PCR - Spring 2022

PCR. Spring 2022.

Research Week.

Special Edition.

Contents.

AP Biology Impact of Phosphate and Nitrate Supplemented Diet on Drosophila melanogaster 9 Antibacterial Properties of Ginger Extract and Lactobacillus fermentum 9 Relation between Sugar Growth and Saccharomyces cerevisiae 10 Phytoremediation with Lemna minor 10 Relationship between Lemna minor and Heavy Metals 11 Impact of Environmental Salinity on Saccharomyces cerevisiae 11 Effect of UV Radiation Exposure on Saccharomyces cerevisiae 12 Effect of Glucose Concentration on CO2 Production 12 Effect of Ethanol on the Mass of Tenebrio molitor 13 Effect of Red Bull on Tenebrio molitor 13 Efficacy of Lemna minor to Detect Fragrance Chemical Levels 14 Modeling the Mutagenic Effect of Escherichia coli through the Ames Test 14 Analyzing Lemna minor Absorption of CO2 15 Effect of pH on Protein Concentration found in Saccharomyces cerevisiae 15 Cryphonectria parasitica and Hypoviruses 16 2

Ideal pH for Growth of Lemna minor 16 Effect of Agrobacterium tumefaciens on Lemna minor 17 Effect of Polystyrene on Tenebrio molitor Development and Mortality 17 Effect of Different Saline Concentrations on Lemna Minor Growth 18 Efficacy of Venturia inaqeualis on Malus domestica 18

Methods in Molecular Biology Research Summary of Research Class 19 Dnase1L3 Mutations Provide Novel Insight into the Catalytic Mechanism of Dnase1 Family Endonucleases 20 Bioinformatic Characterization of 3L1W 21

MAPS Team The Role of Ideonella sakaiensis PETase in the Degradation of PET Plastics: a Structural Comparison of the Wild Type and S238F/W159H Double Mutant 22

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Contents.

HIRT American Modernism 23 Creativity During Covid-19 24 Cultural Racism and Its Imperceptible Impacts 24

IRT Accelerated Molecular Dynamics for Complex Physical Systems 25 Artificial Collagen for as a Biomaterial 26 Drover: Drone-Rover Communication for Pathfinding 27 Effect of Dietary Restrictions on the Lifespan of Drosophila melanogaster 28 Expression and Analysis of Fluorescent Fish Proteins in Vitro 29 Metagenomic Analysis of a Compost Microbiome 29 PETase - Plastic Degrading Enzymes 30 Protein Purification of Shigella dysenteriae Transcription Factor VirF 31 Quantifying the Effect of Structure on Neural Network Training 32 Quantifying the Phototactic Memory of Chlamydomonas reinhardtii 32 4

Smart Heart: Deep Learning-Based Analysis of 12-Lead Electrocardiograms for Cardiovascular Disease Risk Prediction 33 The Effect of KIF11 Activity on YAP Localization 34

Club Highlights Anatomy Club 35 FYI Sci 35-36 Pingry Community Research 37 Girl Code 37 Robotics 38

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Editor’s Note.

Welcome to the Research Week edition of the Pingry Community Research

(PCR) Journal. We are excited to showcase Pingry’s top scientific talent, both in terms of research skills and knowledge of scientific concepts and discoveries.

The PCR journal provides students the opportunity to publish novel research. Through a written medium, students demonstrate their in-depth understanding of complex, collegiate-level scientific topics, and their applications in research at Pingry.

This special edition of PCR serves as a written complement to the in person poster presentations occurring throughout research week. Readers can preview abstracts, figures, and summaries reflecting the research conducted in advanced courses and extracurriculars, such as AP Biology, Independent Research Teams (IRT), and the Methods in Molecular Biology Research Class, among others.

Through the PCR journal, we hope to spark intellectual curiosity and promote scientific inquiry amongst the next generation of Pingry researchers.

Dive into the wonders of Pingry Research through this special edition of PCR: Pingry’s foremost journal of scientific research.

Mirika Jambudi (V), Editor-in-Chief Kristin Osika (VI), Former Editor-in-Chief

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Editorial Staff.

Editor-in-Chief: Mirika Jambudi (V), current Kristin Osika (VI), former

Head Layout Editor: Sarah Gu (IV)

Head Copy Editor: Evan Xie (IV)

Layout Editors: Leon Zhou (IV) Sriya Tallapragada (III)

Copy Editors: Max Watzky (V) Leon Zhou (IV) Hansen Zhang (IV) Chelsea Peng (IV) Sophia Odunsi (IV) Ali Santana (IV) Brielle Marques (IV) Annabelle Shilling (IV)

Faculty Advisor: Mr. Maxwell

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AP Biology.

