Theses and Dissertations from UMD
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Item FEEDBACK-CONTROLLED BIOELECTRONIC HYBRID SYSTEM ENABLED BY ELECTROGENETIC CRISPR(2023) Wang, Sally Patricia; Bentley, William E; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)With the rise of concepts like the “internet of things” and the advances in electronic technologies, our lives have now been occupied with smart devices that easily communicate with one another. These devices, however, lack the ability to freely exchange information with the world of biology, since electronics and biology possess very different communication modalities. Recently, the field of “electrogenetics” was introduced by enlisting redox mediators like hydrogen peroxide as a novel signaling medium to facilitate the connection between electronics with biology. In this dissertation, we expanded the electrogenetic framework and established a complete network of Bio-Nano Things, which collectively allowed automated, algorithm-based feedback control of electrogenetic CRISPR activity. First, we engineered the abiotic/biotic interface in order to improve information transfer between electronics and biological systems. Inspired by nature, we created an “artificial biofilm” that immobilized living cells on the surface of the electrode by electrochemically assembling bacteria and thiolated polyethylene glycol (PEG-SH) to form a thin film. We then endowed the PEG-SH hydrogel with redox capabilities via conjugation to generate an interactive material that can autonomously synthesize hydrogen peroxide to initiate communication with a bacterial population. Additionally, a polycysteine-tagged Streptococcal protein G was introduced for PEG-SH hydrogel surface decoration to enable the recognition of cells and other biological molecules. Next, we developed oxyRS-based electrogenetic CRISPR to broaden the bandwidth of electrochemical signaling, allowing multiplexed transcriptional regulation on various genetic targets. These include two crucial quorum sensing genes that controlled the relay of electrochemical signals to a broader yet selective audience of microbial populations through quorum sensing communication. We then integrated the engineered interface with eCRISPR-mediated transcriptional regulation to present “Biospark”, a full electrogenetic system including custom-made hardware and software, for algorithm-governed automated control of gene expression. Finally, we demonstrated a network of Bio-Nano Things by connecting the Biospark system with another custom bio-electrochemical device and even users to achieve remote feedback control of eCRISPR activity and more importantly, multidirectional communication between living systems regardless of physical distance. Together, we believe this work represents a huge leap toward making “smarter” devices and networks that can seamlessly guide biological processes with electronic input and can spawn various applications in the fields of biotechnology.Item Electrogenetic communication structures to encode, propagate, and decode information through synthetic biology(2022) VanArsdale, Eric S; Bentley, William E; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Cellular communication, the exchange of information between cells, is a process in which cues are encoded, propagated, and decoded through shared chemical and physical semantics. This general process forms the basis for homeostatic decision-making to which the cell dedicates substantial resources. Central to these processes are abstract communication structures each which serve a defined role and create semantic compatibility. “Transmitters” relay messages away from the source, the “channel” propagates the message through a network, and “receivers” are the terminal site of relay. When a particular semantic code of a message is not fit to the structure it must interact with there is a loss in communication capacity. Overcoming this barrier requires transformation of semantic variants to fit the context of a particular communication channel. For bioelectronics, communication structures must interconvert electron-oriented codes, such as current and voltage, into molecularly-oriented codes, such as genetic inducers and molecular gradients. Here, I present the creation of a redox electrogenetic communication channel which contains the principal transmitter, channel, and receiver structures, each of which is made through the combination of electrochemistry, materials science, and synthetic biology. Each of these structures is the focus of separate sections within my dissertation. First, I will present receiver structures that convert molecular information into electrochemical redox signals. These systems are comprised of engineered bacteria that recognize molecular cues and catalytically synthesize redox-active components that are received using electrochemistry. Second, I will present transmitter structures that encode electronic inputs into redox-active inducers that are perceived by genetic circuits. These circuits interact with the message to ensure faithful replication of the message and constitute electrogenetic noise suppression at the single-cell level and second messenger homogenization on the population level. Lastly, I will present channel propagation which links transmitter and receiver structures. These systems link the prior two sections and network communication into a greater context of biological control. I will demonstrate these interactions through controlled production of small molecules, on-demand cell lysis, and restructuring of cell consortia. Together, these methodologies constitute a bioelectronic communication paradigm that can directly mediate information exchange between electronic and molecular formats. Future application of these communication structures will enable logic-driven control systems and computer-learning models in a medical and environmental internet of bionano-things and true cybernetic integration.