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- All Subjects: Health
- All Subjects: Synthetic Biology
- Creators: Harrington Bioengineering Program
- Member of: Theses and Dissertations
Description
This thesis project is the result of close collaboration with the Arizona State University Biodesign Clinical Testing Laboratory (ABCTL) to document the characteristics of saliva as a test sample, preanalytical considerations, and how the ABCTL utilized saliva testing to develop swift COVID-19 diagnostic tests for the Arizona community. As of April 2021, there have been over 130 million recorded cases of COVID-19 globally, with the United States taking the lead with approximately 31.5 million cases. Developing highly accurate and timely diagnostics has been an important need of our country that the ABCTL has had tremendous success in delivering. Near the start of the pandemic, the ABCTL utilized saliva as a testing sample rather than nasopharyngeal (NP) swabs that were limited in supply, required highly trained medical personnel, and were generally uncomfortable for participants. Results from literature across the globe showed how saliva performed just as well as the NP swabs (the golden standard) while being an easier test to collect and analyze. Going forward, the ABCTL will continue to develop high quality diagnostic tools and adapt to the ever-evolving needs our communities face regarding the COVID-19 pandemic.
ContributorsSmetanick, Jennifer (Author) / Compton, Carolyn (Thesis director) / Magee, Mitch (Committee member) / School of Life Sciences (Contributor) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
Currently in synthetic biology only the Las, Lux, and Rhl quorum sensing pathways have been adapted for broad engineering use. Quorum sensing allows a means of cell to cell communication in which a designated sender cell produces quorum sensing molecules that modify gene expression of a designated receiver cell. While useful, these three quorum sensing pathways exhibit a nontrivial level of crosstalk, hindering robust engineering and leading to unexpected effects in a given design. To address the lack of orthogonality among these three quorum sensing pathways, previous scientists have attempted to perform directed evolution on components of the quorum sensing pathway. While a powerful tool, directed evolution is limited by the subspace that is defined by the protein. For this reason, we take an evolutionary biology approach to identify new orthogonal quorum sensing networks and test these networks for cross-talk with currently-used networks. By charting characteristics of acyl homoserine lactone (AHL) molecules used across quorum sensing pathways in nature, we have identified favorable candidate pathways likely to display orthogonality. These include Aub, Bja, Bra, Cer, Esa, Las, Lux, Rhl, Rpa, and Sin, which we have begun constructing and testing. Our synthetic circuits express GFP in response to a quorum sensing molecule, allowing quantitative measurement of orthogonality between pairs. By determining orthogonal quorum sensing pairs, we hope to identify and adapt novel quorum sensing pathways for robust use in higher-order genetic circuits.
ContributorsMuller, Ryan (Author) / Haynes, Karmella (Thesis director) / Wang, Xiao (Committee member) / Barrett, The Honors College (Contributor) / School of Mathematical and Statistical Sciences (Contributor) / Department of Chemistry and Biochemistry (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
MPV17-related hepatocerebral mitochondrial DNA depletion syndrome, previously known as Navajo Neurohepatopathy (NNH), is a rare genetic disease affecting Navajo children of the American Southwest. These children can suffer from several severe symptoms like brain damage and liver disease, and a diagnosis leads to death by age 10, on average. The only known effective therapy for NNH is a liver transplant. Currently, the disease is diagnosed through a lengthy and expensive process of gene sequencing, but oftentimes patients with the most severe forms of NNH deteriorate quickly; thus a rapid diagnostic would be beneficial to beginning the transplant process as early as possible. Here, Tentacle Probes, a novel technology to detect genetic mutations, were proposed to rapidly and accurately diagnose NNH. Because of Tentacle Probes' double binding site kinetics, they can detect mutations more accurately than other types of genetic probes. Probes specific to the NNH mutation were designed for use with a real-time polymerase chain reaction (PCR) detection platform. Initial synthetic DNA testing of Tentacle Trobes showed capable differentiation between mutated and non-mutated samples. However, experiments to validate those results at Phoenix Children's Hospital before moving to patient samples showed that test viability decreased over time. Efforts to diagnose the issues that led to decreased viability suggested four possible explanations that are as follows (in order of decreasing likelihood): first, undesired products from improper PCR primer design was supported by double bands in DNA gel electrophoresis; second, DNA may have degraded over time or due to repeated cycles of freezing and thawing stock solutions, and this was supported by smeared DNA gel electrophoresis; third, probe degradation, specifically of the fluorescent reporter, is possible; finally, contaminants that inhibit the PCR reaction may have been introduced. A combination of these factors may also have caused the change in assay viability. As a result of these most likely possibilities, new primers were designed and steps suggested to return viability to the assay. Thus, the various limitations and requirements for this Tentacle Probe diagnostic have been identified, and as assay development continues following the promising initial results achieved, we are confident that a rapid method if diagnosing NNH is on its way to help the children afflicted with this devastating disease receive timely access to treatment.