Impact of Phosphate and Nitrate Supplemented Diet on Drosophila melanogaster

by James Houghton (VI), Mirika Jambudi (V), Rachel Zhang (V) It has been established that dietary restriction through simple caloric reduction elongates lifespan in Drosophila melanogaster, a model organism used for studies across a variety of disciplines.

A number of different nutritional factors have shown to impact the lifespan of D. melanogaster, but not much is known about their individual roles.

It has been previously demonstrated that phosphate and nitrate play a role in elongation in mice and have been linked to preventing age-related locomotor decline in D. melanogaster.

In this study, the impacts of phosphate and nitrate on the lifespan of D. melanogaster are explored.

Results demonstrate that a nitrate-supplemented diet slightly increased the lifespan of D. melanogaster and that a phosphate restricted diet had no statistically significant effect on the lifespan of D. melanogaster. This indicates that nitrate may be a key nutritional factor involved in lifespan extension, and further research could lead to implications for dietary choices in humans.

Antibacterial Properties of Ginger Extract and Lactobacillus fermentum

by Olivia Hung (VI), Katie Lin (V), Morgan McDonald (V) The purpose of this project is to research the antibacterial properties of ginger extract and antibacterial resistance in Lactobacillus fermentum.

L. fermentum sourced from ATCC was grown in MRS liquid medium and incubated for 4 days until significant growth was determined based on the light permeability of stock tubes. Batches of MRS agar were mixed with the directed water, boiled, autoclaved, and then poured and frigid to produce approximately 35 MRS agar plates for L. fermentum to grow on. In the initial procedure, we intended to compare the growth rates of 10 small control colonies to the growth rate of 10 experimental colonies treated with ginger extract.

The growth rate would be determined by measuring the diameter of the colonies before and

after a determined period of time of 24 hours.

Due to minimal growth of the 5 colonies with a diameter of about 2 mm after 4 days of incubation at 40 degrees celsius, procedural changes were made. In the continuation of the experiment, the growth rate will be measured based on the expansion of these colonies outside of the designated diameter. It is hypothesized that as the concentration of ginger increases, the resistance level of L. fermentum will decrease, supporting the hypothesis that ginger extract has antibacterial properties. The results of this research may provide insight for advancements in probiotics and antibiotic medication, microbial resistance, and safety in the dairy industry.

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Relation between Sugar Growth and Saccharomyces cerevisiae

by Carson Clunis (V), Alexis Matthews (V) More commonly known as yeast, Saccharomyces cerevisiae is a eukaryotic single-celled organism in the fungus kingdom found on plants, animals, and soil and functions as a natural decomposer in the environment. To survive, S. cerevisiae converts carbohydrates and breaks them down into carbon dioxide, ethanol, flavor molecules, and energy in a process called fermentation. Since yeast is a chemoorganotroph, it utilizes organic compounds as an energy source. Yeast must contain carbon to perform fermentation, typically from carbohydrate sugars from hexose sugars (i.e., glucose and fructose) or disaccharides (i.e., sucrose and maltose), and oxygen from the air. This experiment tests S. cerevisiae’s anaerobic respiration

in different sugar environments. The two different sugars, glucose and sucrose, were put into two different large beakers and mixed in with a packet (equal to about 2 ¼ tablespoons) of yeast.

Then, water at 40 degrees celsius was added to each beaker. The difference in the release of carbon dioxide would be measured 20 minutes after a balloon is placed on the beaker. The circumferences of the balloons were measured before and after to show the balloon’s change in volume.

The results of this experiment have shown that, on average, after 20 minutes, the circumference of the sucrose balloon grew 1.2 inches larger than the glucose ballon, which proves that yeast expresses more carbon dioxide when

Phytoremediation with Lemna minor

by Cayden Barrison (V), Kristen Lytle (V), Luigi Zavala (V) Wastewater can be treated with Lemna minor in a more environmentally friendly and cost-effective manner than current techniques, thus allowing L. minor to be be a possible solution to global water pollution.

This study looks to evaluate the efficacy of L. minor in minimizing nitrate concentration in wastewater. Three different concentrations of L. minor were placed into nitrate-concentrated water in order to demonstrate the natural ability of the duckweed to reduce nitrate level in the simulated wastewater. The results revealed that, despite low concentrations of duckweed in comparison to the simulated wastewater, the higher concentration of duckweed reduced the concentration of nitrate better. Phytoremediation with L. minor in a controlled setting can be

positively corroborated and recommended as a potential approach for the treatment of natural environments based on the results of this study.

