Presenter: Anna Hickey
Faculty Mentor: Victoria DeRose, Geri Richmond
Presentation Type: Poster 69
Primary Research Area: Science
Major: Biochemistry
Funding Source: Presidential Undergraduate Research Scholars Program, University of Oregon, $5,000.00 research stipend; Scholarships for Oregon Scientists, University of Oregon and National Science Foundation, $2,000.00 research stipend
Cisplatin is a commonly used anti-cancer therapeutic; however, its mechanism of inducing cell death is not well understood. In order to identify and isolate cisplatin’s cellular targets for characterization, our lab utilizes the
“click” reaction (a physiologically stable and high yielding reaction that produces no harmful byproducts) to attach fluorescent compounds or other small molecules to platinated cellular targets such as DNA, RNA, and proteins. In this project, I optimized an in vitro pull-down procedure using streptavidin-coated magnetic beads to separate platinated cellular targets from unplatinated molecules. I first treated target DNA with a click-functionalized platinum reagent, then clicked that compound to a double-stranded DNA linker. The opposite end of this linker contains a biotin molecule, which interacts strongly with the streptavidin-coated magnetic beads through the streptavidin-biotin interaction. Using a powerful magnet, I separated platinated and clicked DNA attached to the beads from unreacted DNA, then confirmed the desired species of DNA was pulled down using polyacrylamide gel electrophoresis (PAGE), a method by which DNA or proteins can be separated by size. I determined that increasing the incubation time of the beads with the platinated DNA increased elution yields. Furthermore, elution temperatures above 90° C also increase the elution yield. Optimizing this pull-down technology will allow us to better characterize platinated molecules, and will ultimately improve our understanding of cisplatin’s cell-death inducing mechanisms.