Plant Propagation
Laboratory Exercise 1
Seed Propagation of Woody Perennial Plants ö Germination and Seed Dormancy
Reference: Text Chapter 7 pages 212-218 and pages 220-240
Key words and terms from CD: bottom heat, dormancy, germination, gibberellin, hard seeds, imbibition, intermittent mist, parts per million, plug tray, radicle, scarification, seeds, stratification.
Objectives:
1. Learn how to scarify and stratify seeds and to observe the results of these treatments.
2. To observe the effects of dormancy breaking chemicals such as gibberellic acid.
3. To become familiar with several different seed germination containers and methods.
4. Evaluate the importance of temperature, mist and growing conditions in seed germination and seedling growth.
5. Compare several woody plant species with respect to ease of seed germination.
Introduction:
Seeds of most temperate-zone woody species are incapable of germination upon harvest. This condition, known as seed dormancy, acts to ensure that seeds will not germinate during periods when they would be killed by harsh environments. Consequently, in order to induce successful and uniform germination, it is often necessary to apply one or more dormancy-breaking treatments. Two treatments are very common in commercial nursery crop production. These include (a) stratification ö the imposition of a moist, cold period (not freezing) to break physiological embryo dormancy and (b) scarification ö various chemical and mechanical treatments that are used to overcome hard or impervious seed coats so that the imbibition of water and exchange of gases necessary for germination can occur.
Changes in naturally occurring hormones such as gibberellins play a role in breaking physiological seed dormancy during the stratification process. We will do an experiment to see if supplying gibberellic acid directly to the seed can be used to overcome dormancy and stimulate germination of otherwise physiologically dormant seeds.
There are a number of specialized containers that are used in propagation. In this laboratory session, you will be introduced to germination flats and Cone-tainers¨. The germination flat is designed to germinate large numbers of seedlings that are transplanted bare-root to larger containers while they are still quite small. Cone-tainers are designed to allow individual seeds to germinate and grow to a larger size before being transplanted directly outdoors or to a larger pot. They are widely used in the forestry industry and in ecological plantings. Cone-tainers come in various sizes and are held in racks prior to transplanting. The vertical ridges inside the Cone-tainer are designed to encourage roots to grow downward rather than circling the container. Openings at the bottom are designed to air-prune the roots as they reach the bottom of the pot. This results in a good root system that can be removed with the media intact when the plant is transplanted. Specialized planting dibbles are used to create holes that match the shape of the Cone-tainer. Plug trays are an additional type of container used to germinate individual seeds that are later removed and transplanted to larger containers.
Temperature and intermittent mist have a significant effect on germination of seeds. We will germinate seeds under several different temperatures and growing conditions to give you an idea of the effects of these important environmental factors. The role of bottom heat in propagation is particularly important.
The germination and dormancy breaking requirements of woody perennial seeds vary with the species. We have selected a number of species that will give you broad exposure to seed dormancy and germination requirements of these plants. Make sure to observe the appearance of these seeds and the resulting seedlings as nature has many ways to propagate plants from seed. There are several excellent resources that will give you detailed information about how to propagate woody perennial plants from seed. A few are listed below.
Young, J. A. and C.G. Young (1992) Seeds of Woody Plants in North America. Revised and enlarged edition. Dioscorides Press, Portland, OR. (This is the most complete and useful book on woody plant seeds and their germination requirements ö a ãmust haveä for any woody plant propagation library.)
MacDonald, B. (1986) Practical Woody Plant Propagation for Nursery Growers. Portland, OR. Timber Press, Inc. (Good resource on nursery practices used in raising seedlings of woody plants.)
Seeds of Woody Plants in the
This week's lab has 5 parts:
A. Effects of stratification and scarification on germination of Eastern redbud and honeylocust
B. Effects of gibberellic acid (GA3) on germination of Eastern redbud seeds
C. Effects of mechanical scarification on germination of Kentucky coffee tree seeds
D. Seed propagation of other deciduous trees without cold stratification.
E. Seed propagation of selected conifers
All flats and cone-tainers are filled with Premier PRO-MIX 'PGX', a sphagnum peat moss-based growing media that allows easy penetration of roots and resists compaction. This media is particularly well-suited for seed germination.
A. The effects of stratification and scarification on germination of Eastern redbud (Cercis canadensis) and honeylocust (Gleditsia triacanthos).
