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Germination Study of Dormant Cercis canadensisL. SeedGlenn HosokawaEastern Illinois UniversityThis research is a product of the graduate program in Botany at Eastern Illinois University. Find out moreabout the program.

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Recommended CitationHosokawa, Glenn, "Germination Study of Dormant Cercis canadensis L. Seed" (1977). Masters Theses. 3266.https://thekeep.eiu.edu/theses/3266

G ERMINATION STUDY OF DORMANT -

Cercis canadensis L. SEED {TITLE)

BY

Glenn Hosokawa �

B. S. , Loyola University, 1975

THESIS

SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS

FOR THE DEGREE OF

Master of Science

IN THE GRADUATE SCHOOL, EASTERN ILLINOIS UNIVERSITY

CHARLESTON, ILLINOIS

1977 YEAR

I HEREBY RECOMMEND THIS THESIS BE ACCEPTED AS FULFILLING

THIS PART OF THE GRADUATE DEGREE CITED ABOVE

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allow my thesis be reproduced because ��--������������

Date Author

pdm

GERMINATION STUDY O F DORMANT

Cercis canadensis L. SEED

by

Glenn Hosokawa

B.S., Loyola University, 1975

ABSTRACT O F A THESIS

Submitted in partial fulfillment of the requirements for the degree of Master of Science at the Graduate School of

Eastern Illinois University

CHARLESTON, I L L I NO IS 1977

ABSTRACT

Cercis canadensis L. (redbud ) seeds have been shown to require

both scarification and stratification before germination occurs.

Seeds which were leached prior to the scarification-stratification

treatments were found to germinate more quickly, and Caso4 solution

was found to enhance germination. However, the percent germination

remained approximately the same and leaching failed to substitute

for the stratification requirement. Extraction studies indicated

that germination inhibitors were present in scarified-non stratified

seeds, however, the presence of germination promoters in scarified­

stratif ied seeds could not be demonstrated. Germination occurred

in either light or dark conditions. Gibberellic acid was found to

substitute for stratification, and synergistic effects were noted

when 10-2

M thiourea and 10-2 M gibberell ic acid were combined.

lndole acetic acid, kinetin, and abscisic acid failed to substitute

for the stratification requirement. When seeds were stratified for

four weeks, rather than the usual six weeks, germination was decreased

by fifty percent.

G55739

ACKNOWLEDGMENTS

The author is greatly indebted to Dr. Roger L. Darding whose

guidance, everlasting patience, encouragement, and advice made the

completion of this research possible. I wish to thank Dr. T. M.

Weidner and Dr. W. C. Whiteside for reviewing and their helpful

suggestions in completion of the manuscript. My gratitude also

goes to Dr. W. A. Weiler whose interest and many helpful suggestions

concerning the project made the research interesting. A special

thanks goes to a friend, a fellow graduate student, E. A. Traylor,

for technical assistance in laboratory. Lastly, I wish to thank

my parents for their encouragement, interest, and in developing the

photographs.

i i

TABLE OF CONTENTS

INTRODUCTION . . . . . . . . . . . . . . . . . . . . .

MATER IALS AND METHODS . . . . . . . . . . . . . . . . . . . . .

RESULTS AND DISCUSSION . . . . . . . . . . . . . . . . . . . . .

SUMMARY . . . . . . . . . . . . . . . . . . . . . . . . . . . .

L ITERATURE CITED . . . . . . . . . . . . . . . . . . . . . . . .

i i i

1

6

10

26

32

GERMINATION STUDY OF DORMANT Cercis canadensis L. SEED

When a viable seed fails to germinate, given appropriate amounts

of wat er, light, temperature, or suitable atmospheric conditions which

normally induce germination, the seed is said to be in a state of

dormancy (Evenri, 1956; Wareing, 1963; 1965).

Among many legume seeds one of the more common factors attributed

to dormancy appears to be associated with the presence of a very hard,

waxy seed coat (Amen, 1963). McKeever ( 1937) with black locust and

Afanasiev (1944) with redbud reported that the seed coat of legumes

inhibit ed germination. The seed coat may also be impermeable to gases.

In cocklebur, the oxygen requirements appear to be much greater for

the upper dormant seed than the lower non-dormant seed (Wareing and

Foda, 1957). Roberts ( 1963, 1964) reported that anaerobic conditions

in dormant rice seeds was due to the presence of an impermeable seed

coat.

