February 22, 2021

Seed Experiment #1 - Survival Methods for Early Germinators

This article was supposed to go out before my last one, but apparently finding time to photoshop memes is much easier than reading research articles on acorn germination… Anyways, let’s just pretend this article came out a month ago so I don’t have to slow it down by rewriting the introduction.

-------------------------------------------------------------------------------------------------------------------

Source content: 12/20/2020

Hellooooooo bonsai world! It’s been a minute since I posted but I'm happy to report I am alive and well in Ohio. To celebrate surviving the first semester of my Ph.D. program, it's only fitting that my first blog post of 2021 focuses on a bonsai experiment I'm designing - the first of many I have in mind.

Of course, throughout the move to Columbus and starting graduate school, maintaining my bonsai collection has remained a priority even if new articles and videos were put on hold. In August, I packed my trees into a UHaul and drove my trimmed-down collection across the country. Since then, I’ve also found time to scout for local Ohio yamadori, service trees for my first local client, attend two workshops at Yume-en (Rob Hoffman’s new bonsai nursery in Marysville, OH), assemble my first grow tent for overwintering my tropicals, collect new seeds stock for 2021, and join the Columbus Bonsai Society’s Board for 2021. All of these and more from my archive could be future blog posts so stay tuned!

Sections:


A young Brewer's oak seedling emerges!

Purpose

In homage to the parallel universe version of me as a plant scientist, and owing to my own background in microbiology research, I have some ideas for bonsai experiments (paging Dr. Jekyll? No, I’m not quite there yet). In the name of learning, these experiments will be public-facing, academic exercises fully documented here on my blog. Queue the Adam Savage quote “Remember kids, the only difference between screwing around and science is writing it down! For my part, I will ask a question about the intersection of bonsai and biology/horticulture, review past publications to see if other bonsai hobbyists or plant scientists have already come up with an answer, and then if needed I will design my own experiment to test past assumptions or to fill what knowledge void I find in the archives of bonsai care practices. I will also try to share the experimental design before carrying them out so that flaws in my logic can be caught or necessary feedback can be solicited and I will of course share the data here no matter the findings. In this way, I can collect feedback on the experimental designs, methods, and conclusions along the way if others in our community has insight to add. Science is best done collaboratively, after all.

The purpose of this specific experiment is to test storage conditions for early-germinating oak seeds and compare the 4 and 12-month survival of those seeds.

Introduction

While I enjoy spending time in nature looking for native yamadori/wild bonsai, in the name of sustainable bonsai, I am also growing a variety of native American species from seed for bonsai to find which can be grown as easily (as quickly and to as refined of a product) as the famous bonsai species from Japan. For several species of Japanese origin, they have been grown for bonsai for so long that everything from establishing attractive roots to building a mature trunk to creating ramified branches is now well understood and streamlined – I’m talking Japanese black/red/white pines, Chinese/Japanese shimpaku junipers, Japanese maples, and trident maples. To that end, after I encountered a small-leaf variety of oak in southern Oregon (see Figure 1 below), I kept it in the back of my mind as an attractive option until I recently discovered a source to buy Quercus garryana var. breweri seeds. These seeds were purchased from a bulk-seller and have since gone out of stock; however, their arrival included a small population of early-risers which catalyzed today’s experiment. On the eve of an Ohio winter full of freeze/thaw cycles and the occasional polar vortex, what should be done for the long-term survival of oak seeds that germinate early?

Figure 1: The naturally dwarfed leaves of a Brewer's oak yamadori I attempted collecting in 2017.