ContributorsThompson, Emily Rose (Author) / Caplan, Michael (Thesis director) / Carpentieri, David (Committee member) / School of Mathematical and Statistical Sciences (Contributor) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
Description
The main objective of this research is to develop and characterize a targeted contrast agent that will recognize acute neural injury pathology (i.e. fibrin) after traumatic brain injury (TBI). Single chain fragment variable antibodies (scFv) that bind specifically to fibrin have been produced and purified. DSPE-PEG micelles have been produced and the scFv has been conjugated to the surface of the micelles; this nanoparticle system will be used to overcome limitations in diagnosing TBI. The binding and imaging properties will be analyzed in the future to determine functionality of the nanoparticle system in vivo.
ContributorsRumbo, Kailey Michelle (Author) / Stabenfeldt, Sarah (Thesis director) / Kodibagkar, Vikram (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2014-05
Description
Transgene expression in mammalian cells has been shown to meet resistance in the form of silencing due to chromatin buildup within the cell. Interactions of proteins with chromatin modulate gene expression profiles. Synthetic Polycomb transcription factor (PcTF) variants have the potential to reactivate these silence transgenes as shown in Haynes & Silver 2011. PcTF variants have been constructed via TypeIIS assembly to further investigate this ability to reactive transgenes. Expression in mammalian cells was confirmed via fluorescence microscopy and red fluorescent protein (RFP) expression in cell lysate. Examination of any variation in conferment of binding strength of homologous Polycomb chromodomains (PCDs) to its trimethylated lysine residue target on histone three (H3K27me3) was investigated using a thermal shift assay. Results indicate that PcTF may not be a suitable protein for surveying with SYPRO Orange, a dye that produces a detectable signal when exposed to the hydrophobic domains of the melting protein. A cell line with inducible silencing of a chemiluminescent protein was used to determine the effects PcTF variants had on gene reactivation. Results show down-regulation of the target reporter gene. We propose this may be due to PcTF not binding to its target; this would cause PcTF to deplete transcriptional machinery in the nucleus. Alternatively, the CMV promoter could be sequestering transcriptional machinery in its hyperactive transcription of PcTF leading to widespread down-regulation. Finally, the activation domain used may not be appropriate for this cell type. Future PcTF variants will address these hypotheses by including multiple Polycomb chromodomains (PCDs) to alter the binding dynamics of PcTF to its target, and by incorporating alternative promoters and activation domains.
ContributorsGardner, Cameron Lee (Author) / Haynes, Karmella (Thesis director) / Stabenfeldt, Sarah (Committee member) / Barrett, The Honors College (Contributor) / Department of Finance (Contributor) / Harrington Bioengineering Program (Contributor)
Created2015-05
Description
Markov Chain Monte-Carlo methods are a Bayesian approach to predictive statistics, which takes advantage of prior beliefs and conditions as well as the existing data to produce posterior distributions of relevant parameters. This approach, implementable through the JAGS packaging in R, is promising for its impact on the diagnostics space, which is a critical bottleneck for pandemic planning and rapid response. Specifically, these methods provide the means to optimize diagnostic testing, for example, by determining whether it is best to test individuals in a certain locale once or multiple times. This study compares the expected accuracy of single and double testing under two specific conditions, a general and Icelandic test case, in order to ascertain the validity of MCMC methods in this space and inform decisionmakers and future research in the space. Models based on this platform may eventually be tailored to the priors of specific locales. Additionally, the ability to test multiple regimes of real or simulated data while maintaining uncertainty widens the pool of researchers that can impact the space. In future studies, ensemble methods investigating the full range of parameters and their combinations can be studied.
