Browsing by Author "Hillman, Ethan T."
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Item A Genetic Engineering Toolbox for the Lignocellulolytic Anaerobic Gut Fungus Neocallimastix frontalis(ACS Synthetic Biology, 2023-04-21) Hooker, Casey, A.; Hanafy, Radwa; Hillman, Ethan T.; Muñoz Briones, Javier; Solomon, Kevin V.Anaerobic fungi are powerful platforms for biotechnology that remain unexploited due to a lack of genetic tools. These gut fungi encode the largest number of lignocellulolytic carbohydrate active enzymes (CAZymes) in the fungal kingdom, making them attractive for applications in renewable energy and sustainability. However, efforts to genetically modify anaerobic fungi have remained limited due to inefficient methods for DNA uptake and a lack of characterized genetic parts. We demonstrate that anaerobic fungi are naturally competent for DNA and leverage this to develop a nascent genetic toolbox informed by recently acquired genomes for transient transformation of anaerobic fungi. We validate multiple selectable markers (HygR and Neo), an anaerobic reporter protein (iRFP702), enolase and TEF1A promoters, TEF1A terminator, and a nuclear localization tag for protein compartmentalization. This work establishes novel methods to reliably transform the anaerobic fungus Neocallimastix frontalis, thereby paving the way for strain development and various synthetic biology applications.Item Top-down enrichment of oil field microbiomes to limit souring and control oil composition during extraction operations(AIChE Journal, 2022-10-17) Hillman, Ethan T.; Caceres-Martinez, Louis Edwards; Kilaz, Gozdem; Solomon, Kevin V.Microbial processes sour oil, corrode equipment, and degrade hydrocarbons at an annual global cost to the oil and gas industry of nearly $2 billion. However, top-down control of these microbial processes can reduce their damage and enhance oil recovery. Here, we screened microbial communities from five oil wells in the Illinois basin and evaluated nutrient injection strategies to control metabolism and community composition. Molasses with molybdate supplementation stimulated gas and organic acid production while suppressing corrosive H2S formation in samples from two wells. These changes were accompanied with significant reshaping of the microbiome community. Simulations of field operations via a lab-scale mini-coreflood validated that oil well microbiomes can be engineered to inhibit deleterious H2S and shape oil hydrocarbon composition in situ. These pilot studies validate the economic potential and sustainability of top-down approaches for microbiome engineering to control microbes in oil extraction and enhance the economic viability of oil recovery.