Photo by The Ozark Chinquapin Foundation American Chestnut (Castanea dentata)

Allegheny Chinquapin (Castanea pumila)

Ozark Chinquapin (Castanea ozarkensis) The Potential of Biotechnology for Ozark Chinquapin Conservation Hannah Pilkey; SUNY-College of Environmental Science & Forestry Blight Tolerant American Chestnut “Darling 58”

Graph courtesy of Andy Newhouse, SUNY-ESF Research Objective

Can the same methods used to produce a transgenic American chestnut, be used to develop a transgenic Ozark chinquapin tree? Regenerating Ozark Chinquapin Embryos

Somatic embryo mass Shoots emerging from Micropropagation “OC001-14” somatic embryos of shoots Oxalate Oxidase (OxO) Gene From Wheat

Ubiquitous , found in many plants

Detoxifies the oxalic acid produced by the fungus

NOT a pesticide, does not kill the fungus

Changes the fungus’ lifestyle from a pathogen, to a saprotroph (like on Chinese chestnut & some oaks)

Slide by Bill Powell Genetic Transformation

• Agrobacterium (AGL1) -mediated transformation

• p35s-OxO binary vector

• “OC001-14” somatic embryos

• 10-week period of selection in bioreactors OxO Detected in Ozark Chinquapin Embryos

100 bp 100 bp

+ AC + OC (Darling) wt AC wt OC Natural Blight-Resistance in Ozark Chinquapin

Research by Leslie Bost https://ozarkchinquapinmembership.org/blight-screening/ Oxalic Acid Leaf Disc Assay Final Thoughts

Does Ozark chinquapin have a gene similar to OxO?

Other enzymatic pathways? • Oxalate-CoA • Oxalyl-CoA decarboxylase • Formyl-CoA • Formate dehydrogenase

Genes can be put into the genome of Ozark chinquapin if it is needed in the future, but protocol should be optimized to increase number of transformants. Thank you!

Dr. Scott Merkle

Steve Bost and The Ozark Chinquapin Foundation

The American Chestnut Foundation Questions? www.esf.edu/chestnut Contact: [email protected]