaves, whereas flowering was induced

when they were applied during growth stage 2 with 10–12 leaves (Table 4, Figs. 4,

5). These results indicated that this plant has a juvenility and require sufficient

vegetative growth for perceiving cold temperature. But, cold treated plants that had

4–6 leaves could flower in Exp. 1 (Fig. 2). These contrasting results might have been

brought about from the difference between seedling periods. The 4–6 leaves seedling

periods in Exp. 1 and 2 were 8 and 6 weeks, respectively (Fig. 1). This difference

might affect the juvenility and the contrasting results. Thus, additional studies about

the growth stage in this plant are required to judge seedling by adult phase.

In conclusion, Lysimachia mauritiana requires LD condition following cold

temperature for floral induction, indicating that this species is obligate vernalization

requiring LD plants. Additionally, this species needs a sufficient vegetative growth

to be sensitive to vernalizing temperature for flowering. These results can provide a

foundation for manipulating flowering in Korean native wild plants and help

commercial growers to cultivate wild flowers.

27

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33

ABSTRACT IN KOREA

본 연구는 갯까치수염(Lysimachia mauritiana Lam.)의 일장과 저온

처리(실험 1) 및 유년성(실험 2)에 대한 생장과 개화 반응을 알아보기

위해서 진행되었다. 실험 1 에서 파종 8 주 후 본엽이 4~6 매일 때,

9/15 시간 일장 조건에서 각각 0, 3, 6, 9, 12 주 동안 5oC 저온 처리가

진행되었다. 저온 처리 이후에는 온실로 옮겨져 총 5 가지의 일장

처리(9/15, 12/12, 14/10, 16/8, 24/0 시간)가 진행되었다. 영양 생장

지표에서는 저온 처리와 관계없이 일장이 증가함에 따라 엽수, 초폭,

초고 및 바이오매스가 모두 증가하는 것으로 나타났다. 저온 처리를

받지 않은 식물들은 16/8 시간 이상의 장일 조건에도 불구하고 개화가

유도되지 않았다. 9 주 저온처리구에서 16/8 시간과 24/0 시간의

일장에서 개화율은 각각 83%와 82%였으며, 9 주와 10 주

저온처리에서는 16/8 시간의 이상의 일장 조건에서 100%의 개화율이

관찰되었다. 6 주 이상의 저온 처리 후, 14/10 시간의 일장 조건에서도

개화가 진행되었으나 개화율은 20%에 미치지 못했다. 실험 2 는 본엽

수에 따라 생육 단계가 2 개의 처리로 구분되었고, 생육 단계 1 과 2 의

본엽 수는 각각 4~6 매, 10~12 매였으며 육묘 기간은 각각 6 주,

34

10 주였다. 실험 2 의 저온 처리는 기간을 0, 5, 10 주, 총 3 개로

구분하였고, 나머지 환경 조건 및 일장 처리는 실험 1 과 동일하게 진행

되었다. 다만 통제된 온도 환경에서 정확한 반응을 확인하기 위해서

실험 2 는 밀폐형 시스템에서 이루어졌다. 이전의 결과와 비슷하게 영양

생장은 일장이 길어질수록 유의하게 증가하였고, 반면에 저온 처리

기간이 길어질수록 감소하는 양상을 나타냈다. 게다가 5oC 저온에

노출이 되었음에도 불구하고 생육 단계 1 에서는 어떠한 개화 반응도

관찰되지 않았다. 개화는 생육 단계 2 에서만 관찰되었으며 개화율은

16/8, 24/0 시간 일장 조건에서 5 주 저온 처리시 각각 40%, 47%, 10 주

저온 처리 시 각각 71%, 91%였다. 결과적으로 개화에 5oC 저온과 장일

조건을 둘 다 필수적으로 요구하므로, 이 식물은 절대적 춘화 요구도가

있는 질적 장일 식물이라 할 수 있다. 또한 갯까치수염에서 개화 유도를

위한 저온에 감응하기 위해서는 일정 수준 이상의 영양 생장을

요구한다고 할 수 있지만, 유년성과 관련된 보다 구체적인 추가 실험이

필요하다.