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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

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
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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

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
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Description
Synthetic biology is an emerging engineering disciple, which designs and controls biological systems for creation of materials, biosensors, biocomputing, and much more. To better control and engineer these systems, modular genetic components which allow for highly specific and high dynamic range genetic regulation are necessary. Currently the field struggles to

Synthetic biology is an emerging engineering disciple, which designs and controls biological systems for creation of materials, biosensors, biocomputing, and much more. To better control and engineer these systems, modular genetic components which allow for highly specific and high dynamic range genetic regulation are necessary. Currently the field struggles to demonstrate reliable regulators which are programmable and specific, yet also allow for a high dynamic range of control. Inspired by the characteristics of the RNA toehold switch in E. coli, this project attempts utilize artificial introns and complementary trans-acting RNAs for gene regulation in a eukaryote host, S. cerevisiae. Following modification to an artificial intron, splicing control with RNA hairpins was demonstrated. Temperature shifts led to increased protein production likely due to increased splicing due to hairpin loosening. Progress is underway to demonstrate trans-acting RNA interaction to control splicing. With continued development, we hope to provide a programmable, specific, and effective means for translational gene regulation in S. cerevisae.
ContributorsDorr, Brandon Arthur (Author) / Wang, Xiao (Thesis director) / Green, Alexander (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
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Description
Smartphones have become an integral component of lifestyles worldwide, acting as mobile computers capable of life organization. They remain the most quickly cycled consumer electronic, owned for no more than 3 years on average. Individuals continue to upgrade their smartphones quickly, stemming from the desire for more power and better

Smartphones have become an integral component of lifestyles worldwide, acting as mobile computers capable of life organization. They remain the most quickly cycled consumer electronic, owned for no more than 3 years on average. Individuals continue to upgrade their smartphones quickly, stemming from the desire for more power and better features. In 2016, there were 1.15 billion smartphone upgrades, resulting in a growing used smartphone market valued at \$18 billion. Individuals continue to invest time and effort to sell their smartphone, receiving payment of less than market value. In regards to value-minded users with solidified schedules, I created Trusted Trade-in. This startup provides the bustling middle class with the ability to upgrade their smartphone in an efficient and valuable manner. Compared to current solutions, Trusted Trade-in offers an all-in-one upgrade system. The creation of this startup involved the complete creation of a business model in addition to the coding of a responsive website. An online-based business, customers will be able to visit the Trusted Trade-in website and be given the options to trade-in or trade-up. Competing against Craigslist, eBay and Verizon, Trusted Trade-in features a combined smartphone resale and upgrade process. If the decision is made to trade-in, the customer will be quoted for their current smartphone according to specific physical criteria. The trade-up option will request the same information from the customer and allow them to select a new model for their upgrade. This exciting and innovative marketplace will completely transform the way people upgrade their smartphones through financial and time-based savings.
ContributorsWoods, Quintin Delane (Author) / Sebold, Brent (Thesis director) / Lin, Elva S. Y. (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2017-05
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Description
Military personnel are affected by muscle fatigue during the various missions and training regimens for their work. Muscle fatigue is caused by the overuse and lack of nutrients to muscles. When a soldier is fatigued, they are unable to perform at their maximum potential and are also more susceptible to

Military personnel are affected by muscle fatigue during the various missions and training regimens for their work. Muscle fatigue is caused by the overuse and lack of nutrients to muscles. When a soldier is fatigued, they are unable to perform at their maximum potential and are also more susceptible to injury. For military personnel to save time and money as well as become more efficient within the missions they deploy soldiers, muscle fatigue should be predicted. Predicting fatigue will allow for a reduced rate of negative exercise-related impacts. This means that soldiers will be able to avoid potential life threatening situations they encounter due to the muscle fatigue. The newest technology in wearable devices includes clothing that incorporates heart rate monitors, breathing rate and breathing depth sensors, and a database that converts this information into the amount of calories burned during a workout. Fatigue can be tracked and predicted in the military using wearable clothing with activity sensors, preventing further injury to the soldiers and optimizing performance output at all times. For military personnel, the ability to predict fatigue using this technology would be beneficial to the soldiers and the military as a whole.
ContributorsFalk, Brady Thomas (Author) / Lockhart, Thurmon (Thesis director) / Williams, Deborah (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
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Description
Current research into live-cell dynamics, particularly those relating to chromatin structure and remodeling, are limited. The tools that are used to detect state changes in chromatin, such as Chromatin Immunoprecipitation and qPCR, require that the cell be killed off. This limits the ability of researchers to pinpoint changes in live