1. Each group of 3 students should obtain the following seeds for each species and treatment as listed below (donât mix them up). These seeds have been cold stratified for > 90 days.
Treatments:
Eastern redbud Honeylocust
a. untreated - 30 seeds d. untreated - 30 seeds
b. acid scarified 30 minutes - 30 seeds e. acid scarified 30 minutes - 30 seeds
c. acid scarified 60 minutes - 30 seeds f. acid scarified 60 minutes - 30 seeds
2. Subdivide the seeds from each treatment (a, b, c, d, e, f ) into 3 batches of 10 seeds each.
Batch 1 (10 seeds) will be planted today in germination flats
Batch 2 (10 seeds) will be cold stratified 4 weeks then planted - Lab 5 (Feb 5)
Batch 3 (10 seeds) will be cold stratified for 10 weeks then planted ö Lab 9 (Mar 18)
3. Plant Batch 1 of each treatment and species, each in a separate row in the germination flats provided (a total of six rows per group). Plant the seeds so each seed is covered by about ¹ inch of media. Be sure to label each row with plant species, treatment, date, lab section number, and a name identifying your lab group. You will score germination results in Lab 3 (Jan 22).
4. Take Batch 2 and 3 seeds from each treatment and place each batch in its own individual mesh bag. Note: Please place redbud and honeylocust seeds that have received the same treatment into the same mesh bag. The two species can be separated by size after 4 or 10 weeks of stratification. Mesh bags are made by taking a square of mesh, placing the seeds in the center, then enfolding the mesh around the seeds and use a twisty to close up the mesh bag. Write the date, treatment, your lab section #, and a name identifying your lab group on a label and tie the label to the neck of each mesh bag. When completed each group will have 6 bags as follows:
1. redbud (10) / honeylocust (10) ö untreated ö 4 week stratification
2. redbud (10) / honeylocust (10) ö untreated ö 10 week stratification
3. redbud (10) / honeylocust (10) ö 30 min acid ö 4 week stratification
4. redbud (10) / honeylocust (10) ö 30 min acid ö 10 week stratification
5. redbud (10) / honeylocust (10) ö 60 min acid ö 4 week stratification
6. redbud (10) / honeylocust (10) ö 60 min acid ö 10 week stratification
5. While wearing rubber gloves stir the slurry of Captan (a broad spectrum fungicide) then dip each bag briefly into the slurry, then allow the bags to drain briefly on paper towels. Place the mesh bags into the appropriate container which will have moist sphagnum moss in it. Batch 2 seeds will be stored at 40¡ F (4.5¡ C) for 4 weeks, while Batch 3 seeds will be stored at this temperature for 10 weeks.
By the end of Lab 11 (April 1), when final germination counts are scored, you should have a clear understanding of scarification and stratification.
B. Seed treatments that may facilitate germination: Eastern redbud (Cercis canadensis) only
Various chemical stimulants and synthetic analogs of naturally occurring plant hormones have been used to stimulate seed germination. A partial list of treatments includes:
a. gibberellic acid at 500 to 1000 mg/L for 48-72 hours
b. cytokinins (including 6-benzylaminopurine (BAP)) at 100 to 200 mg/L for 24 hours
c. hydrogen peroxide (1% solution for 72 hours)
d. high pressure treatments
e. Hot or boiling water has been shown effective in treating impermeable redbud seeds (Submerge in boiling water for 1 minute or start at 180¡ F, allowing seeds to soak overnight in gradually cooling water.)
f. Fusicoccin appears to remove the inhibitory effect of abscisic acid, a plant hormone present in seeds that delays or prevents germination.
g. Ethylene, a plant hormone, has been shown to stimulate seed germination in some species. Ethrel (Ethephon) is a commercially-available compound that releases ethylene.
As an example, your lab instructor will provide seeds of Eastern redbud (Cercis canadensis) that have been cold stratified > 90 days and have been acid scarified for 30 minutes and treated with different levels of gibberellic acid (GA3). We will test if GA3 will increase germination rates. Perform the experiment as you are advised, making certain to compare the effect of each treatment with a proper control. You will be asked to hand in your group's germination results so that a class summary can be prepared.
Part B (continued): Effects of gibberellic acid (GA3) on germination of dormant Eastern redbud (Cercis canadensis) seeds.