ContributorsSuresh, Tarun (Author) / Naufel, Mark (Thesis director) / Panchanathan, Sethuraman (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
The dilemma of the lack of prenatal and neonatal healthcare has been prevalent among third world countries for many years. The lack of prenatal healthcare has been shown to have direct links to spontaneous preterm births from which low-birth weight in babies can be a result. The World Health Organization has identified preterm birth as one of the biggest overseen burdens in developing countries.
This study seeks to answer the research questions: What are the major risk factors associated with the lack of prenatal and neonatal healthcare in developing countries? What are potential routes of intervention (ROI) to help these countries? The goal is to analyze the risk factors and determine if there are any ROIs available to minimize potential incidents or accidents associated with complications of preterm birth.
A few potential risk factors include: poverty, a mother’s lack of education, a lack of professional visitation during pregnancy, having a short cervix, and routine use of Ultrasound. This research paper has identified that keeping ultrasound diagnostics to a minimum, seeking professional help during pregnancy, incorporating corticosteroids for preterm births, implementing Kangaroo Mother Care, and Cervical Cerclage are interventions that can reduce preterm births and the associated complications that come with it. We believe that further research, regarding compliance of each of these interventions, would show reduction of preterm births and low birth weight in developing countries.
This study seeks to answer the research questions: What are the major risk factors associated with the lack of prenatal and neonatal healthcare in developing countries? What are potential routes of intervention (ROI) to help these countries? The goal is to analyze the risk factors and determine if there are any ROIs available to minimize potential incidents or accidents associated with complications of preterm birth.
A few potential risk factors include: poverty, a mother’s lack of education, a lack of professional visitation during pregnancy, having a short cervix, and routine use of Ultrasound. This research paper has identified that keeping ultrasound diagnostics to a minimum, seeking professional help during pregnancy, incorporating corticosteroids for preterm births, implementing Kangaroo Mother Care, and Cervical Cerclage are interventions that can reduce preterm births and the associated complications that come with it. We believe that further research, regarding compliance of each of these interventions, would show reduction of preterm births and low birth weight in developing countries.
ContributorsHuapaya, Eduardo Luciano (Author) / Muthuswamy, Jitendran (Thesis director) / Comar, William (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
The primary goal of this paper is to analyze a tool used for measuring human
development on a global scale. Originally, development within a country was solely judged by the degree of economic growth by way of Gross National Product (GNP) and per capita income. Holistically, GNP measures the total extent of economic activity of a country’s people within a given time period. (Rutherford, 2012). Critics found several issues with this one-dimensional approach of measuring human development. What failed to be recognized was the distribution of income among the country’s citizens. Higher incomes often favor men within the majority when compared to women and people of minority groups (Feiner & Roberts, 1990). GNP also failed to recognize the social limitations under a government. In other words, are there limitations as to what goods can be bought and who can buy them?
development on a global scale. Originally, development within a country was solely judged by the degree of economic growth by way of Gross National Product (GNP) and per capita income. Holistically, GNP measures the total extent of economic activity of a country’s people within a given time period. (Rutherford, 2012). Critics found several issues with this one-dimensional approach of measuring human development. What failed to be recognized was the distribution of income among the country’s citizens. Higher incomes often favor men within the majority when compared to women and people of minority groups (Feiner & Roberts, 1990). GNP also failed to recognize the social limitations under a government. In other words, are there limitations as to what goods can be bought and who can buy them?
ContributorsSteeley, Emily Anne (Author) / Parmentier, Mary Jane (Thesis director) / Grgich, Traci (Committee member) / Harrington Bioengineering Program (Contributor) / School of Human Evolution & Social Change (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
The global population over the age of 60 is estimated to rise to 23% by 2050 only increase the prevalence of functional neurological disorders and stroke. Increase in cases of functional neurological disorders and strokes will place a greater burden on the healthcare industry, specifically physical therapy. Physical therapy is vital for a patient’s recovery of motor function which is time demanding and taxing on the physical therapist. Wearable robotics have been proven to improve functional outcomes in gait rehabilitation by providing controlled high dosage and high-intensity training. Accurate control strategies for assistive robotic exoskeletons are vital for repetitive high precisions assistance for cerebral plasticity to occur.