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Relationship between Lemna minor and Heavy Metals

by Lleyton Lance (VI), Allen Wu (VI) Duckweed, a flowering aquatic plant, has been continually affected by rising heavy metal concentrations in natural bodies of water caused by a multitude of factors including industrial pollution. This study seeks to illustrate the effect of various heavy metals on the growth of duckweed. Duckweed was grown in 0.035g of zinc chloride (ZnCl2), 0.035 g of magnesium chloride (MgCl2), or 0.035 g of copper(II) chloride (​​CuCl2) along with 1.5 mL of tap water in a 24-well microplate. Additionally, three wells of duckweed were grown in a combination of 0.035g ZnCl2, 0.035g MgCl2, and 0.035g ​​CuCl2 and 1.5 mL of tap water for three weeks. This study hypothesized that the growth of duckweed in a combi-

nation of metals would stunt the growth greater than growing the duckweed with individual metals. The results showed that adding heavy metals generally decreased the percent growth of duckweed. In comparison to the control group which displayed a 66.67% increase in duckweed fronds, the ZnCl2, MgCl2, ​​CuCl2, and combination groups had a 9.03%, 8.33%, -2.38%, and -14.74% change in number of duckweed fronds (all p < 0.05). However, the data was not statistically significant (p > 0.05) when comparing individual metals to the combination group, meaning that there is no conclusive evidence that a combination of heavy metals stunts the growth of duckweed to a greater degree than individual metals.

Impact of Environmental Salinity on Saccharomyces cerevisiae

by Adrian Kurylko (V), Gordon Oatman (V) Fermentation is an anaerobic process that converts simple sugars into ATP for cellular energy and produces ethanol and carbon dioxide as byproducts. The rate of yeast fermentation can vary based on different environmental factors.

This study aimed to measure the effects of various salinity levels on the rate of yeast fermentation and use the results to determine the optimal saline environment for yeast fermentation. We hypothesized that low salinity environments are preferable because the hygroscopic nature of salt will dehydrate the yeast cells, thus slowing fermentation. Our setup used a flask containing the yeast-salt-sucrose solution dissolved in water and a tube to collect and measure the carbon dioxide produced. Previous research has shown glucose to be the most efficient sugar for fermentation

but sucrose is a viable alternative. As hypothesized previously, the control group without salt produced the most carbon dioxide after 30 minutes with the highly concentrated 0.5 M solution producing the least amount of carbon dioxide.

The solution’s volume and the masses of yeast and sucrose were kept constant for all groups.

The rate of fermentation increased more rapidly as time passed with little to no water displacement occurring during the first 10 minutes of each trial in each group. This is likely due to a design flaw in which carbon dioxide was getting stuck in the tube but nonetheless, the results of our experiment were consistent with our prediction that fermentation rate would decrease as salt concentration increased.

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Effect of UV Radiation Exposure on Saccharomyces cerevisiae

by Ram Doraswamy (VI), Sankar Gollapudi (VI), Emily Shen (VI) Ultraviolet (UV) radiation is known to damage living cells and organisms. Over 2 months, multiple trials were conducted to examine the impact of UV exposure on the yeast species Saccharomyces Cerevisiae. The process includes growing the yeast cells before placing samples of yeast under UV light for 1, 3, and 5 minutes respectively. Samples are then mixed with methylene blue chloride 1% solutions, which distinguishes the dead cells from the alive ones, as dead cells are stained while alive cells remain uncolored. Because UV radiation is known to damage DNA, more yeast cells are expected to die when they are exposed to UV radiation for longer periods of time. The trial results confirm the hypothesis, revealing a positive correlation between dead yeast cells and time

of UV exposure being observed. This experiment was conducted in order to gain a deeper understanding of how UV light affects living organisms.

Effect of Glucose Concentration on CO2 Production

by Abigail Rubino (VI), Amanda Pfundstein (V) Sugar is one of the various factors shown to impact the rate of fermentation. The rate of fermentation can be determined by carbon dioxide production, a by-product of the cellular process. The study observes the effects of four different glucose concentrations: 0.5 ml, 2.5 ml, 4.5 ml, and 6.5 ml and their respective effects on carbon dioxide production in yeast cells, specifically Saccharomyces cerevisiae (also known as the “sugar-eating fungus”). Across these different glucose concentrations, a constant amount of H2O and Saccharomyces is maintained. Initial results indicate that altering glucose concentrations affects the process of fermentation, more specifically, the amount of carbon dioxide produced. Furthermore, these results reveal a direct relationship between sugar concentra-

tion and carbon dioxide production; higher glucose concentrations increase CO2 production.

