Viability of Magnolia seed at -20C or -196C

[Melissa Spearing]

Hello there CPC – happy to have this wonderful resource and expertise knowledge base available.

I’m working at the National Tree Seed Centre in Canada and drafting a protocol note for a special Applications in Plant Science issue I’d like to ask anyone for additional results to compare or cite. I’m looking for any arboreta or seed banks that have been able to successfully or routinely store and recover Magnolia acuminata seedlings from either -20C freezer storage >3 months or -196C cryogenic storage. I’m hoping there are more results in the United States where it is not a rare species as it is here (federally listed as endangered, has been since the 1980s); its very hard to get much viable seed from our wild populations.

Of interest, in 2019, we had 11 single-tree seed collections of Magnolia acuminata come to our centre from the Guelph Arboretum’s gene bank collection. We also had Guelph Arboretum bulk seed that had not yet been tested from 2017 acuminata and M. tripetala in -20C at <5% moisture content. Evidence from our earlier germination testing records, Kew’s Seed Information Database, and consulting with a number of expert magnolia growers/seed exchanges, everyone suggested it was “impossible” to dry and freeze this seed, in spite of repeated Woody Plant Seed Manuals stating eastern US Magnolia species are orthodox. I was also curious whether the 1937 paper noting storing “dry” seeds with the sarcotesta on actually retains higher humidity in the seed or repels fungi vs cleaning it off – he did not do oven moisture content tests to define “dry” by today’s standards!

So out of curiosity, I first imbibed our 2017 stored seedlots in -20C and found 30% of the acuminata embryos stained with tetrazolium, though 100% of the endosperm did not; we were able to grow a few normal embryos on woody plant medium and agar after removing the seed coat – regular germination on Kimpak resulted in whole seeds rotting before embryos could grow. Better yet, the 2017 tripetala seedlot stained at 70% viable embryos and we recovered more of those that acclimated to the greenhouse in 2020, including a few from Kimpak with seed coat on. But its still not definitive (to me, by existing original studies) whether cucumber as a whole is orthodox, desiccation sensitive/intermediate as some still call that category, or if certain parents impart better orthodox seed behaviour than others.

Back to the 2019 single-tree seedlots: we cleaned and put fresh seed into damp peat for 7 months stratification in the cooler as a standard procedure control. We germinated according to the 1937 26C/15C treatment saw large variability in germination rates Summer 2020 of 1-97%. Of the largest four seedlots, we also dried and split fractions into slow drying vs fast drying methods over lithium chloride solutions before freezing at -20C and -196C. We destructively tested samples to determine a seed moisture desorption isotherm and ensure all seedlots were equilibrated to ~20% eRH / <6% MC before going into -20C and -196C.

Due to COVID, I did not get to finish my 2019 stored seed germination tests to compare to fresh, but hope to finish that this fall, if its equal or close to fresh by either treatment. Regardless of the complexity of my inquiry, tts been a fun side-of-my-desk experiment so far and we have 3x the number of seedlings as exist in the wild in Ontario. We hope to return these in a balanced parental mix to declining populations once they are big enough to be transported/transplanted.

I welcome comments, other research papers I may have missed or discussion here or you can email melissa.spearing@canada.ca

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