Major
Physics
Anticipated Graduation Year
2024
Access Type
Open Access
Abstract
Proper three-dimensional organization of the genome is required for cells to coordinate and control gene activity. Supercoiling is a critical process in compacting DNA into tightly wound structures to organize the genome. Formation of DNA mini circles via looping is an important, early step in this process. Here, we are working to develop a single-molecule fluorescent assay to monitor DNA loop formation in real-time. We are using polymerase chain reaction (PCR) to generate dye-labeled constructs of different sequences. High and low levels of NaCl are used to induce looped and unlooped formations of the DNA that give distinct high and low FRET signals, respectively. Non-helical regions associated with genetic disorders, such as spinocerebellar ataxia and ALS, disrupt genomic organization. With this assay, we aim to track looping activity in real time and study how defects within the helical domain associated with such genetic disorders affect the looping process and how it can lead to genomic disorganization.
Faculty Mentors & Instructors
Brian Cannon, Associate Professor, Department of Physics; Cole Geinosky, Research Assistant, Department of Physics
Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License.
Single Molecule DNA Looping Assay
Proper three-dimensional organization of the genome is required for cells to coordinate and control gene activity. Supercoiling is a critical process in compacting DNA into tightly wound structures to organize the genome. Formation of DNA mini circles via looping is an important, early step in this process. Here, we are working to develop a single-molecule fluorescent assay to monitor DNA loop formation in real-time. We are using polymerase chain reaction (PCR) to generate dye-labeled constructs of different sequences. High and low levels of NaCl are used to induce looped and unlooped formations of the DNA that give distinct high and low FRET signals, respectively. Non-helical regions associated with genetic disorders, such as spinocerebellar ataxia and ALS, disrupt genomic organization. With this assay, we aim to track looping activity in real time and study how defects within the helical domain associated with such genetic disorders affect the looping process and how it can lead to genomic disorganization.