Some seeds require scarification of seed coat before germination

can occur. Amen ( 1965) with woodrush, Toole et al. ( 1964) experimenting

with peanuts, and Westra and Loomis ( 1966) with sea oats have reported

that scarification or removal of the seed coat was required for or to

enhance germination.

In similar studies (Kahn, 1966; Toole et al., 1964; Wareing and

Foda, 1957; and Westra and Loomis, 1966) have indicated that the removal

of soluble germination inhibitors can promote germination.

2

Other seeds require stratification before dormancy is broken.

Germination of cow-wheat (Curtis and Cantlon, 1963; Cantlon et al. ,

1963) requires exposure to cold temperature (4C) for a period of 5-6

weeks. Related germination studies with Elberta peaches (Donoho and

Walker, 1957) and hazel (Bradbeer, 1968; Bradbeer and ·Colman, 1967;

Frankland and Wareing, 1966) also showed a requirement for cold

temperatures. Frankland (1961) reported that gibberellic acid can

substitute for the cold treatment in seeds of hazel and beech. During

stratification of cherry seeds, there have been reported changes in

cell number, dry weight, oxygen uptake, and length of embryo axis

(Olney and Pollack, 1960; Pollack and Olney, 1959). Afanasiev (1944)

reported, in redbud seeds, that catalase and peroxidase activity

increased, as well as starch content. However, protein and fat content

decreased during the stratification period. Thus, increases in meta­

bolic processes appear to accompany the process of stratification (Amen,

1963).

Studies by lkuma and Thimann (1963, 1964) indicate that light

affects germination i� certain species. Their research showed that

red light promoted and far-red light inhibited lettuce seed germination;

but far-red inhibition could be reversed by red light. This photo­

sensitivity appears to be located in the endosperm layer of the lettuce

seed (lkuma aBd Thimann, 1963). Researchers Kahn (1960), Toole and

Cathey (1961), and Toole et al. (1955a, b) reported that red light

can reverse the inhibitory effects of improper temperature on germina­

tion in lettuce and peppercress seeds. Investigations (lkuma and

Thimann, 1960; Kahn, 1960; Kahn et al. , 1957; Toole and Cathey, 1961;

Toole et al., 1955a, b) have shown that gibberellic acid may be substituted

3

for red l ight in germ ination of lettuce and peppercress seeds. In

related stud ies with pea seedl ings, Kohler and Lang (1963) and Simpson

and Wa in (1961) reported that exogenous or endogenous gibberellic acid

could counteract the germination inh ibitory substances produced in the

stems. Thus, a dormant seed may require light, scarif.ication, strati­

f ication, leaching, or application of growth regulators to break dormancy.

The concept of inhibitor-promoter interactions relative to seed

dormancy suggested by Hemberg (1949) has been supported by several

researchers (Amen, 1963; 1968; Black and Naylor, 1959; Frankland,

1961; lkuma and Thimann, 1960; Kahn, 1960; Kahn et al. , 1957; Overbeek,

1966; Paleg, 1960; 1962; Toole and Cathey, 1961; and Villiers and

Ware ing, 1960). Studies by Pillay (1966) with mazzard cherry seeds,

Hashimoto and Rappaport (1966a, b) with bean seeds, Roberts (1964)

w ith r ice seeds, and Thomas et al. (1965) with sycamor� suggest that

seed dormancy is initiated by either decrease in synthesis of promoter,

or accumulation of inhibitor, or inhibition of promoter by inhibitor.

The presence of inhibitor in seed has been supported by Lipe and Crane

(1966) with isolation of an endogenous hormone 11dormin11 from the integu­

ments of Lovell peach seeds. The inhibitor appears to have similar

antagonistic interactions with gibberellic acid3 described by Thomas

et al. (1965), and the germination of the peach seed coincides with

decreased 11dormin11 level in the seed (Lipe and Crane, 1966). Conforth

et al. (1965) reported that the 11dormin11 seems to be identical in func­

t ion to abscisin I I. Other cases where naturally occurring inhibitors

have been counteracted by hormones have been reported. Overbeek et al.