This relationship provides valuable knowledge for industries that rely on fermentation, such as the baking industry, which could use these results to maximize fermentation in their products.

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Effect of Ethanol on the Mass of Tenebrio molitor

by Allie de Asla (VI), Cece Korn (V), Sydney Puntus (V) There has been much debate about the true effects of ethanol on human weight, as it is high in caloric content, but can also increase metabolic rate.The experiment presented attempts to understand how small concentrations of ethanol affect the masses of T. molitor. Experimentally, T. molitor were given their normal diet of potatoes infused with different concentrations of ethanol with their masses being recorded periodically over the trials.

Of the concentrations given, results demonstrated that the mass of T. molitor, on average, decreased in the group given the highest concentration, while mass increased in groups given the lower concentrations. These results may provide convincing evidence in favor of ethanol’s ability to cause weight loss by increasing metabolic rate rather than causing weight gain due to its caloric content.

Effect of Red Bull on Tenebrio molitor

by Claire Fernicola (VI), Nav Jha (VI), Grant Myers (VI) Energy drinks are prevalent among humans because of their ability to increase energy; however, are the effects the same among all species? The experiment attempted to answer the question “what are the effects of Red Bull on mealworms?” There were two control groups, in which mealworms were placed inside a choice chamber and given types of grass to eat.

We also used an experimental group where we gave mealworms Red Bull. The results showed an increase in the mealworms’ movement speed after consuming the Red Bull. We suspect this was due to the caffeine in the drink, which indicates that humans may react similarly.

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Efficacy of Lemna minor to Detect Fragrance Chemical Levels

by Connor Chen (VI), Evan Haidri (VI), Franklin Zhu (VI) In today’s world, pollution is a problem that is spreading globally, with massive manufacturing and production spreading harmful pollutants that have irreversible effects on the planet’s ecosystems. As of recently, synthetic musks and fragrance chemical byproducts released by factories producing fragrance products such as perfume and cologne have been identified as a major source of pollution, particularly in aquatic environments. As a result of these harmful substances, nearby bodies of water are polluted, posing a threat to marine life. The purpose of this study was to know whether Lemna minor will be effective at detecting fragrance chemicals levels in order to guide efforts to control and reduce pollution. The presence of L. minor has been used in the past to detect pollution from other compounds in bodies of water. For a study eval-

uating the applicability of L. minor to pollution caused by synthetic musks and fragrance chemicals, we examined the growth and survival of L. minor samples at various concentrations of fragrance pollution over time. Unlike other bacteria, L. minor is unable to grow in low concentrations of fragrance chemicals in the water. As a result, the bacteria usually die a few days after being introduced to the water. The results show that duckweed can be a reliable natural method for determining if fragrance chemical pollutants are present in water. More testing and experiments can be done to determine the exact concentration of chemical pollutants L. minor can withstand, but our results demonstrate that duckweed can be an effective natural method for testing for the presence of fragrance chemical pollutants.

Modeling the Mutagenic Effect of Escherichia coli through the Ames Test

by Jack Martin (VI), Maria Loss (V), Milenka Men (V) Substance abuse has become a public health crisis; while studies on addictive behavior began in the 1930s, those only aimed to decipher addiction behaviorally. Extensive testing has not been done on all chemicals involved, leading to the need for mutagenic testing to be performed. The research project focuses on modeling the mutagenic effects of specific components found in addictive substances through the Ames Test procedure.

The project utilizes E. coli instead of the more commonly utilized Salmonella due to lab safety restrictions. The wild-type E. coli strain requires tryptophan to grow, but if the cells are mutated after exposure to a mutagen, growth without tryptophan occurs. The results showed that cer-

tain chemicals will cause revertant growth without the presence of tryptophan and therefore should be considered mutagenic. Through analyzing the chemical breakdown of vape liquid and assessing risk factors implicated through individual compounds, the results help clear up the current ambiguity surrounding the mutagenicity of electronic cigarettes and deconstruct the possible health concerns regarding the products.

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Analyzing Lemna minor Absorption of CO2

by Israel Billups (VI), Fara Odunlami (VI) lutant. It can be concluded that L. minor would be a more viable option to aid in the control of carbon emissions within our atmosphere.

