By Kendra Gurney
TACF® New England Region Science Coordinator

In spring of 2009 Dr. Fenny Dane, a professor at Auburn University, presented her research on the evolutionary history of Castanea species at CT-TACF's Annual Meeting. Dr. Dane has used the DNA of chloroplasts, small organelles within the plant cell, to trace populations of Castanea species over millions of years. The study of chloroplast-DNA is a common tool for plant scientists interested phylogeography because the DNA of the chloroplast is effectively haploid (only has one set of chromosomes) and inherited maternally. This provides the ability to trace seed-mediated migration and colonization of plant populations over time. Differences in chloroplast DNA can be used to designate haplotypes, or different genetic lineages.

It has been shown that each Castanea species has several distinct chloroplast types. More differences can be detected among Asian species than within European or North American species using chloroplast DNA analysis. Dr. Dane hypothesizes that the genus originated in Asia, migrated to Europe and then continued west to North America. From an analysis of samples from the Appalachian range of the US we can hypothesize that Castanea found refuge in the southern region of the Appalachian Mountains during glacial periods. Following the retreat of the ice sheet, one lineage, the C. dentata D1 haplotype, migrated in a north-easterly direction.

Much of Dr. Dane's work with Castanea in North America has focused on samples from the southern and central Appalachians, where more diversity in chloroplast type has been detected. Dr. Dane suggested that it would be helpful to her work if the CT Chapter could provide her with samples of genetic material from known, validated Castanea dentata (American chestnut) examples from Connecticut. Currently, such specimens are validated by scientists who use a dichotomous key to analyze a sample for traits indicating C. dentata lineage. Positive validation leads to the selection of mother trees appropriate for inclusion in the CT chapter's breeding program. Analyzing the chloroplast DNA of validated C. dentata samples from CT would benefit Dr. Dane's work by providing more samples to support the single north-easterly linage hypothesis. Such an analysis would also benefit the CT Chapter, by verifying that the current process of sample validation for selecting mother trees is in fact identifying pure C. dentata from the local population. This past summer, CT Chapter President Bill Adamsen took on the task of collecting samples for Dr. Dane from the chapter's mother trees, as well as the backcross offspring that have been produced. His total journey was just over three hundred miles and he collected a total of collection totaled thirty five mother tree samples and twelve orchard back-cross samples.

Thirty three of those samples collected were submitted to Dr. Dane (some samples were omitted because of collection timing issues) and the results are in! Her graduate student, Xiaowei Li, isolated DNA from the leaf samples and amplified small sections of the chloroplast genome using the polymerase chain reaction, followed by sequencing analysis. All samples showed the C. dentata D1 haplotype, most commonly detected in populations of the American chestnut (see Figures). This means that all samples are from the linage that travelled north-east following the retreat of the ice sheet. The analysis cannot pin-point the origin of the trees any further, but does provide evidence that the mother trees used in the CT Chapter's breeding program are, in fact, Castanea dentata, or American chestnut, which is a nice validation of the careful selection process used.

For more information see: Dane, F. 2009. Comparative phylogeography of Castanea species. Proceedings of the Fourth International Chestnut Symposium. Acta Horticulturae 844: 211-222.

Home page of Fenny Dane at Auburn

Article by Kendra Gurney
The American Chestnut Foundation®
New England Regional Science Coordinator
USFS Northern Research Station
705 Spear Street
South Burlington, VT 05403
Tel: 802.951.6771 x1350
Cell: 802.999.8706
Kendra@acf.org