(1967) reported that 11dormin11 activity in duckweed could be counteracted

by cytokinin benzyladenine, but not with auxin or gibberellin. In

4

cocklebur seed, the naturally occurring inhibitors coumarin and

xanthatin are reversed by exogenous kinetin and red light (Kahn, 1966;

Kahn and Talbert, 1966).

The possibility of more than one hormone being involved in the

germination of seeds is supported by works of lkuma and Thimann (1963)

and Leff (1964) working with Grand Rapids lettuce seeds. They reported

the combination of gibberellin or red light with kinetin, but not kinetin

alone, greatly enhanced germination. The same study showed that a com­

bination of kinetin and red light, but not far-red light, stimulated

cotyledon expansion, and gibberellic acid stimulated hypocotyl elonga­

tion and cotyledon expansion in d ark.

Many researchers have reported that hormones increase enzyme

activity during germination ( Bonner and Varner, 1965; Dreman and Berri,

1963; lkuma and Thimann, 1963; Overbeek, 1966; Simpson and Naylor, 1962;

and Varner, 1964). In lettuce seeds, kinetin and/or red light increased

pectinase and cellulase activity (lkuma and Thimann, 1963). Similarily,

Simpson and Naylor (1962) reported an increase in amylase and maltase

activities in oat seeds after exogenous application of gibberellic acid3.

The present study will attempt to clarify the reasons for seed

dormancy in Cercis canadensis L. Afanasiev (1944) reported that dormancy

in redbud seeds (Cercis canadensis L.) is controlled by two factors; 1) a

seed coat impermeable to water and 2 ) a stratification requirement. He

also noted that increased oxygen tensions, or application of formic acid,

or NaOH resulted in an increased germination rate. Afanasiev futther

observed during stratification that enzymatic activity increased along

with starch content. The aspects of light, leaching, promoter�inhibitor

interactions, and exogenous application of hormones involving germination

of redbud seeds have not been adequately covered in the literature.

During this study each of the above factors will be investigated.

5

6

MATER IALS AND METHODS

Cercis canadensis L. (F. W. Schumacher Co. , Sandwich, Mass. ) seeds

were used in this study. To help prevent fungal contamination ( Plate 1 )

all glassware was washed in detergent, rinsed with 0. 5 N HCl, followed

by distilled water, and autoclaved for 20 minutes. Whatman #1 filter

paper and distilled water were autoclaved for sterilization. Surface

sterilization of Cercis canadensis L. seeds was accomplished by placing

the seeds in cheesecloth bags and soaking in agitated solution of 10 per­

cent Clorox for 30 seconds. The seeds were then rinsed in distilled

water for 20 minutes before being used in all subsequent experiments.

All the experiments were replicated three times.

Leaching, Scarification, and Stratification Experiments

Leaching was accomplished by placing the surface sterilized seeds

in cheesecloth bags (50 seeds/bag ) , and suspending them in a beaker

with water continuously running from a faucet for a period of seven

days. The temperature of tap water ranged between 10-llC during the

leaching period. A portion of the leached seeds was then placed on

9. 0 cm Whatman #1 filter paper in a sterilized 100 x 15 mm Petri dish.

The remaining seeds were scarified with concentrated H2

so4 for 30 minutes,

washed with sterilized distilled water five times, and then soaked in

the sterilized water for three hours. The water was drained off and

the leached-scarified seeds were placed on Whatman #1 sterile filter

papers in a sterilized Petri dish containing 10 ml sterile distilled

7

water. Those leached-scarified seeds requiring stratification were

placed in a dark chamber at 4C for 6 weeks. Seeds not requiring strati­

f ication (Table 1) were placed in a dark chamber at 21C for 6 weeks .

. Prior to placing the seeds in the chambers, the chambers were washed

w ith detergent, rinsed with water, wiped clean with 50 percent Clorox

solution to minimize fungal contamination. Whenever seeds required

addit ional water, 5 ml sterile distilled water was added to all the

Petri dishes. After 6 weeks of stratification, the seeds were taken

from the two chambers and placed in the constant temperature room. The

temperature in the room fluctuated between 26-28C. The seeds were placed

under continuous illumination with 40-watt cool-white fluorescent light,

emitting light intensity of 220 ft-c at the Petri dish surface. As soon

as the radicle protruded (approximately 2 mm) from the seed coat, the

seed was determined to be germinated; and the number of seeds germinated

every 24 hours were recorded.·

The above experiments were repeated again with three modifications

(Table 2): 1) seeds were leached with agitated sterile distilled water

(26-29C); 2) seeds were germinated in sterilized 10-J M Caso4 solution,

instead of sterile distilled water; and 3) seeds with combined treatments

of a) leaching, scarification, and stratification, and b) scarification

and stratification were placed in a constant temperature room and covered

with aluminum foil, so that germination occurred in the dark, rather than

1 i ght.

