"This season, TACF and affiliated scientists are working to clone and preserve pollen from a handful of wild American chestnut trees which have been observed to contain genetic resistance to the chestnut blight fungus. Out of an estimated 4 billion American chestnut trees which once populated the Appalachian Mountains in the late 1800s, only 24 trees have been identified to be what scientists call 'Large, Surviving American chestnuts' or LSAs. The 'S' for surviving doesn't just mean that they are alive, but that they are 'surviving with chestnut blight infection'. There are a few thousand 'large American chestnuts', but they tend to be 'Lucky' instead of 'Surviving'. Once those Large Lucky American chestnut trees get hit with the blight, they tend to succumb in as little as 1-5 years whereas the Large Surviving American chestnuts are documented to have survived multiple decades with chestnut blight infection.

Some of the better known LSA trees are the Amherst tree in VA, the Ort tree in PA, and the Adair County tree in Kentucky. To clone these trees, grafters at multiple locations are using scion wood collected from the LSA trees and then grafting them onto some manner of rootstock. This grafting process creates an exact genetic replica. The clones can then be used in various types of experiments.

The primary experiment TACF is aiming to instigate with these LSAs is something called 'RNASeq'. There are several steps to that experiment. First, the DNA and RNA from all the grafted trees (the clones) will be fully sequenced. Then, the clones will be infected with the chestnut blight fungus and their RNA will be isolated and fully sequenced right after the infection with blight. Finally, the RNA will be fully sequenced again at a few days following infection. This sequence of RNA Sequencing, hence the name 'RNA Seq', will be key to better understanding the underlying genetic mechanisms of disease resistance in these LSAs.

The grafting process is taking place in 2022. Once the grafts are well-established, the RNA Seq will take place in 2023 or 2024.

In addition to the cloning and RNA Seq experiment, pollen from these LSAs will be collect to use both this season and additional pollen will be frozen for long-term storage and use. Using that pollen, the LSAs can be bred together.

The resulting seeds and seedlings can then be used in additional experiments to better understand the underlying mechanisms of blight-resistance in these unique trees. That breeding and the subsequent experiments will take place across the next 5-10 years."