Thermal characterization of DNA phase change phenomena at the microscale
Abstract/Contents
- Abstract
- There are continuing efforts within the biological community to develop new and advanced DNA characterization techniques that use reduce sample volumes, increase throughput during detection of reactions and processes, and optimize characterization methods. Such scaling efforts introduce new challenges as many of these techniques rely on fluorescent detection methods, which become limited due to drastic reductions in fluorescence signal. This limitation requires the development of alternative detection schemes. Due to the important role of thermal phenomena in DNA reaction processes, we investigate the capability of utilizing thermal characterization of small volume fluid samples to detect DNA melt curves for sequencing and genotyping purposes. Additionally, we explore the use of frequency-domain thermal metrology to simultaneously probe DNA reaction kinetics during melt curve detection. Thermal detection offers a potential DNA characterization alternative that does not suffer from the same scaling limitations as more traditional techniques. In this work, we first apply the 3 [omega] measurement technique, an established thermal property characterization methodology, to characterize thermal properties of µL volumes by using heaters with 200-700 [mu] m length and 2-5 [mu] m width. Sensitivity and uncertainty analyses show the ability of these devices to uniquely and simultaneously extract thermal conductivity and heat capacity to sub-percent accuracy levels. We investigate fluid samples of deionized (DI) water, silicone oil, and a salt buffer solution to experimentally determine their temperature-dependent thermal properties from 25°C to 80 °C. We also conduct temperature-dependent measurements down to pL droplet volumes based on an analysis of the actual thermal volume probed in the experiments, independent of droplet boundary conditions, and offer insight into the tradeoffs between volume, sensitivity, and operational frequency regime, which are essential to consider in optimal device design for each application. Secondly, we apply the 3 [omega] technique to investigate phase change phenomena and the frequency-dependent specific heat of DNA melt transitions from double-stranded to single-stranded DNA. We incorporate a relaxation function to simultaneously probe the relaxation times of the DNA reaction kinetics in the linear response regime of the measurement. We then compare these relaxation times resolved by our frequency-domain thermal spectroscopy to previous works' empirical data, achieved with intricate laser temperature-jump measurement systems. We also demonstrate a high-throughput melt curve detection scheme, achieved by focusing on the single-frequency out-of-phase temperature component of the 3 [omega] experimental data in the temperature domain. Lastly, we present an extension of the 3 [omega] method to exploit the nonlinear temperature response exhibited by the DNA melting and annealing processes when amplifying the temperature excursion, which may provide a novel high throughput method to detect the presence of complementary DNA segments.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2015 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Roy, Shilpi |
|---|---|
| Associated with | Stanford University, Department of Electrical Engineering |
| Primary advisor | Goodson, Kenneth E, 1967- |
| Thesis advisor | Goodson, Kenneth E, 1967- |
| Thesis advisor | Poon, Ada Shuk Yan |
| Thesis advisor | Pop, Eric |
| Advisor | Poon, Ada Shuk Yan |
| Advisor | Pop, Eric |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Shilpi Roy. |
|---|---|
| Note | Submitted to the Department of Electrical Engineering. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2015. |
| Location | https://purl.stanford.edu/rh889md8035 |
Access conditions
- Copyright
- © 2015 by Shilpi Roy
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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