Germination Inhibitor-Promoter Experiments

An attempt was made to determine if germination inhibitors or promoters

were present in the seeds. The solvents H20, 95% ethanol, acetone, and

benzene were used in the extraction of scarified stratified (4 weeks) seeds

8

Table 1. Treat ment of Cercis canadensis L. seeds. The seeds were germinated in sterile distil led water.

Experiment

1-A

2-B

3-c

4-D

5-E

6-F

7-G

8-H

Leaching

+

+

+

+

Treatment s

Scarification Strat if ication

+

+

+ +

+

+

+ +

Table 2. Treatment of Cercis canadensis L. seeds. Th e seeds were germinated in sterile 10-3 M CaS04 solution. The seeds germinated in dark are designated with DK.

Treat ments

Experiment Leaching Sear if i cat ion Stratification

1-A

2-B +

3-c +

4-D +

5-E + +

5-E (DK) + +

6-F + +

7-G + +

8-H + + +

8-H (DK) + + +

9

and scarified-non stratified seeds. Five hundred scarified-stratified

seeds or scarified-non stratified seeds were macerated in a motar prior

to solvent extraction. Fifty mls of various solvents were added to the

macerated seeds and further maceration was continued. The homogenate

was filtered through Whatman #1 filter paper after 30 minutes. The fil-

trate was then evaporated to one ml volume, and the filtrate was brought

to 50 ml with sterilized 10-3 M Caso4. This procedure was followed for

all the solvents, and the extracts were applied to seeds. The extracts

from scarified-stratified seed s were applied to scarified-non stratified

seeds, whereas the extracts from scarified-non stratified seeds were

applied to scarified-stratified seeds. The controls, scarified-stratified

or scarified-non stratified seeds, were applied with 10-3 M C�so4 solution

(one ml volume of various solvents brought to 50 mls with sterilized 10-3 M

Caso4). These treated seeds were placed in the constant temperature room

(26-28C, continuous illumination with cool-white fluorescent light, 220

ft-c at the Petri dish surface) and observed for germination.

Growth Regulator Experiments

The growth regulating agents gibberellic acid3 (grade C, 80% K salt),

kinetin (grade A), indole 3 acetic acid (grade C), abscisic acid (racemate,

grade A), and thiourea (grade A) were purchased from CALB IOCHEM, LaJolla,

Calif. The desired concentrations were prepared by dissolving hormones

and thiourea in 5 ml 95% ethanol and 10-3 M Caso4 solution. Kinetin and

abscisic acid required heating and stirring to dissolve in the solution.

The prepared solutions were applied to scarified-stratified seeds, and

scarified-non stratified seeds. These seeds were placed in the constant

temperature room (26-28C, cool-white fluorescent light, constant illumina-

tion 220 ft-c at Petri dish surface) and observed for germination.

10

RESULTS AND DISCU SSION

Leaching, Scarification, and Stratification Experiment

As shown in Table 3 and Plate 2, only those seeds which were both

scarified and stratified germinated ( 5- D, 8-H ) . This data verifies

the experiments conducted by Afanasiev (1944). It is interesting to

note that germination occurred more quickly (8- H ) when seeds were leached

prior to the scarification-stratification treatments ( Fig. 1). However,

there is little difference in total percent germination (63% for 5-E and

69% for 8- H ) . Figure 2 indicates that the greatest number of seeds ger-

minated at approximately day four, and also the enhancement effects of

the leaching treatment occurred during days 2, 3, and 4.

The above experiments were repeated with the modification of the

germination phase occurring in 10-3 M Caso4 rather than distilled water.