            Many species of oak (genus Quercus) have been studied over the years for timber, cork production, and conservation purposes. Among the unique traits which must be considered when storing and germinating oak seeds, we must acknowledge that oaks have highly metabolically active seeds which need oxygen and moisture throughout their storage (1). Multiple studies have verified that acorns that were dried prior to storage had significantly lower germination rates, even though drying seeds of other species is typically helpful to elongate their storage lives (2, 3, 4, 5). This characterizes oak seeds as “recalcitrant” (sensitive to drying and freezing) whereas other “orthodox” seeds are resistant to these conditions (3).  However, within the recalcitrant classification, oaks contain further complexity. The “white oak” subgenus Quercus Quercus aka Quercus Lepidobalanus/Leucobalanus (why are there so many names for one subgenus?!) has seeds that mature in 6 months (rather than Red/Black oaks which mature in >1 year) and whose seeds have short storage shelf typically under 1 year before germination rates starts to drop off (3). The reason why white oak acorns have cannot be stored for long periods is primarily their shorter seed dormancy periods; multiple white oak species have now observed this manifest through the phenomenon of seeds germinating early during cold stratification (1, 3, 6, 7, 8). Some studies went on to find that storage of acorns at -1C would prevent early germination among white oaks, but as always species must be considered. While seeds from the bur oak, Quercus macrocarpa,  and English oak, Quercus robur, have been found to tolerate subfreezing temperatures (4, 2), multiple other studies have found subfreezing storage temperatures (even just -1C) result in embryo damage and less germination relative to +1 or +4 C storage conditions (3, 8). While ideal storage conditions have been studied across multiple species of white oak, preserving early germinators is quite unstudied matter. The studies mentioned so far all left them in the cold stratifying population and only made note of them when the cold stratification period ended.

Figure 2: As you can see, Brewer’s oak (bottom) has substantially smaller leaves and seeds compared to wild type Garry oak – and this isn’t even factoring in the reduction that can occur in a bonsai setting. 

In the context of our Garry oak (aka Oregon white oak) subspecies, the early germinating acorns we observed now make sense. While we know white oak storage suggestions for preventing early germination differ by species, storage via subfreezing temperatures to prevent early germination have not been tested in the Garry oak in any published work probably due to its regional distribution. Garry Oak is found between northern California and Southwestern British Columbia and is naturally found in low-altitude, semi-wet environments (9). A recent study conducted on ideal storage conditions for acorns of Quercus garryana wild type (not Brewer’s variety – see Figure 2 above) found that the best course of action was to refrigerate seeds as soon as possible after collection to prevent unwanted germination in cold storage, however, after 6 months of storage, unplanned germination rates rose in all groups tested (1). Meanwhile, the closely related Brewer’s oak subspecies we are working with survives best in its limited natural range of northern California to southwestern Oregon at higher altitudes from 1400-2000m where maximum snowpack ranges from 50”-90”, depending on elevation (10, 11). Although Garry oak germinates in fall with the arrival of autumnal rains and has no dormancy period (1), the more winter-hardy native range of Brewer’s oak indicates our seeds may not behave identically. These differences combined with the utter lack of peer-reviewed research on Brewer’s oak germination and lack of detailed information given from the seed bulk-seller leaves us to chart our own course on how to store our Brewer’s oak seeds and what to do with the early germinating seeds.

In some contexts, early germination is not problematic but rather is encouraged. Vegetable growers who start their plants indoors with grow lights or in a protected greenhouse a few weeks early can see larger plants by the end of the year because of the head start (Mark Comstock does this with his pine seedling-cuttings too; 12, 13). However, growing a temperate tree indoors could have negative consequences too. As with the infamous ‘mallsai’ juniper procumbens which die after being deprived of their natural dormancy for too long, it remains to be seen if a seed that sprouts one full season too early could fall to the same fate or if it would be flexible enough to survive and adjust back into a natural rhythm next fall. Furthermore, because my grow tent space is already overflowing with tropical trees and I do not have a sheltered greenhouse or polytunnel to extend my growing season, my plants are best served by being in sync with Ohio’s seasons and by germinating at the appropriate time. 

Figure 3: Seed anatomy terminology

Now that we know about the fundamentals of oaks and acorn germination and the possible advantages and disadvantages we can expect to see from giving early-germinating acorns special treatment, we must delve deeper into the fundamentals of seed biology before we can formulate specific hypotheses. Upon the arrival of my acorns, I noticed condensation in the bag and white radicles bursting from the pointed tips of ~15 acorns. As you can see in Figure 3 above, a radicle is a scientific word for the first root that emerges. For most seed types, shortly after the radicle emerges, the hypocotyl emerges (the first stem), followed by unfurling of the cotyledons once above ground (the seed’s first leaf structures) and then followed by the plant’s true leaves developing. If my acorns were shipped in spring and planted straight after their required cold stratification, this sequence of events would happen in short succession. However, according to my reading, it is more natural for Quercus garryana and other fall-germinating oak seeds to have their radicle emerge first, grow for an undetermined amount of time, and arrest hypocotyl growth until warmer spring conditions arrive.