This thesis presents a preliminary determination and design of a control algorithm for an assistive ankle device developed by the ASU RISE Laboratory. The assistive ankle device functions by compressing a spring upon heel strike during gait, remaining compressed during mid-stance and then releasing upon initiation of heel-off. The relationship between surface electromyography and ground reactions forces were used for identification of user-initiated heel-off. The muscle activation of the tibialis anterior combined with the ground reaction forces of the heel pressure sensor generated potential features that will be utilized in the revised control algorithm for the assistive ankle device. Work on this project must proceed in order to test and validate the revised control algorithm to determine its accuracy and precision.
This thesis presents a preliminary determination and design of a control algorithm for an assistive ankle device developed by the ASU RISE Laboratory. The assistive ankle device functions by compressing a spring upon heel strike during gait, remaining compressed during mid-stance and then releasing upon initiation of heel-off. The relationship between surface electromyography and ground reactions forces were used for identification of user-initiated heel-off. The muscle activation of the tibialis anterior combined with the ground reaction forces of the heel pressure sensor generated potential features that will be utilized in the revised control algorithm for the assistive ankle device. Work on this project must proceed in order to test and validate the revised control algorithm to determine its accuracy and precision.
ContributorsGaytan-Jenkins, Daniel Rinaldo (Author) / Zhang, Wenlong (Thesis director) / Tyler, Jamie (Committee member) / Harrington Bioengineering Program (Contributor, Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
Cell fate is a complex and dynamic process with many genetic components. It has often been likened to “multistable” mathematical systems because of the numerous possible “stable” states, or cell types, that cells may end up in. Due to its complexity, understanding the process of cell fate and differentiation has proven challenging. A better understanding of cell differentiation has applications in regenerative stem cell therapies, disease pathologies, and gene regulatory networks.
A variety of different genes have been associated with cell fate. For example, the Nanog/Oct-4/Sox2 network forms the core interaction of a gene network that maintains stem cell pluripotency, and Oct-4 and Sox2 also play a role in the tissue types that stem cells eventually differentiate into. Using the CRISPR/cas9 based homology independent targeted integration (HITI) method developed by Suzuki et al., we can integrate fluorescent tags behind genes with reasonable efficiency via the non-homologous end joining (NHEJ) DNA repair pathway. With human embryonic kidney (HEK) 293T cells, which can be transfected with high efficiencies, we aim to create a three-parameter reporter cell line with fluorescent tags for three different genes related to cell fate. This cell line would provide several advantages for the study of cell fate, including the ability to quantitatively measure cell state, observe expression heterogeneity among a population of genetically identical cells, and easily monitor fluctuations in expression patterns.
The project is partially complete at this time. This report discusses progress thus far, as well as the challenges faced and the future steps for completing the reporter line.
A variety of different genes have been associated with cell fate. For example, the Nanog/Oct-4/Sox2 network forms the core interaction of a gene network that maintains stem cell pluripotency, and Oct-4 and Sox2 also play a role in the tissue types that stem cells eventually differentiate into. Using the CRISPR/cas9 based homology independent targeted integration (HITI) method developed by Suzuki et al., we can integrate fluorescent tags behind genes with reasonable efficiency via the non-homologous end joining (NHEJ) DNA repair pathway. With human embryonic kidney (HEK) 293T cells, which can be transfected with high efficiencies, we aim to create a three-parameter reporter cell line with fluorescent tags for three different genes related to cell fate. This cell line would provide several advantages for the study of cell fate, including the ability to quantitatively measure cell state, observe expression heterogeneity among a population of genetically identical cells, and easily monitor fluctuations in expression patterns.
The project is partially complete at this time. This report discusses progress thus far, as well as the challenges faced and the future steps for completing the reporter line.
ContributorsLoveday, Tristan Andre (Author) / Wang, Xiao (Thesis director) / Brafman, David (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05