Table 4 and Figure 3 both show that leaching prior to the scarification-

stratification treatment still enhanced the rate of germination. The

. . . 10-3 M C SO t (84% f 5 E 82� germ1nat1on percentage 1n . a 4 was grea er or - , �

for 5-E [DK], 98% for 8- H, and 85% for 8-H [DK]) than in sterile distilled

water (63% for 5-E and 69% for 8- H ) for both leached and non leach�d seeds

(Table 5). Although leaching increased the percent germination in

scarified-stratified seeds, it failed to eliminate the stratification

requirement in scarified seeds, which had been leached for seven days.

Apparently light is not required for redbud seed germination, since

germination occurs in either light or darkness ( Fig. 3). However, light

11

conditions do increase the rate of germination (Fig. 3) and as expected,

the etiolated seedlings grown in dark conditions were tall and chlorotic

(Plate 3).

12

Table 3. Germination of Cercis canadensis L. seeds in sterile distilled water. The numbers are averages for three replications, each containing fifty seeds. See Table 1 for explanation of experiment designation.

Days After Treatment

Experiment 2 _ 3_ 4 _ s _ 6 _7 _ 8 _L 10

1-A 0 0 0 0 0 0 0 0 0 0

2- B 0 0 0 0 0 0 0 0 0 0

3-c 0 0 0 0 0 0 0 0 0 0

4- D 0 0 0 0 0 0 0 0 0 0

S-E 0 0. 3 3 19 30 30 31 31 32 32

6-F 0 0 0 0 0 0 0 0 0 0

7-G 0 0 0 0 0 0 0 0 0 0

8-H 0 9 31 32 33 34 34 34 34

1 00 -

90 -

Bo -

70 -

c: 60 -0 ....... IU c: E I... 50 -Q.)

(.!j ....... c: Q.) u I... 40 Q.)

0..

30 -

20 -

1 0 -

Experiment 8-H --- • Experiment 5-E --- •

Days After Treatment

13

I

10

Fig. 1. Germination of Cercis canadensis L. seeds in sterile-distilled water. Experiment 8-H represents leached-scarified-stratified seeds. Experiment 5-E represents scarified-stratified seeds.

14

23 -

22 -Experiment 8- H --- .

2 1 - Experiment 5-E --- .

20 -

1 9 -

1 8 -

1 7 -

>-rtJ 16

0

I.. cu 1 5 -0..

"'C cu 14 -..... rtJ c: E 13 I.. cu

t!l 12 -U'I

"'C cu 1 1 -cu

Vl � 10 -0

I.. cu 9 -..0 E ::i

z 8 -

rtJ ..... 7 -0 I-

6 -

5 -

4 -

3 -

2 -

1 -

I I I 4 5 6 8 10

Days After Treatment

Fig. 2. Cercis canadensis L. seeds germinated per day. The points on the graph represent the average number of seeds germinated each day.

15

Table 4. Germination of Cercis canadensis L. seeds in sterile 1 0- 3 M CaS04 solution. The numbers are aver.ages for three replications, each containing fifty seeds. See Table 2 for explanation of ex periment designation.

Days After Treatment

Experiment 2 _ 3_ 4 _ 5 _ 6 _7 _ 8 _9_ 10

1 -A 0 0 0 0 0 0 0 0 0 0

2-B 0 0 0 0 0 0 0 0 0 0

3-c 0 0 0 0 0 0 0 0 0 0

4-D 0 0 0 0 0 0 0 0 0 0

5-E 0 4 11 1 6 24 29 39 4 1 42 42

5-E (DK) 0 2 5 8 13 1 5 23 30 38 4 1

6-F 0 0 0 0 0 0 0 0 0 0

]-G 0 0 0 0 0 0 0 0 0 0

8-H 0 21 32 39 49 49 49 49 49 49

8-H (DK) 0 9 16 20 33 36 40 41 42 42

100 -

90 -

80 -

70 -

c 60 -0 � � c

E 50 -� �

c.!J � c � u 40 � -�

0..

30 -

20 -

1 0 -

I I

2 3 I I I

4 5 6

16

Experiment 8- H -------- & Experiment 8- H (DK) --- 0 Experiment 5-E -------- 4t Experiment 5-E (DK) --- •

I I I I

7 8 9 10

Day s After Treatment

Fig. 3. Germination of Cercis canadensis L. seeds in sterile 10-3 M CaS04 solution. Experiment 8-H represents leached- scarified­stratified seeds, 8-H (DK) represents leached-scarified­stratified seeds germinated in dark, 5-E represents scarified­stratified seeds, and 5-E (DK) represents scarified-stratified seeds germinated in dark.