The behavior of these Garry oak seeds in their natural range suggests that the best course of action is to plant all these seeds outside through the winter, but we have several unknown factors to consider. First, as I already described, we are dealing with Quercus garryana var. breweri – a subspecies of Garry oak that could behave differently than the Quercus garryana species which had been studied previously. Second, Ohio is outside the natural range (though within the recommended USDA Hardiness zones) of Brewer’s oak and therefore our winters may get to colder temperatures or have more freeze-thaw cycles than the seeds are accustomed to enduring. And if the seeds cannot germinate as they normally would through our unique midwestern winters, would the seeds benefit from continued humid cold storage until spring, or would they do better if to be planted indoors early? All of this is to say – do early germinating seeds benefit from any special treatment, or are they adaptable enough to endure any condition?

I hypothesize three potential methods could underly survival mechanisms for the early germinators. First, by returning the seeds to cold storage, germination might pause until spring – resulting in only minor radicle growth and no hypocotyl growth which will resume when planted in warm temperatures. Alternatively, allowing the early germinators to continue germinating in an indoor environment a few months ahead of time could also result in healthy plants which sprout earlier and potentially grow larger by the end of the first year due to their artificially longer growing season. Also possible, the naturally cold-stratified seeds could prove resistant to the midwestern winters and germinate without issue.  Or finally, none of these conditions matter and the early seeds could be doomed to die by their seasonal dysphoria.

Materials and Methods

Through this experiment, we test these hypotheses regarding whether early germinating Brewer’s oak seeds require any special treatment to ensure continued germination and 1-year survival of those seeds. At the onset of the experiment on 12/12/20, I received 1 pound of Quercus garryana var. breweri from Sheffield’s seeds (now out of stock). The photo series below (Figures 4-12) shows how seeds were sorted by radicle length (using a malleable wire to gauge lengths of curved radicles), assigned an identification number, and planted. Non-germinators were placed into a sealed Ziploc bag in the crisper drawer of my refrigerator with moist paper towels (1, 14). The sorted seeds were then randomized into the three conditions and were either planted outside, planted indoors (grow tent), or placed in a new sealed Ziploc bag with moist paper towels back into the crisper. The group which returned to the refrigerator would be a control for no special treatment to early germinators - as all past artificial cold stratification studies have left early germinators undisturbed until planting time. The randomization was done by alternating which condition received a seed, starting from the shortest emerged radicle to the longest radicle (see table below). Additionally, each group included one seed that had a radicle emerge but had that radicle broken off. This scenario mimics the natural strategy of Siberian chipmunks (Tamias sibiricus) who deal with the problem of early germinating white oak (Quercus mongolica) acorns by pruning off the emerging radicles to preserve their food caches and lends me to believe that radicle regeneration and healthy germination is still possible (7). 

Figure 4: Measuring radicle length 

Figure 5: The seeds for the experiment - you can see the emerging radicles on 1-9 as well as the broken radicles on 10-12.

Figure 6: The continued moist cold stratification condition (no treatment control).

All seeds were picked for immediate planting were put into a mix of 50% pumice, 50% composted pine bark (see Figure 8 for the exact product) which will be a moist yet aerated bonsai mix for the seedlings to grow in upon germination. The planting was done into 32oz plastic yogurt containers with added drainage holes in a clockwise fashion – lowest to highest - with the number of the seed also labeled on an outside position of the pot. The indoor condition was placed in the lower level of my grow tent intended for tropical trees. The outdoor condition was placed in the well of a cinderblock with mulch between the walls of the cinderblock and the pot to insulate the roots from cold temperatures and with moss placed on top of both the indoor and outdoor conditions to retain moisture. Both indoor and outdoor seeds were not given special watering regimens; the indoor seeds were watered 3-5 times a week when the rest of my tropical trees were watered, and the outdoor seeds were watered sparingly – only when there were extended periods without snow or rain, and the rest was left to nature. 

Figure 7: Lifehack for making holes in yogurt training containers - stab with bonsai scissors! A drill bit would be safer but I lost the ones I used to use for this.

Figure 8: This is the brand of aged pine used in the soil mix today. There are many like it, but this one is mine!

Figure 9: Resulting soil mix.

Figure 10: Planting arrangement.

Figure 11: After covering with more soil and moss.

Figure 12: The outdoor condition with mulch and cinderblock insulation.

During the first year of growth, the dates of hypocotyl emergence will be recorded as well as the date of entrance into dormancy (total leaf drop/browning) to check for any impacts of the different conditions on the plants' seasonal circadian rhythms.

