Aspen trees are dioecious, bearing male and female flowers on separate trees providing interesting breeding opportunities to leverage their genetic potential. An Open4st process protocol was developed describing how to produce aspen seed indoors under fluorescent lights. Controlled pollinations are made using cut branches from male and female aspens.
Introduction
Aspen flower bearing branches are cut from desirable trees before bud expansion and placed indoors in 5 gallon buckets of water under constant fluorescent lights with temperatures between 60 and 75 degrees. Male flower branches are started first and allowed to shed pollen, which is cleaned and stored for later fertilization of the female aspen flowers. The female flowers shed seed within a few weeks, which can be separated from the cotton using low pressure air flow and screens. Seed can be sown directly or frozen for several years over Drierite desiccant in a sealed container.
5 gallon buckets: Five gallon buckets are filled with clean water to sustain the cut branches. The buckets must be clean. Use bleach as needed to remove the bacterial slime that accumulates during the flushing period.
Fluorescent lights: Hang 4' shop lights with 40 watt cool white lamps at about 6.5' from the floor and spaced about 4' apart. Shop lights can also be in a vertical position, within a few feet of the branches but it does not appear as efficient. Leave the lights on 24/7. LED lights have not been tried yet.
Room Conditions: The "breeding" room should have a door and be capable of being sealed off. Be sure to seal any heating vents. Ideally, the floor should have a drain to allow washing the floor between pollen collections and fertilizations. The heating system should be room controlled to allow 60 to 75 degree temperatures. Try to limit humidity when the pollen is shedding.
Tools: Lopping and/or pruning shears are needed to cut 1/2" of the branch ends when changing the water. Use a high quality face mask when working with the pollen. Several stainless steel mesh strainers of various sizes to filter pollen and seed from unwanted debris.
Seed Macerator: Use a medium sized shop vac with an exhaust port and a flexible hose that tapers to about a 1.5" end then secure 2 plastic 1/4 hoses to the end. These hoses should be inserted top and bottom of a 1 quart clear plastic tumbler that fits atop the macerator having 3 PVC couplers with filter cloths and mesh holes sized to trap cotton or seed for that filtering level. See: Aspen-Cotton-Seed-Macerator photos.
Journal: Record breeding activities in a journal for historical purposes. This will save significant time and effort when the process is repeated.
1. This process is based on the document: "Procedures for Crossing Bigtooth and Quaking Aspen" [1].
2. Create a Breeding Plan: Plan the breeding schedule around known limiting factors. These generally are the natural pollen shedding dates of the males and the receptive dates of the females. This information should be tracked in a journal for easy reference. The actual dates are subject to change depending upon various environmental and clonal factors. Therefore, allow enough time to collect the male pollen before the females are receptive. It is possible to delay the process by cutting the branches before bud break and store them in 5 gallon buckets of water at 32 degrees. There is a limit on how early branches can be collected. In some years, with some clones the males will not shed pollen unless they are harvested close (eg. 1 - 2 weeks) to their natural shedding date.
3. Determine the Sex of Aspen Trees: See: How to Determine the Sex of Aspen Trees.
4. Branch Collection: Use common sense and all safety precautions when cutting tree limbs! Lower branches can be cut with conventional pole pruners. Higher branches can be cut using a double sided Pocketech hand chain saw positioned with a Pocketech Limb Hand Chain Saw with Sling Shot And Weight Bag. McGovern made a portable extension saw by purchasing round aluminum tubing in 6' lengths with diameters from 1 1/8" to 3/4", inserted them end to end into each other, drilled 1/4" holes on each end and secured with 1/4-20 nuts and bolts and bolted a curved hand pruning saw on the end. There are 6 tube sections providing a 36" long pole that is capable of reaching branches 40' high. See Extendable Pole Pruner photos.