All seeds have been cold stratified for > 90 days and scarified in concentrated H2SO4 for 30 min and triple rinsed with water, then placed in different concentrations of (GA3) for 72 hours.
Treatments:
1. GA3 1000 mg/L soaked 72 hr. 20 seeds
2. GA3 500 mg/L soaked 72 hr. 20 seeds
3. GA3 250 mg/L soaked 72 hr. 20 seeds
4. GA3 125 mg/L soaked 72 hr. 20 seeds
5. control distilled H2O soaked 72 hr. 20 seeds
1. Each group should obtain 20 seeds of each of the 5 treatments.
2. Plant all 100 seeds in separate rows of 10 seeds per row in the appropriately labeled germination flats. Label each row with plant species, treatment, date, lab section number, and a name identifying your lab group
3. You will score germination results two weeks from today.
Each germination flat from Experiments A and B will be marked with colored tape. When the flat is full, place the flat in the location indicated in the table below. Please record on your data sheet where your flat is placed. We will compare germination results from these different conditions to illustrate the effects of temperature and mist on germination of woody plant seeds.
| Tape color |
Location |
Mist |
Bottom heat |
Temperature |
| Green |
Zone 16 bench 1 |
yes |
No |
75¡ F day & night |
| Red |
Zone 16 bench 1 |
yes |
Yes - 80¡ F |
75¡ F day & night |
| Blue |
Zone 21 bench 1 |
no |
Yes - 80¡ F |
75¡/65¡ F (day/night) |
| Yellow |
Zone 21 bench 1 |
no |
No |
75¡/65¡ F (day/night) |
C. Effects of mechanical scarification on Kentucky coffee tree (Gymnocladus dioicus).
Mechanical scarification is an alternate method to chemical scarification when seed coats are particularly hard and thick. Kentucky coffee tree requires scarification for germination. This species exhibits hypogeal germination where the cotyledons remain in the ground, unlike, for example the garden bean, which exhibit epigeal germination where the cotyledons come out of the ground
1. Each group of 3 students should obtain 10 seeds of Kentucky coffee tree.
5 seeds will be unscarified controls
5 seeds will be mechanically scarified
2. Scarify 5 seeds by removing a section of seed coat with emery sanding paper to expose a small spot of inner tissue.
3. Plant 5 seeds per row in the large Cone-tainers labeled for Kentucky coffee tree located in Zone 21. Plant seeds about 1 inch deep. Label each row with species name, date sown, whether they are scarified or unscarified, lab section #, and a name to identify your lab group. Cone-tainers will be placed in Zone 16 (mist bench) without bottom heat. You will score germination in two weeks.
D. Seed propagation of other deciduous trees without cold stratification.
Different woody plant species have different embryo dormancy breaking requirements. Germination results of the following three species will demonstrate differences in their germination potential without cold stratification.
Common horsechestnut (Aesculus hippocastanum) typically requires 60-90 days stratification for germination for maximum germination rates. The seed for our lab was purchased from F.W. Schumacher Co. (www.treeshrubseeds.com) and has not been cold stratified. As an alternative to prechilling, these seeds have been acid scarified 60 minutes and then soaked in water 24 hours.
Each group should plant 5 seeds (one row) in the large Cone-tainers in Zone 21. Plant seeds about 1 inch deep. Label each row with species name, date sown, lab section #, and a name to identify your lab group.
D. Seed propagation of other deciduous trees without cold stratification (continued).
Ginkgo (Ginkgo biloba) To maximize germination ginkgo typically requires 30-60 days warm stratification followed by 60-90 days cold stratification. During warm stratification the immature embryo completes development. The seeds for this lab have been purchased from Schumacher and have only been warm stratified.
Each group should plant 10 seeds (one row) of Ginkgo in the medium-sized Cone-tainers labeled for Ginkgo. Plant seeds about 1 inch deep. Label each row with species name, date sown, lab section #, and a name to identify your lab group.
Red oak (Quercus rubra) typically requires 30-60 days cold stratification for maximum germination rates. We will compare germination rates of acorns not cold stratified (purchased from Schumacher) vs. acorns harvested the first week of January 2007 to see if Nov/Dec outdoor temperatures in Lafayette have cold stratified these acorns sufficiently to increase their germination potential.