Table 5. Percent germination of Cercis canadensis L. seeds.

Germination solution Experiment

5-E

8-H

5-E

5-E (DK)

8-H

8-H (DK)

Leached Non-leached

63

69

84

82

85

17

18

Germination Inhibitor-Promoter Experiments

The presence of germination inhibitors in scarified-non stratified

Cercis canadensis L. seeds was investigated. Ethanol and acetone extracts

from scarified-non stratified seeds completely inhibited germination of

scarified-stratified seeds (Table 6); while water and benzene extracts

reduced the germination of scarified-stratified seeds (Table 6). Thus

there appears to be germination inhibitors present in scarified-non

stratified seeds. It should be mentioned that the seeds were stratified

for four weeks, rather than the usual six weeks. This could possibly

account for the poor germination of the controls relative to other experi­

ments (Table 4) .

In order to investigate the possibility of germination promoters

being present in scarified-stratified seeds, water, ethanol, acetone,

and benzene extracts from these seeds were applied to scarified-non

stratified seeds. Since all seeds failed to germinate, it appears that

extractable germination promoters are not present in scarfied-stratified

seeds.

19

Table 6. Effects of extracts from scarified-non stratified Cercis canadensis L. seeds on germination of scarified-stratified (4 weeks) seeds. The numbers are averages for three repl i-cations, each containing fifty seeds.

Days After Treatment

Experiment 2 _ 3_ 4 _ 5 _ 6 _7_ 8 _9 _ 10 -- --

Scarified-stratified seeds (contro 1) 0 4 6 10 14 15 16 17 17

Water extract 0 0 0 2 3 3 3 3

Scarified-stratified seeds (control ) 0 0 2 6 11 13 14 16 16 16

95% ethanol extract 0 0 0 0 0 0 0 0 0 0

Scarified-stratified seeds (cont ro 1) 0 0 4 7 11 12 14 .15 15

Acetone extract 0 0 0 0 0 0 0 0 0 0

Scarified-stratified seeds (cont ro 1) 0 0 3 5 9 12 15 18 18 18

Benzene extract 0 0 2 7 8 10 11 11 11 11

20

Growth Regulator Experiments

Table 7 shows that gibberellic acid can substitute for stratifi­

cation. Increasing gibberellic acid concentration from 10-6 M to 10-3 M

-2 increases germination, while 10 M slightly inhibits germination relative

to 10-3 M. Frankland (1961) reported gibberell ic acid can substitute

for stratification in hazel and beech seeds. It is interesting to note

that the percent and rate of germination follow exactly the same gibber-

ellic acid concentration effects as discussed above ( Fig. 4). Also scari-

fied seeds germinated in gibberellic acid required approximately fifteen

days for the completion of germination, relative to the usual ten days

for scarified-stratified seeds. lndole 3 acetic acid, kinetin, abscisic

acid, and thiourea all failed to eliminate the stratification requirement;

-2 -2 however, a synergistic effect was noted for 10 M thiourea plus 10 M

gibberell ic acid (Table 7).

When gibberellic acid is applied to scarified-stratified seeds

(Table 8) , the number of seeds germinated increased with gibberellic

acid concentration of 10-3 M and 10-2 M. However, concentrations of

10-6 M and 10-5 M reduced germination.

lndole 3 acetic acid at concentration of 10-2 M and 10-3 M completely

inhibited germination, while concentrations of 10-6 M, 10-5 M, and 10-4 M

reduced germination.

Kinetin concentrations at 10-6 M, 10-5

M, 10-4 M, and 10-2 M reduced

germination, while 10-3 M completely inhibited germination.

Abscisic acid completely inhibited germination at 10-2 M, 10-3 M,

and 10-4 M, while it reduced germination at 10-5

M and 10-6 M.

Thiourea reduced germination at all concentrations.

2 1

Table]. Germination of scarified Cercis canadensis L. seeds germinated in growth regulator solution. The numbers are averages for three replications, each containing fifty seeds.