5. Male Branch Processing: Place the cut male branches in 5 gallon buckets of water and under fluorescent shop lights left on constantly. The branches can be bundled with garden wire with 3-5 branches per "set" for easier handling. The buds will expand and produce pollen within a few days to 1.5 weeks at 60-75 degrees temperature. Adjust the room temperature from cool (60 degrees) to warm (75 degrees) to slow or speed up the development, depending upon the expected receptivity of the female flowers. Normally, male buds do not need to be pruned. However, some clones with larger male catkins tend to require more resources and will perform better if 1/2 to 2/3 of the buds are removed from the branch. Replace the water, cut off 1/2" of stem and clean the bucket with a bleach rinse when the water gets cloudy. Try to avoid cleaning the buckets when the male catkins are fully extended and about to shed pollen. Allow the pollen to shed onto a newspaper. Optionally, tap or cut the loaded catkins with a scissors onto a paper plate if there a few catkins or need to limit pollen dispersal. Pour the pollen through a very fine kitchen screen to remove contaminants then refrigerate in a vial or pill container and inside a sealed jar with Calcium Chloride desiccant (Drierite) until the pollen is needed. It should store for about 4-5 weeks in the refrigerator. Pollen storage for multiple years should use a vacuum freeze drying process [1].
6. Female Branch Processing: Bundle the cut female branches in the field after they have been cut from the tree. Each bundle should contain 3 branches per "set" that are at least 1/2" in basal caliper and 4' tall. Secure each bundle with gardeners wire so that each branch has room for the flowers to be accessed. Prune excess set flower buds to about 10 clusters that will naturally point down so the flowers aren't as likely to break under their own weight. Place the flower buds in the buckets with clean water and under 24/7 lights. Adjust the room temperature from cool (60 degrees) to warm (75 degrees) to slow or speed up the development of the flowers. Replace the water and scrub the bucket with a bleach/water rinse every 2-3 days and cut off 1/2" of branch stem ends. This is very important! Some clones are more tolerant to dirty water and a lazy breeder, others will fail all of the flowers, sometimes at the very end.
7. Female flower fertilization: The flowers are fertilized with pollen when they are receptive which is is generally around 15 mm long but varies with each clone and species. Use a lighted magnifying glass to view the stigmas that will be pinkish red and once pollinated successfully become green in 24-36 hours. If the pollen is dead or is incompatible with the female the stigmas will remain red for a few days. Fertilization is accomplished by lightly brushing the pollen from a 1 oz disposable cup with a "Testers" model paint brush. The cup is used to support the flower during pollen application. Do not re-use the pollen. After fertilizing, swing the branch sets in the air a few times to emulate "wind pollination", impregnate pollen into the stigmas and remove excess pollen drift as the branches are transported to the buckets. Inspect the stigmas with a magnifying glass after the "wind treatment" to verify that pollen has adhered to the sticky stigmas. Only fertilize enough flowers that the branch can support. Typically this means to eventually prune to 4-8 flowers per branch. The actual flower bearing capability is dependent upon many factors including water cleanliness, branch vigor and clonal variation.
8. Seed collection: It is important to monitor the seed development process. A properly maintained and vigorous female branch may disperse seed on its own, but sometimes the seed may not disperse. Watch the ovary tip just below the dried stigmas for splitting that indicates the seed may be ready to shed. This is a good time to open a few ovaries and look for dry cotton and seeds that separate easily from the cotton. The process can also be controlled by cutting the catkins when 1/4 of the ovaries have shed seed and the cotton and seeds are dry to the touch. Gently press the capsules with your fingers to split open the ovaries. Place the catkins on a paper plate and allow the flower to naturally shed the cotton/seed. Place the cotton/seed in a custom macerator with several layers of screens. Apply forced air from the blower side of a vacuum cleaner to separate the seed from the cotton. Place the seed in paper packets marked with the cross name and dated. Store the seed packets in the refrigerator with a desiccant for a few weeks or in the freezer for up to 5 years.
9. Growing the seed: Contract with Rakers Greenhouse to grow aspen seed (preferred) or start them indoors via: Aspen Seed Propagation Under Fluorescent Lights.
Results
It is important to update the breeding success and failures of the breeding project in the respective journals, photo albums and databases. These results will be invaluable for reference or in preparation for the next breeding project.
Aspen Breeding Journal Examples:
2010-Phase3-Crosses-Orignal: See worksheets "BranchPollinations" and "BranchCollections".
2006-A502xGrober-Project: See all worksheets
A listing of links to photos or galleries to help describe the process.
1. Brent A. Stewart (1975) Procedures for Crossing Bigtooth and Quaking Aspen. 13p.
2. John Fiveash, James L. McConnell (1989) A Storage Method for Pollen Using Freeze Drying.
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