Publication Date
2022
Document Type
Thesis
Committee Members
Michael Leffak, Ph.D. (Advisor); Mike Kemp, Ph.D. (Committee Member); Michael Markey, Ph.D. (Committee Member)
Degree Name
Master of Science (MS)
Abstract
Microsatellite repeat sequences have been shown to induce replication stalling, fork collapse, double-strand breaks (DSBs), and possibly stimulate break-induced replication. In this study we use a dual-fluorescent HeLa model that is designed to monitor recombination at an ectopic site through use of flow cytometry and inverse PCR with a microsatellite in the lagging strand for DNA synthesis. To test the stability of the 78 bp polypurine/pyrimidine repeat from the PDK1 locus, we subjected cells to replication stress drugs designed to induce DSBs and measure break-induced replication (BIR). The study revealed that polypurine repeat cells undergo endogenous stress contributing to instability at the ectopic site as well as slow cell growth. Further, we show that there is an orientation dependency for instability with the (Pu)78 cells being more unstable. Lastly, we present a novel candidate for a protein involved in break-induced replication, COPS2. Preliminary experiments show this protein produces unique recombination patterns when knocked down.
Page Count
77
Department or Program
Department of Biochemistry and Molecular Biology
Year Degree Awarded
2022
Copyright
Copyright 2022, some rights reserved. My ETD may be copied and distributed only for non-commercial purposes and may not be modified. All use must give me credit as the original author.
Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License.
ORCID ID
0000-0002-5595-5491