Each group should plant 10 unstratified acorns (2 rows) and 10 stratified acorns (2 rows) in the large Cone-tainers labeled for red oak located in Zone 21. Plant acorns about ¹ inch deep. Label each row with species name, date sown, if stratified or unstratified, lab section #, and a name to identify your lab group.
All Cone-tainers with horsechestnut, ginkgo, and red oak seeds will be placed in Zone 16 (mist bench) without bottom heat. You will score germination in two weeks.
E. Seed propagation of Conifers
Conifers have unique growth habits and various dormancy breaking requirements. We have selected three species of conifers to plant. Blue spruce (Picea pungens var. glauca) has no stratification requirement. Cedar of Lebanon (Cedrus libani) requires 28 days cold stratification. White pine (Pinus strobus) requires 30-45 days cold stratification. Both the white pine and cedar of Lebanon seeds used in this lab have been cold stratified.
Each group of three students should obtain the following:
1. Blue spruce ö 10 seeds
2. White pine ö 10 seeds
3. Cedar of Lebanon ö 10 seeds
Plant 10 seeds (1 row) of each species in the small white Cone-tainers. Spruce and pine seeds should be planted to a soil depth of 1-2x the seedâs diameter. Plant cedar about ¹ inch deep. Label each row with species name, date, lab section #, and name to identify your lab group.
These Cone-tainers racks will be placed in Zone 16 (mist bench) without bottom heat. You will score germination in two weeks.
HORT 201 - Results and Data from Laboratory # 1
Seed Propagation of Woody Perennial Plants ö Germination and Seed Dormancy
Name: ______________________________ Lab section and group: ____________________
Results, Part A: Eastern redbud (Cercis canadensis)
Germination environment: _______________________________________________________
_____________________________________________________________________________
Treatment # seeds planted # germinated % germination
per batch
_____________________________________________________________________
A. No stratification (collect data during lab #3, Jan 22)
Control
30 min acid
60 min acid
_____________________________________________________________________
B. 4 weeks stratification at 40¡ F (4.5¡ C) (collect data during lab #7, Feb 19)
Control
30 min acid
60 min acid
_____________________________________________________________________
C. 10 weeks stratification at 40¡ F (4.5¡ C) (collect data during lab # 11, Apr 1)
Control
30 min acid
60 min acid
_____________________________________________________________________
Results, Part A: Honeylocust (Gleditsia triacanthos)
Germination environment: _______________________________________________
_____________________________________________________________________
Treatment # seeds planted # germinated % germination
per batch
_____________________________________________________________________
A. No stratification (collect data during lab #3, Jan 22)
Control
30 min acid
60 min acid
_____________________________________________________________________
B. 4 weeks stratification at 40¡ F (4.5¡ C) (collect data during lab #7, Feb 19)
Control
30 min acid
60 min acid
_____________________________________________________________________
C. 10 weeks stratification at 40¡ F (4.5¡ C) (collect data during lab #11, Apr 1)
Control
30 min acid
60 min acid
_______________________________________________________________________
Results, Part B: Eastern redbud (Cercis canadensis) (collect data during lab #3, Jan 22)
Germination environment: _______________________________________________
______________________________________________________________________________
Treatment # seeds planted # germinated % germination
per batch
______________________________________________________________________________
1. GA3 1000 mg/L
2. GA3 500 mg/L
3. GA3 250 mg/L
4. GA3 125 mg/L
5. Control (distilled water)
Comments and observations:
______________________________________________________________________________
Results, Part C: Kentucky coffee tree (Gymnocladus dioicus) (collect data during lab #3, Jan 22)
Treatment # seeds planted # germinated % germination
______________________________________________________________________________
No scarification
______________________________________________________________________________
Results, Part D: Deciduous Tree Seed Germination (collect data during
lab # 3, Jan 22)
Tree species # seeds planted # germinated % germination
Common horsechestnut
Ginkgo
(Ginkgo biloba)
Red oak - unstratified
(Quercus rubra)
Red oak - stratified
(Quercus rubra)
Comments and observations:
_____________________________________________________________________________
Results, Part E: Conifer seed germination (collect data during lab #3, Jan 22)
Species # seeds planted # germinated % germination
Blue spruce
(Picea pungens var. glauca)
White pine
(Pinus strobus)
Cedar of Lebanon
(Cedrus libani)
______________________________________________________________________________