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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 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
Description
Insects are able to navigate their environments because they can detect hydrocarbons and volatile odors, but it is not clear which one has the fastest reaction when detected, or how much of a response can be produced due to either one. In order to determine which category of odorant is detected first as well as which one causes the highest response rate, data on electrophysiological responses from ants was analyzed. While the statistical tests can be done to understand and answer the questions raised by the study, there are various hydrocarbons and volatile odors that were not used in the data. Conclusive evidence only applies to the odorants used in the experiments.
ContributorsDarden, Jaelyn (Author) / Gerkin, Richard (Thesis director) / Liebig, Juergen (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
Cardiovascular disease (CVD) remains the leading cause of mortality, resulting in 1 out of 4 deaths in the United States at the alarming rate of 1 death every 36 seconds, despite great efforts in ongoing research. In vitro research to study CVDs has had limited success, due to lack of biomimicry and structural complexity of 2D models. As such, there is a critical need to develop a 3D, biomimetic human cardiac tissue within precisely engineered in vitro platforms. This PhD dissertation involved development of an innovative anisotropic 3D human stem cell-derived cardiac tissue on-a-chip model (i.e., heart on-a-chip), with an enhanced maturation tissue state, as demonstrated through extensive biological assessments. To demonstrate the potential of the platform to study cardiac-specific diseases, the developed heart on-a-chip was used to model myocardial infarction (MI) due to exposure to hypoxia. The successful induction of MI on-a-chip (heart attack-on-a-chip) was evidenced through fibrotic tissue response, contractile dysregulation, and transcriptomic regulation of key pathways.This dissertation also described incorporation of CRISPR/Cas9 gene-editing to create a human induced pluripotent stem cell line (hiPSC) with a mutation in KCNH2, the gene implicated in Long QT Syndrome Type 2 (LQTS2). This novel stem cell line, combined with the developed heart on-a-chip technology, led to creation of a 3D human cardiac on-chip tissue model of LQTS2 disease.. Extensive mechanistic biological and electrophysiological characterizations were performed to elucidate the mechanism of R531W mutation in KCNH2, significantly adding to existing knowledge about LQTS2. In summary, this thesis described creation of a LQTS2 cardiac on-a-chip model, incorporated with gene-edited hiPSC-cardiomyocytes and hiPSC-cardiac fibroblasts, to study mechanisms of LQTS2.
Overall, this dissertation provides broad impact for fundamental studies toward cardiac biological studies as well as drug screening applications. Specifically, the developed heart on-a-chip from this dissertation provides a unique alternative platform to animal testing and 2D studies that recapitulates the human myocardium, with capabilities to model critical CVDs to study disease mechanisms, and/or ultimately lead to development of future therapeutic strategies.
ContributorsVeldhuizen, Jaimeson (Author) / Nikkhah, Mehdi (Thesis advisor) / Brafman, David (Committee member) / Ebrahimkhani, Mo (Committee member) / Migrino, Raymond Q (Committee member) / Plaisier, Christopher (Committee member) / Arizona State University (Publisher)
Created2021
Description
Interdigitated back contact (IBC) solar cells have achieved the highest single junction silicon wafer-based solar cell power conversion efficiencies reported to date. This thesis is about the fabrication of a high-efficiency silicon heterojunction IBC solar cell for potential use as the bottom cell for a 3-terminal lattice-matched dilute-nitride Ga (In)NP(As)/Si monolithic tandem solar cell. An effective fabrication process has been developed and the process challenges related to open circuit voltage (Voc), series resistance (Rs), and fill factor (FF) are experimentally analyzed. While wet etching, the sample lost the initial passivation, and by changing the etchant solution and passivation process, the voltage at maximum power recovered to an initial value of over 710 mV before metallization. The factors reducing the series resistance loss in IBC cells were also studied. One of these factors was the Indium Tin Oxide (ITO) sputtering parameters, which impact the conductivity of the ITO layer and transport across the a-Si:H/ITO interface. For the standard recipe, the chamber pressure was 3.5 mTorr with no oxygen partial pressure, and the thickness of the ITO layer in contact with the a-Si:H layers, was optimized to 150 nm. The patterning method for the metal contacts and final annealing also change the contact resistance of the base and emitter stack layers. The final annealing step is necessary to recover the sputtering damage; however, the higher the annealing time the higher the final IBC series resistance. The best efficiency achieved was 19.3% (Jsc = 37 mA/cm2, Voc = 691 mV, FF = 71.7%) on 200 µm thick 1-15 Ω-cm n-type CZ C-Si with a designated area of 4 cm2.