At the end of year 1 (spring 2022), the surviving seeds will be separated, weighed, leaf # recorded, height recorded, germination/survival rate recorded, and be planted in individual pots or the ground for further training as done in my previous post about training seedlings for bonsai.

Results

Table 1: Pre-planting info


Table 2: Post-planting results


Discussion/Conclusions

TBD
            All this reading and planning comes with the major caveat that 12 seeds are not a great sample size for making conclusions. This is intended as a pilot study for further investigation by others in the future, but if any clear trends emerge, it may still be informative.

References

1. Devine, W. D., Harrington, C. A., & Kraft, J. M. (2010). Acorn storage alternatives tested on Oregon white oak. Native Plants Journal (Indiana University Press), 11(1), 65–76. https://doi.org/10.2979/NPJ.2010.11.1.65
2. Ozbingol, N., & O’Reilly, C. (2005). Increasing acorn moisture content followed by freezing-storage enhances germination in pedunculate oak. Forestry, 78(1), 73–81. https://doi.org/10.1093/forestry/cpi007
3. Connor, K. (2004). Storing acorns. Native Plants Journal, 5(2), 161–166. https://doi.org/10.2979/NPJ.2004.5.2.160
4. Schroeder, W. r., & Walker, D. s. (1987). Effects of moisture content and storage temperatures on germination of Quercus macrocarpa acorns. Journal of Environmental Horticulture, 5(1), 22–24. https://doi.org/10.24266/0738-2898-5.1.22
5. McCreary, D., & Koukoura, Z. (1989). The effects of collection date and pre-storage treatment on the germination of blue oak acorns. New Forests, 3(4), 303–310. https://doi.org/ttps://doi.org/10.1007/BF00030040
6. Tilki, F. (2010). Influence of acorn size and storage duration on moisture content, germination and survival of Quercus petraea (Mattuschka). Journal of Environmental Biology, 31(3), 325–328.
7. Yang, Y., Yi, X., & Yu, F. (2012). Repeated radicle pruning of Quercus mongolica acorns as a cache management tactic of Siberian chipmunks. Acta Ethologica, 15(1), 9–14. https://doi.org/10.1007/s10211-011-0102-0
8. Hoss, G. (2006). Successful 1-y storage of swamp white oak acorns. Native Plants Journal, 7(1), 69–71. https://doi.org/10.2979/NPJ.2006.7.1.69
9. Renninger, K. (n.d.). What Is A Garry Oak? Oak Harbor Garry Oak Society. Retrieved February 22, 2021, from https://ohgarryoaksociety.org/what-is-a-garry-oak/
10. Quercus garryana var. Breweri. (n.d.). Retrieved February 22, 2021, from https://ucjeps.berkeley.edu/eflora/eflora_display.php?tid=64727
11. 2019 Klamath Mountain Snowpack Updates. (2019, April 11). Bigfoot Trail. https://www.bigfoottrail.org/2019/04/2019-klamath-mountain-snowpack-updates/
12. Shepherd, L. (2011). The Complete Guide to Growing Vegetables, Flowers, Fruits, and Herbs from Containers: Everything You Need to Know Explained Simply. Atlantic Publishing Company.
13. Mark Comstock | Facebook. (n.d.). Retrieved February 22, 2021, from https://www.facebook.com/mark.comstock.5283
14. Díaz-Pontones, D., & Reyes-Jaramillo, I. (2009). Producción y almacenamiento de bellotas de Quercus hintonii Warburg (Fagaceae) de la Depresión del Balsas, México. Polibotánica27, 131–143.

Blog Announcements

  1. Submit your trees for critique or advice here. I need new trees for the next Bonsai Buds episode! Guest announcement TBD.
  2. Contact me if you ordered seeds from me last year and they did not germinate. As you can see, I've been doing much more research on long-term seed storage and germination and am testing to ensure higher, more uniform rates of germination this year. I'm happy to send new seeds at no charge.
  3. New seeds types with my 10-year bonsai growing guide will be on sale in March.
  4. On behalf of the Columbus Bonsai Society, I'm putting together an Yamadori Study Group. Those of you interested in joining and who live nearby can read more and fill out this brief survey here.

No comments:

Post a Comment

ADBLOCKER DETECTED
It looks like you're using an ad blocker. That's okay, sometimes I do too.
Please disable your ad blocker and reload the page to support our work making free bonsai content for readers like you.
Reload Page
Powered By - BlogAntiAdblock