Pacific McGeorge School of Law
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Comprehensive analysis of antibiotic susceptibility across Variovorax to prepare for genetic analysis of horizontal gene transfer between strains.
No full textGenetic manipulation of bacteria often involves the use of recombinant DNA techniques such as heterologous gene expression from plasmids and insertional mutagenesis. In order to effectively use these techniques it is necessary to use selectable markers, the most common of these being antibiotics. In our lab we commonly use two antibiotics, Kanamycin and Gentamicin, to select for recombinant strains. As we expand our studies other isolates within the genus Variovorax, it is necessary to text for susceptibility to these compounds to ensure that we can use them for selection. In this work we report the results of minimum inhibitory concentration (MIC) experiments using these two antibiotics against multiple Variovorax isolates in a broth microdilution assay. The strain was first inoculated into YE broth and incubated for about 24 hours prior to MIC testing. The diluted antibiotics, Kanamycin and Gentamicin, were prepared in 96-well plates with concentrations spanning from 1024 µg/mL- 0.5 µg/mL. Rows A-D were treated with Kanamycin, while rows E-H were treated with Gentamicin. After performing dilution across all columns, all wells were inoculated with a standardized bacterial dose. Column 12 was left free of antibiotic and acted as a positive control for bacterial growth, and rows D and H were uninoculated to ensure the test medium was sterile. Incubation took place at 30°C for 24–48 hours without shaking, and the endpoint growth was observed by eye and measured using a Spectramax250 plate reader. The MIC determined to be the lowest concentration of antibiotic that completely inhibited visible growth. This experiment will be the basis going forward for subsequent genetic experiments using these strains
Exploring the Impact of AI in Higher Education: Efficacy, Ethics, Risks, and Future Trends
No full textGenerative Artificial Intelligence is a transformative tool reshaping today’s society and driving unprecedented changes. As Stanford University puts it, there are “...emerging areas of exploration for AI and learning, including collaborative classroom tools,” and “...teachers want to understand how to use AI to teach, how to teach about AI, and how AI actually works” (Sacks, 2025). This research examines case studies, interviews, and existing scholarly literature to identify AI\u27s efficacy, ethicality, risks, and benefits in higher education. According to the Educause Review, “these efficiencies will lead to increased efficacy – to more effective teaching, learning, institutional decisions, and guidance” (Zeide, 2019). Through Functionalist Theory, we explore the potential risks and benefits of using AI in higher education settings.
This research will investigate potential future trends, including the evolution of AI technology, its integration into various sectors, and the societal consequences of its widespread adoption. As Dr. Andrew Martin, a professor at the University of New South Wales, states, “The more you rely on generative AI to help you with your schoolwork, the less you might be inclined to meet up with friends in person or online after school to brainstorm around an essay” (Abrams, 2025). With this, our research should contribute to the ongoing debate about the responsible development and deployment of AI in our increasingly digital world
The Carlson Lab Fraction Library for Secondary Metabolite Screening of Marine Bacteria
No full textActinomycetes and cyanobacteria are prolific producers of secondary metabolites commonly found in natural product libraries. The Carlson Lab Fraction Library contains fractionated chemical extracts from bacterial and fungal strains isolated from aquatic environments. These extracts are organized in 96-well plates for long-term storage and efficient bioassay testing. Approximately 300 crude extracts have been prepared by liquid-liquid partitioning of culture media with ethyl acetate. For actinomycetes and cyanobacteria, the crude extract is fractionated over silica gel column chromatography, with four different solvent mixtures used to elute different compounds according to their polarity. Each fraction or partition is dried and weighed, and a 7.5 mg subsample is dissolved in DMSO at 10 mg/mL for cryopreservation in an indexed 96-well plate at -20°C. This library has amassed 650 extracts and fractions, enabling rapid bioassay testing without requiring sample preparation each time an assay is executed. Undergraduates here at Pacific have screened for antibiotic activity against 4 human pathogens and, through a collaboration with UC Merced, testing against a panel of human pathogenic fungi is underway. By facilitating high-throughput biological testing and analysis of natural product fractions, the creation of this library is instrumental in our search for novel bioactive compounds
Synthesis and Preliminary Characterization of the Antimicrobial Peptide P-113 Using SPPS
No full textPRESENTED BY: JUNE CHOI, WILEY WINKLER, BRENDEN, ALLEN BRYAN
Title: Synthesis and Preliminary Characterization of the Antimicrobial Peptide P-113 Using SPPS
Introduction:
Peptides are short chains of amino acids that play diverse and vital roles within the human body. As the fundamental units that make up proteins, they play a crucial role in the growth, repair, maintenance, and even defense of cells and our body. The focus of the research was on the synthesis of the fragments of the P-113 antimicrobial peptide (AKRHHGYKRKFH). Found in human saliva,it exhibits activity against various oral pathogens. Through the selection of this peptide sequence, we sought to investigate how easily the peptide fragments (AKRH, HGYK, and RKFH) could be detected with mass spectrometry, as well as set up future experiments for testing the antimicrobial efficacy of these fragments against a range of bacterial strains.