ContributorsMoeini Rizi, Mansoure (Author) / Goodnick, Stephen (Thesis advisor) / Honsberg, Christina (Committee member) / Goryll, Michael (Committee member) / Smith, David (Committee member) / Bowden, Stuart (Committee member) / Arizona State University (Publisher)
Created2022
Description
Skin wounds can be caused by traumatic lacerations or incisions which disrupt the structural and functional integrity of the skin. Wound closure and primary intention treatment of the wound as soon as possible is crucial to avoid or minimize the risk of infection that can result in a compromised healing rate or advanced functional intricacy. The gold standard treatment for skin wound healing is suturing. Light-activated tissue sealing is an appealing alternative to sutures as it seals the wound edges minimizing the risk of infection and scarring, especially when utilized along with biodegradable polymeric biomaterials in the wound bed. Silk fibroins can be used as a biodegradable biomaterial that possesses properties supporting cell migration and proliferation in the tissue it interacts with. In addition, histamine treatment is shown to have extensive effects on cellular functions promoting wound healing. Here, the evaluation of Laser-activated Sealants (LASE) consisting of silk fibroin films induced with Indocyanine Green dye in a wound sealed with laser in the presence of Histamine receptor agonists H1R, H2R and H4R take place. The results were evaluated using Trans-epidermal Water Loss (TEWL), histological and analytical techniques where immune cell biomarkers Arginase-1, Ly6G, iNOS, Alpha-SMA, Proliferating Cell Nuclear Antigen (PCNA), and E-Cadherin were used to study the activity of specific cells such as macrophages, neutrophils, and myofibroblasts that aid in wound healing. PBS was used as a control for histamine receptor agonists. It was found that TEWL increased when treated with H1 receptor agonists while decreasing significantly in H2R and H4R-treated wounds. Arginase-1 activity improved, while it displayed an inverse relationship compared to iNOS. H4R agonist escalated Alpha-SMA cells, while others did not have any significant difference. Ly6G activity depleted in all histamine agonists significantly, while PCNA and E-Cadherin failed to show a positive or negative effect.
ContributorsPatel, Dirghau Manishbhai (Author) / Rege, Kaushal (Thesis advisor) / Massia, Stephen (Committee member) / Brafman, David (Committee member) / Arizona State University (Publisher)
Created2022
Description
The future will be replete with Artificial Intelligence (AI) based agents closely collaborating with humans. Although it is challenging to construct such systems for real-world conditions, the Intelligent Tutoring System (ITS) community has proposed several techniques to work closely with students. However, there is a need to extend these systems outside the controlled environment of the classroom. More recently, Human-Aware Planning (HAP) community has developed generalized AI techniques for collaborating with humans and providing personalized support or guidance to the collaborators. In this thesis, the take learning from the ITS community is extend to construct such human-aware systems for real-world domains and evaluate them with real stakeholders. First, the applicability of HAP to ITS is demonstrated, by modeling the behavior in a classroom and a state-of-the-art tutoring system called Dragoon. Then these techniques are extended to provide decision support to a human teammate and evaluate the effectiveness of the framework through ablation studies to support students in constructing their plan of study (\ipos). The results show that these techniques are helpful and can support users in their tasks. In the third section of the thesis, an ITS scenario of asking questions (or problems) in active environments is modeled by constructing questions to elicit a human teammate's model of understanding. The framework is evaluated through a user study, where the results show that the queries can be used for eliciting the human teammate's mental model.
ContributorsGrover, Sachin (Author) / Kambhampati, Subbarao (Thesis advisor) / Smith, David (Committee member) / Srivastava, Sidhharth (Committee member) / VanLehn, Kurt (Committee member) / Arizona State University (Publisher)
Created2022
Description
The advent of CRISPR/Cas9 revolutionized the field of genetic engineering and gave rise to the development of new gene editing tools including prime editing. Prime editing is a versatile gene editing method that mediates precise insertions and deletions and can perform all 12 types of point mutations. In turn, prime editing represents great promise in the design of new gene therapies and disease models where editing was previously not possible using current gene editing techniques. Despite advancements in genome modification technologies, parallel enrichment strategies of edited cells remain lagging behind in development. To this end, this project aimed to enhance prime editing using transient reporter for editing enrichment (TREE) technology to develop a method for the rapid generation of clonal isogenic cell lines for disease modeling. TREE uses an engineered BFP variant that upon a C-to-T conversion will convert to GFP after target modification. Using flow cytometry, this BFP-to-GFP conversion assay enables the isolation of edited cell populations via a fluorescent reporter of editing. Prime induced nucleotide engineering using a transient reporter for editing enrichment (PINE-TREE), pairs prime editing with TREE technology to efficiently enrich for prime edited cells. This investigation revealed PINE-TREE as an efficient editing and enrichment method compared to a conventional reporter of transfection (RoT) enrichment strategy. Here, PINE-TREE exhibited a significant increase in editing efficiencies of single nucleotide conversions, small insertions, and small deletions in multiple human cell types. Additionally, PINE-TREE demonstrated improved clonal cell editing efficiency in human induced pluripotent stem cells (hiPSCs). Most notably, PINE-TREE efficiently generated clonal isogenic hiPSCs harboring a mutation in the APOE gene for in vitro modeling of Alzheimer’s Disease. Collectively, results gathered from this study exhibited PINE-TREE as a valuable new tool in genetic engineering to accelerate the generation of clonal isogenic cell lines for applications in developmental biology, disease modeling, and drug screening.
