Explorations in Real-Time Polymerase Chain Reactions
Arts and Sciences
Department of Chemistry
Polymerase Chain Reaction (PCR) is a method by which a specific segment of DNA can be replicated for further analysis or to ascertain the presence of that sequence. Real-time (or quantitative) PCR allows researchers to view the amplification of the DNA segments both quantitatively and in real time, providing faster results and an idea of the process and rate of the reaction. This real-time information is gathered using fluorescence detection, as fluorophores attach to the amplifying, double stranded DNA segments. The fluorescence detected is proportional to the amount of DNA replicated. We have used both conventional and real-time PCR to test the identity and genetic composition of various food samples, and to learn more about the process of PCR and its uses in research. First, we used a BioRad Fish DNA Barcode kit to test the identity of five fish samples. From this experiment, only one sample was successfully amplified and sent for sequencing. Next, we used a kit that tests for the presence of genetically modified genes in plants (GMO Test kit), also from BioRad, to test commercially available corn-based snack foods for the presence of GMOs, using SYBRgreen as the fluorophore for the real-time PCR reaction. The results from this genetically modified food experiment positively identified GMO genes in a DoritosTM sample, and gave expected control values which indicated a reliable analysis. In the future, we will use these tests as the basis for developments of new protocols for use in the classroom or in research.
Mammoser, Claire C., "Explorations in Real-Time Polymerase Chain Reactions" (2015). Symposium on Undergraduate Research and Creative Expression (SOURCE). 419.