Method:
Solid Phase Peptide Synthesis (SPPS) was employed to construct these peptides. In this technique, the C-terminal amino acid is directly attached to resin beads, allowing for the controlled, stepwise chain elongation. Each amino acid has an Fmoc (9-fluorenylmethyloxycarbonyl) protecting group on the -amino group, which prevents unwanted side reactions and ensures selective coupling. The deprotection step with 20% Piperidine in DMF removed this protecting group, and the coupling steps added the amino acid to the peptide. This process continues sequentially from the C-terminus to the N-terminus. Once synthesis was complete, the peptide was cleaved from the resin, releasing it into solution. The crude product was then purified to remove excess reagents and byproducts, and subsequently freeze-dried with a lyophilizer. Finally, the percent yield of the peptide was calculated.
Result:
By following this procedure, the desired antimicrobial peptide fragments were successfully synthesized and their purity was evaluated by mass spectrometry. The peptide sequence will be used for future studies on its properties and antimicrobial efficacy by using mass spectrometry methods and zone inhibition experiments
Preserving Pichia: A Study on Its Longevity on Freeze-Dried Rodent Pellets
No full textOur group has the goal of developing the yeast Pichia pastoris into a probiotic that can colonize human digestive tracts and deliver therapeutic proteins. To introduce Pichia pastoris into the mouse gut, a model for human beings, we freeze dried cultured P. pastoris into the food pellets of mice. This study examined the survival of P. pastoris on two different food pellets, grain-based and purified. We soaked both food pellets in concentrated liquid cultures of the yeast before freeze drying. To determine the viability of the yeast in these pellets, the freeze-dried pellets were ground, resuspended in liquid and then plated on selective medium. P. pastoris growth was tracked weekly by counting colonies on selective medium. While yeast viability was sufficiently high on the grain-based pellets during the first week, it decreased between weeks two and four. Nevertheless, four weeks after freeze drying, the concentration of surviving yeast remained sufficiently high to feed mice more than 1 × 108 colony forming units per day using the grain-based food pellet. The same method was used to freeze dry P. pastoris onto purified food pellets, which consequently showed lower survival of P. pastoris. The viability of P. pastoris in the purified food pellet suggests that significantly lower than 1 × 108 colony forming units per day would be achieved. Our results demonstrate that freeze drying P. pastoris onto a grain-based diet would enable us to treat mice with the desired concentration of living yeast, a key step toward developing P. pastoris into a probiotic that could potentially ameliorate several diseases
Cracking Under Pressure: Vocal Stress and Warm-Up in Humpback Whale Food Calls
No full textHumpback whales (Megaptera novaeangliae) are baleen whales that travel to the northern Pacific Ocean in summer to forage on herring. Some of these whales engage in a cooperative foraging strategy known as bubble-net feeding, in which group members release a net of bubbles which surrounds a school of fish, and produce loud, low-frequency vocalizations, known as food calls, which function to further aggregate prey into denser schools (Hanser, 2009). These calls can be classified into two primary categories: tonal and modulated. The fundamental frequency, or lowest frequency component, of tonal food calls remains relatively stable, but it fluctuates rhythmically in modulated food calls. We observed that sometimes the whale’s voice seems to ‘crack’ during a food call, exhibiting a brief but sudden shift in fundamental frequency. We proposed that the exertion associated with food call production induces stress on the vocal tract, leading to voice cracks (Fitch et al., 2002). Previous research has shown no differences in the occurrence of vocal cracks between tonal and modulated food calls, suggesting both are equally stressful. We hypothesized that longer call series would contain more vocal cracks due to stress generated by repeated production of difficult calls. We further hypothesized that whales may need to ‘warm up’ their vocal tract before producing food calls to avoid vocal cracks. We tested these hypotheses by using Audacity sound software to analyze acoustic recordings of humpback whales performing bubble-net feeding. We matched food call series with cracks to randomly selected call series containing similar call types but without voice cracks to examine the effect of series length and the presence of calls preceding the series on the occurrence of voice cracks. If ‘warming up’ helps prevent vocal cracks, we expect that food calls series without vocal cracks will be preceded by calls containing significant frequency modulation, much like those produced by professional singers prior to performing