Current research into live-cell dynamics, particularly those relating to chromatin structure and remodeling, are limited. The tools that are used to detect state changes in chromatin, such as Chromatin Immunoprecipitation and qPCR, require that the cell be killed off. This limits the ability of researchers to pinpoint changes in live cells over a longer period of time. As such, there is a need for a live-cell sensor that can detect chromatin state changes. The Chromometer is a transgenic chromatin state sensor designed to better understand human cell fate and the chromatin changes that occur. HOXD11.12, a DNA sequence that attracts repressive Polycomb group (PCG) proteins, was placed upstream of a core promoter-driven fluorescent reporter (AmCyan fluorescent protein, CFP) to link chromatin repression to a CFP signal. The transgene was stably inserted at an ectopic site in U2-OS (osteosarcoma) cells. Expression of CFP should reflect the epigenetic state at the HOXD locus, where several genes are regulated by Polycomb to control cell differentiation. U2-OS cells were transfected with the transgene and grown under selective pressure. Twelve colonies were identified as having integrated parts from the transgene into their genomes. PCR testing verified 2 cell lines that contain the complete transgene. Flow cytometry indicated mono-modal and bimodal populations in all transgenic cell colonies. Further research must be done to determine the effectiveness of this device as a sensor for live cell state change detection.
ContributorsBarclay, David (Co-author) / Simper, Jan (Co-author) / Haynes, Karmella (Thesis director) / Brafman, David (Committee member) / School of Life Sciences (Contributor) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
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Description

For many, a long-distance hike on the 2,650+ mile Pacific Crest Trail (PCT) is the adventure of a lifetime. The federally designated National Scenic Trail passes through 48 Wilderness Areas in California, Washington, and Oregon on its way from Mexico to Canada. The trail experience on the PCT has been

For many, a long-distance hike on the 2,650+ mile Pacific Crest Trail (PCT) is the adventure of a lifetime. The federally designated National Scenic Trail passes through 48 Wilderness Areas in California, Washington, and Oregon on its way from Mexico to Canada. The trail experience on the PCT has been changing rapidly over the last 20 years due to two main factors: a four-fold increase in hikers attempting the whole trail each season; and hikers’ rapid adoption of digital technology like smartphones, GPS, and satellite messengers. Through a literature review and accompanying hiker survey, this study aimed to determine how these two factors have combined to alter the trail experience. Despite increased traffic on the trail, hikers appear to still be able to find ample solitude and a feeling of escape from society, and they reported being more likely to form lasting friendships as part of a “trail family”. However, increased traffic has altered many of the sensitive natural landscapes along the trail, contributed to the retirement of some iconic “trail angels” and led to increased conflict between subcultures within the community. Digital technology usage, particularly the use of smartphones and GPS-capable mapping apps, seems to be linked to decreased feelings of solitude, self-sufficiency, and escape. However, digital devices have helped democratize long-distance hiking by simplifying the logistics of long-distance hikes. Users of the devices also did not report reduced feelings of freedom or challenge from their hikes. Moreover, device users still felt that they were disconnecting with technology when hiking on the trail. Acknowledging both positive and negative effects of the changing trail experience, hikers can make more informed decisions about how to mitigate the negative impacts and maximize the positive impacts on the aspects of the trail experience they care the most about.

ContributorsDeSimone, Dante (Author) / Shaeffer, Duncan (Thesis director) / Schmidt, Peter (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description

Phoenix Police officers are required to wear Body-Worn Cameras while out on patrol and must have the cameras turned on when interacting with the public. The Body-Worn Camera (BWC) Policy was initially established as a means of accruing evidence and increasing police accountability when in the presence of the public.

Phoenix Police officers are required to wear Body-Worn Cameras while out on patrol and must have the cameras turned on when interacting with the public. The Body-Worn Camera (BWC) Policy was initially established as a means of accruing evidence and increasing police accountability when in the presence of the public. However, BWC technology has the ability to perform many other useful functions. The information provided by the cameras could be used to reduce the paperwork done by police officers while on duty, thus allowing them to spend more time taking calls from dispatch. The versatility of the body-worn camera and its components also make it an ideal pairing for an electrocardiograph (ECG) device to aid in the health of officers and law enforcement retention.

ContributorsChacon, Elyana (Author) / Ross, Heather (Thesis director) / Scott, Michael (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2023-05
Description

This study investigates the impact of technology and social media on religious practices and beliefs concerning death and the afterlife. As the concept of a "Digital Afterlife" becomes more prevalent, questions surrounding its compatibility with religious belief systems and implications on privacy arise. The COVID-19 pandemic has intensified the issue,

This study investigates the impact of technology and social media on religious practices and beliefs concerning death and the afterlife. As the concept of a "Digital Afterlife" becomes more prevalent, questions surrounding its compatibility with religious belief systems and implications on privacy arise. The COVID-19 pandemic has intensified the issue, prompting social media platforms to develop digital wills, although their usage remains limited. This research seeks to explore how the Information Age is shaping the concept of the afterlife, its alignment with major religious belief systems, and perceptions of the digital afterlife across various societal groups. Furthermore, the study examines the role of social media in redefining religious values, norms, and boundaries, highlighting the importance of engaging in an ongoing conversation about the complex and evolving intersection of religion, technology, and death.

ContributorsAlsabah, Wid (Author) / Hussain, Faheem (Thesis director) / Mostafa, Mashiat (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2023-05
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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

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.
ContributorsLoveday, Tristan Andre (Author) / Wang, Xiao (Thesis director) / Brafman, David (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05