Electrochemical Detection of DNA Methylation and Implications for Detection of Cancer Onset Resulting from Hypermethylation
Gonzalez, Samantha Dawn
Epigenetics refers to the process by which genetic material is modified without changing the base coding sequence1. One such epigenetic process is DNA methylation, employed by cells to regulate gene expression, often by silencing transcription, which subsequently impedes the expression of targeted genes2. Regulation via DNA methylation is a vital process in facilitating normal development. Abnormally high levels of DNA methylation within cells (ie. hypermethylation) can lead to disease, such as the development of various forms of cancer3. For instance, hypermethylation in promoter regions coding for tumor suppressor genes results in deactivation of these regulatory genes, affecting downstream cellular processes, and allowing diseased cells to proliferate2. One strategy for the early detection of cancer is to assess the amount of abnormal methylation at certain genomic loci1-3. This is typically performed via the use of various techniques, such as quantitative polymerase chain reaction (PCR) and reporters of genomic methylation (RGM)4-5. While these processes provide a significant amount of information, they demand substantial amounts of time, labor, and materials. Electrochemical sensor-based methods to detect DNA and DNA-related processes have the ability to remedy these drawbacks6. Here, we developed an electrochemical sensor to monitor DNA methylation in DNA oligomers of known lengths and sequences with varying levels of methylation. Gold electrodes were modified with 21-mer single stranded DNA (ssDNA) and then hybridized with complementary strands, creating surface bound double stranded DNA (dsDNA). The complementary sequences were modified containing up to six 5-methyl cytosine (5-mC) locations. Electrochemical detection of DNA was accomplished utilizing an electrochemically active di-viologen compound, known as C12Viologen, that features differential binding to DNA based on conformation, which is altered by methylation8. Cytosine methylation alters the structure of DNA in solution, which alters the binding of the C12Viologen compound in the oligomers9. These changes in binding were detected using square wave voltammetry (SWV) by monitoring the differences in peak potentials upon exposure to high ionic strength conditions, which forces changes in DNA structure based on 5-mC content9. We show that the electrochemical data demonstrates significant differences among unmethylated and methylated DNA samples.
Gonzalez, Samantha Dawn. (May 2017). Electrochemical Detection of DNA Methylation and Implications for Detection of Cancer Onset Resulting from Hypermethylation (Honors Thesis, East Carolina University). Retrieved from the Scholarship. (http://hdl.handle.net/10342/6234.)
Gonzalez, Samantha Dawn. Electrochemical Detection of DNA Methylation and Implications for Detection of Cancer Onset Resulting from Hypermethylation. Honors Thesis. East Carolina University, May 2017. The Scholarship. http://hdl.handle.net/10342/6234. September 29, 2020.
Gonzalez, Samantha Dawn, “Electrochemical Detection of DNA Methylation and Implications for Detection of Cancer Onset Resulting from Hypermethylation” (Honors Thesis., East Carolina University, May 2017).
Gonzalez, Samantha Dawn. Electrochemical Detection of DNA Methylation and Implications for Detection of Cancer Onset Resulting from Hypermethylation [Honors Thesis]. Greenville, NC: East Carolina University; May 2017.
East Carolina University