Days After Treatment

E>periment __ 5 _ _ 6 _ _ 7 _ _L _L _!Q_ _1_1_ .J..L ..J.L .J.i_ __!2_ _!L

Scarified-strat ified 0 24 29 39 41 42 42 42 42 42 42 42 42

Scarified control

Gibberel 1 ic acid

10-2 M

10-3 M

10-4 M

10..;5 M

10-6 M

lndole 3 acetic acid

10-2 M

10-3 M

10-4 M

10-5 M

10-6 M

Kinetin

10-2 M

10-3 M

10-4 M

10-5 M

10-6 M

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 14 19 21 27 29 29 31 31 31 31

0 3 8 17 22 28 31 32 34 36 38 39 40

0 3 5 7 10 20 22 23 23 23 23 23

0 0 1 3 5 5 12 13 13 13 13 13 13

0 0 3 4

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

0 0 0 0 0 0

9 10 10 11

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

0 0 0 0

11 11 11

0 0 0

0 0 0

0 0 0

0 0 0

0 0 0

0 0 0

0 0 0

0 0 0

0 0 0

0 0 0

22

Table ]. Continued.

Days After Treatment

Experiment _1 _ _ 5 _ _ 6 _ _ 7 _ _ 8 _ __L __!Q_ _1_1 _ _!L _!l_ _!L _lL _1L

Abscis ic acid

1 0-2 M

1 0-3 M

1 0-4 M

1 0-5 M

1 0-6 M

Thiourea

1 0-2 M

1 0-3 M

1 0-4 M

1 0-5 M

1 0-6 M

Thiourea 1 0-2 M plus Gibberellic acid 1 0-2 M

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 0 0 0 0 0 0 0 0 0 0

0 2 7 1 6 25 30 40 42 43 44 45 45 45

c: 0 .µ

70 -

� 60 -E I.. Q.)

CJ .µ c: 50 -Q.) u I.. Q.)

a..

40 -

30 -

20 -

10 -

Lj

Days After Treatment

Fig. 4. Germination of scarified Cercis canadensis L. seeds treated with growth regulator solution.

24

Table 8. Germination of scarified-stratified Cercis canadensis L. seeds germinated in growth regulator solution. Scarified seeds were stratified for four weeks prior to treatment with growth regulator solutions. The numbers are averages for three replications, each containing fifty seeds.

Days After Treatment

Experiment 2 _3_ 4 _5_ 6 _]_ 8 _L 10

Scarified-stratified control 0 0 0 6 7 14 15 16 17 17

Gibberellic acid

10-2 M 0 0 0 2 9 17 19 26 28 28

10-3 M 0 0 2 6 10 24 29 32 34 36

10-4 M 0 0 6 8 11 12 13 17 17

10-5 M 0 0 2 6 7 8 9 9 12 14

10-6 M 0 0 2 4 6 7 8 8 8 8

lndole 3 acetic acid

10-2 M 0 0 0 0 0 0 0 0 0 0

10-3 M 0 0 0 0 0 0 0 0 0 0

10-4 M 0 0 0 2 2 3 3 3 5 7

10-5 M 0 0 1 5 6 7 7 7 7 9

10-6 M 0 0 0 0 0 0 2 3 3

Ki net in

10-2 M 0 0 4 4 5 5 5 5 6

10-3 M 0 0 0 0 0 0 0 0 0 0

10-4 M 0 0 0 0 0 0 0 0 2 2

10-5 M 0 0 2 6 7 7 7 8 10 10

25

Table 8. Continued.

Days After Treatment

Experiment 2 _3_ 4 _5_ 6 _7_ 8 __L 10

Abscisic acid

10-2 M 0 0 0 0 0 0 0 0 0 0

10-3 M 0 0 0 0 0 0 0 0 0 0

10-4 M 0 0 0 0 0 0 0 0 0 0

10-5 M 0 0 4 5 7 7 7 8 9

10-6 M 0 0 5 6 7 7 7 8 8

Thiourea

10-2 M 0 0 2 6 8 12 13 13 14 15

10-3 M 0 0 0 1 1 1 1

10-4 M 0 0 2 5 5 5 6 6 7 7

10-5 M 0 0 5 9 11 12 12 12 12 12

10-6 M 0 0 4 4 6 6 6 6 6

SUMMARY

Cercis canadensis L. (redbud ) seeds required both scarification

and stratification before germination occurred. The results verify

26

the experiments conducted by Afanasiev (1944), and correspond well with

the results of scarification studies with woodrush ( Amen, 1965), peanuts

(Toole et al. , 1964), and with sea oats (Westra and Loomis, 1966).