ContributorsKostes, William Warner (Author) / Brafman, David (Thesis advisor) / Jacobs, Bertram (Committee member) / Lapinaite, Audrone (Committee member) / Tian, Xiaojun (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2022
Description
Written corrective feedback (WCF) has received considerable attention in secondlanguage (L2) writing research. The conducive role of WCF in developing L2 writing and
second language acquisition has been corroborated by a number of theoretical frameworks,
and the findings of empirical studies, meta-analyses, and research syntheses. WCF research
has predominantly addressed its effectiveness in improving learners’ syntactic, lexical, and
orthographic knowledge. This dissertation project extends the scope of this line of research
to formulaic aspects of language and investigates the relative effectiveness of WCF
targeting formulaic vs. non-formulaic constructions in L2 writing. The text-analytic
descriptive aspect of this research design aimed at investigating the extent of L2 learners’
non-target-like use of formulaic vs. non-formulaic forms in L2 writing and writing
teachers’ WCF treatment of non-target (non)formulaic language use. A total of 480 first
drafts of essays written by 33 advanced adult English-as-a-foreign language (EFL) learners
during one semester and 480 drafts of essays corrected through WCF by three EFL teachers
constituted the corpus in this study. Advancing the field of learner corpus research, the
findings demonstrated that whereas learners’ non-target formulaic forms outnumbered that
of non-formulaic ones in their writing assignments, all three teachers provided WCF more
often for erroneous use of non-formulaic forms. The quasi-experimental aspect of the
research design attempts to add new empirical evidence on the L2 learning potential of
accessing and processing WCF provided for formulaic vs. non-formulaic constructions in
L2 writing. To this end, a total of 66 EFL learners in a Test of English as a Foreign
Language preparation course participated in a pretest-posttest design, with 5 experimental groups (those who were provided with direct, indirect, direct plus metalinguistic, and
indirect plus metalinguistic WCF) and a control group (those who were not provided with
WCF). Maintaining a division between formulaic vs. non-formulaic forms, the findings
provide empirical evidence on the interactions between types of WCF, types of linguistic
targets, and the effectiveness of WCF in terms of enhancing L2 learners’ accuracy and
acquisition in their revised writing and new writings in the short and long term.
ContributorsGholami, Leila (Author) / Smith, David (Thesis advisor) / Matsuda, Paul K (Committee member) / James, Mark A (Committee member) / Arizona State University (Publisher)
Created2022
Description
Ecology has been an actively studied topic recently, along with the rapid development of human microbiota-based technology. Scientists have made remarkable progress using bioinformatics tools to identify species and analyze composition. However, a thorough understanding of interspecies interactions of microbial ecosystems is still lacking, which has been a significant obstacle in the further development of related technologies. In this work, a genetic circuit design principle with synthetic biology approaches is developed to form two-strain microbial consortia with different inter-strain interactions. The microbial systems are well-defined and inducible. Co-culture experiment results show that our microbial consortia behave consistently with previous ecological knowledge and thus serves as excellent model systems to simulate ecosystems with similar interactions. Colony patterns also emerge when co-culturing multiple species on solid media. With the engineered microbial consortia, image-processing based methods were developed to quantify the shape of co-culture colonies and distinguish microbial consortia with different interactions. Factors that affect the population ratios were identified through induction and variations in the inoculation process. Further time-lapse experiments revealed the basic rules of colony growth, composition variation, patterning, and how spatial factors impact the co-culture colony.
ContributorsChen, Xingwen (Author) / Wang, Xiao (Thesis advisor) / Kuang, Yang (Committee member) / Tian, Xiaojun (Committee member) / Brafman, David (Committee member) / Plaisier, Christopher (Committee member) / Arizona State University (Publisher)
Created2022
Description
Transitioning into civilian life after military service is a challenging prospect. It can be difficult to find employment and maintain good mental health, and up to 70 percent of veterans experience homelessness or alcoholism. Upon discharge, many veterans pursue higher education as a way to reintegrate into civilian society. However, many studies have shown that veterans encounter multiple challenges during their attempt to reintegrate into civilian life, including anxiety, a lack of relevant skills, post-traumatic stress disorder (PTSD), and other issues that may lead to communication and interaction challenges in the higher education environment. Student veterans also face challenges in the lack of common language and culture clashes due to differences between military and college culture. This study used a mixed-methods approach to examine the challenges military veterans face related to language use in civilian life. The data was collected from 149 student veterans who completed a questionnaire and 11 student veterans who participated in interviews. Detailed analysis of collected data showed that student veterans experienced some challenges in language use, especially when they initially enrolled in their courses, but they seemed to have overcome challenges after spending time in the university setting. The veterans who had prior college education before joining the military seemed to have a slight advantage, having had experience using the academic language. The study also explored how student veterans chose to share their veteran status with other people in their university community. The findings showed that they strongly identified with their veteran identity and was comfortable sharing their status with others, but they also sometimes were reluctant to share their military experience in details because they were afraid that their peers would not understand.
ContributorsObaid, Naji (Author) / Matsuda, Aya (Thesis advisor) / Smith, David (Committee member) / James, Mark (Committee member) / Arizona State University (Publisher)
Created2022