DeRosa and Regan worked in Dr. Powell's lab under the supervision of technician Kathleen Baier and graduate student Kristin Stewart. DeRosa performed DNA extractions and Multiplex PCR (Polymerase Chain Reactions) on leaves from Transgenic American Chestnut trees in the field to help determine the inheritance patterns of two genes, one found to enhance disease resistance and the other a marker gene. Regan analyzed the stability of transgene expression in three-year-old transgenic American chestnut trees using a variation of PCR that is computer-based and quantitatively compares the differences in RNA levels for specific genes.
Buell explained how long the process is – and how very careful and sterile one must be. His summer was spent putting seeds into culture or medium. Those seeds will grow over the winter, and then by next summer, someone else “will be able to perform vector transformations on them.”
Lovier explained: “Transformations are one way to genetically engineer living things. Basically, transformation involves using bacteria, specifically a type called Agrobacterium, that have already been genetically altered to have the genes that we want and allowing the bacteria to give the new genes to the plant for us. Agrobacterium and related bacteria in nature causes disease in plants by giving them some of their DNA, which doesn't mix well with the plants' own genome and tends to be pretty bad for them. But what scientists can do is change out the DNA that the bacteria give so instead of harming the plant, it actually helps them.”
Lovier said that she, Buell and Lane mixed the Agro and the plant tissue they wanted to transform, usually small pieces of leaf, by soaking the pieces in a liquid that had been used to grow the bacteria. “Because leaves have plenty of surface area, a lot of the cells in the tissue can be given the new genes at a time. After the pieces are taken out of the liquid, the pieces are allowed to grow in tissue culture, where the transformed cells can multiply and spread the genes throughout the piece of leaf. Eventually, all the cells have the new DNA and whatever benefit it brings.”