The similar stratification requirements (for 5-6 weeks, at 4C ) were

described in the works of Curtis and Cantlon (1963), Cantlon et al.

(1963) using cow-wheat; and other seeds requiring cold treatment were

described by Donoho and Walker (1957), Bradbeer (1968), Bradbeer and

Colman (1967), and Frankland and Wareing (1966).

However, leaching failed to promote germination for redbud seeds;

and apparently the removal of soluble germination inhibitors, reported

by Kahn (1966), Toole et al. (1964), Wareing and Foda (1957), and Westra

and Loomis (1966), did not occur in redbud seeds. Nevertheless, the

germination of redbud seeds occurred more quickly with leaching prior

to the scarification-stratification treatments.

In addition, Caso4

solution enhanced germination of the seeds com­

pared to distilled water. The enhanced germination may be due to avail­

ability of calcium in the solution, wh�ch appears to be essential in

maintaining and formation of cell-membranes ( Marinos, 1962). The author

strongly suggests the use of Caso4 solution in future seed germination

studies.

27

Since germination occurs in either light or in dark conditions,

light is not a critical factor in the germination of redbud seeds, as

in germination of certain seeds reported by lkuma and Thimann (1963,

1964), Kahn (1960 ) , and Toole and Cathey (1961) . However, better germi­

nation rate occurred in light than in dark.

When presence or absence of germination inhibitors in scarified­

non stratified seeds, and germination promoters in scarified-stratified

redbud seeds were tested, only the presence of germination inhibitors

could be shown. The extraction tests could not verify whether dormancy

in redbud isicaused by either decrease in promoter synthesis, or accumu­

lation of inhibitor, or inhibition of promoter by inhibitor as suggested

by Pillay (1966 ) , Hasimoto and Rappaport (1966a, b ) , Roberts (1964 ) ,

and Thomas et al. (1965 ) . Thus further investigation in the area appears

to be needed.

Just as gibberellic acid can substitute for the cold temperature

in hazel and beech seeds ( Frankland, 1961 ) , the hormone can replace

the cold treatment requirement in redbud seeds. Hormones indole acetic

acid, kinetin, and abscisic acid failed to replace the cold treatment.

When gibberell ic acid and thiourea were combined synergistic effects

were noted in germination of redbud seeds. Thus� gibberell ic acid can

somehow replace the cold temperature requirement, and either gibberell ic

acid or cold temperature along with scarification process are required

for the germination of redbud seeds. Reports of Olney and Pollack (1960 ) ,

Pollack and Olney (1959 ) , and Afanasiev (1944 ) on increased metabolic

processes and enzymatic activities during stratification process, and

increased amylase and maltase activities in oat seeds after exogenous

application of gibberellic acid3 ( Simpson and Naylor, 1962 ) may suggest

28

similar activities in redbud seeds. Therefore, further investigation

would be of interest to determine if during the stratification process

there is increased gibberellic acid content, and effects of gibberellic

acid on the enzymatic activities in redbud seeds during germination.

29

�l ate 1. �erc1s canadensis L. contaminated with fungi.

Plate l.

9 day

1.: , 1 A

!:!:!:£J.!�L.

Treat.rnt

( - 'Ii IA• hJna ( • ) 'r•rifJ t loo ( • "'tnti t ati""

C rmfMticn pf'ri

JlJl\t' l • 197

Cercis canadensis L. seedlings atter 9 days ot germination period.

jU

Plate 3.

( ) J:Aeching + ) lceri i.,. ion + ) N i ice ion

Guu,.tion period

...,.rl

C'erris ('anadensis

Treatment;

( + ) Leaching ( + ) ScarifiC'ation ( + ) Stratification

GPrminetion period

OYPmb(•[' }6, 1976

( 4C )

10 days

No. I - h

Trc>atment:

( +) ( +) ( + )

Leaching Scarification Stratification

Germination period

November 16, 197 6

( lj(')

lQ rleyR

No - h Ill-.)

31

Ce re is period

canadensis L. seedlings after 10 days of germination in either constant light or dark (DK).

32

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34

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35

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