The purpose of this experiment is to use Lemna minor to absorb atmospheric carbon dioxide. L. minor is an organism that absorbs large amounts of carbon dioxide—which makes up a significant portion of its mass. In this experiment, the prime focus was on its relationship with atmospheric CO2. The experimental question was whether L. minor would be able to absorb enough carbon to be an alternative to harmful atmospheric fertilizers and water cleaning methods. An analysis of the growth of L. minor in environments with varying carbon contents was the approach that this experiment focused on. Results from the experiment support the claim that L. minor grows more in the presence of CO2, meaning that it has the ability to readily absorb atmospheric CO2 that is a major environmental pol-

Figure 1: Comparison of duckweed growth on 12/10/21 and 2/22/22

Effect of pH on Protein Concentration Found in Saccharomyces cerevisiae

by Martine Bigos (VI), Zala Bhan (V), Ainsli Shah (V) Nutritional yeast, an inactive form of the Saccharomyces cerevisiae strain, remains active under a variety of environmental conditions, making it a useful organism for experimentation when knowledgeable of its constraints. This research paper aims to measure and understand the effects of pH on the protein concentration of S. cerevisiae. A solution of malt extract and water supplied energy for the process of respiration.

To create basic or acidic environments, NaOH or HCl was added to the solution. Using the Bradford assay and a nanodrop spectrophotometer, we tracked the protein concentration of S. cerevisiae grown in different pHs. We hypothesized that S. cerevisiae grown in conditions with an unusually high or low pH would exhibit lower protein levels

when compared to the S. cerevisiae grown in the control group at a normal pH, while our results showed no change. Our findings provide an increased understanding of protein denaturation and how pH is most likely not a factor when considering the likelihood of an environment being a sustainable one for organisms like S. cerevisiae.

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Cryphonectria parasitica and Hypoviruses

by Willow Palmer (VI), Molly Parker (VI), Jack Taylor (VI) Cryphonectria parasitica is a virulent fungus that has decimated Chestnut tree populations, but history has proven that using toxic chemicals to kill parasites can affect the entire ecosystem. Using a biological agent to kill the fungi is a safer option, and viruses due to their transmissibility have been put on the forefront of botanists’ research. We wanted to know which hypovirus strain would be most effective in limiting fungal growth. As C. parasitica eats away at cellulose and causes rot on plants, we used apples to simulate the rotting process.

Samples of C. parasitica were inserted into bored holes into Malus domestica ‘Gala’. By the end

of the two weeks the inoculated apple’s lesions were measured for the radius that the rot had spread to. As expected the C. parasitica isolates containing hypovirus isolates showed less radial growth. SCI19 and SCI12 had larger radial rot on the apples with hypovirus infected C. parasitica. EP713 and RCL reliably showed diminished radii in hypovirus positive fungi. Since the hypovirus was clearly effective at stopping the spread of C. parasitica, it offers opportunities to help curb further destruction of nature by C. parasitica in the wild. Scientists could use hypoviruses as a method of stopping the C. parasitica and ensuring the safety and prosperity of natural ecosystems.

Ideal pH for Growth of Lemna minor

by Kaitlyn DeVito (VI), Peter Roelke (VI) Duckweed is a staple in aquatic environments considering its value as a food source for certain fish and semi-aquatic birds in addition to providing a habitat for smaller organisms like frogs. Our research explores the ideal pH for lemna minor to grow in, as pH is one of the crucial environmental factors that determines how well duckweed thrives. We tested a range from pH’s from four to eight across five different trials. We inserted a pH solution at each pH level in a row of six wells per trial. To measure growth in each well plate, we took pictures of the individual wells on days one and fourteen of each trial before using ImageJ analysis. The ImageJ application measures the percentage of each well’s area covered by duckweed. After finding each well’s area percentage on day one, we subtracted these values from the day fourteen wells’ final percentage to find the total percent change in growth. Our first trial did not pass ANOVA significance tests, whereas trials two and three saw many of the plants die off. Our fourth and fifth

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trials passed ANOVA tests because their P-values were lower than a 0.05 significance level. Our results from the fourth and fifth trials are therefore statistically significant. While our results from the first three trials were largely inconclusive due to plant death, our results from trials four and five allow us to conclude that duckweed growth is the most optimal in pH 6 and pH 7. After five trials, it can be concluded that growth is very minimal in pH 4, pH 5 and pH 8 as our statistical analysis indicated that many of the plants shrunk.

Effect of Agrobacterium tumefaciens on Lemna minor

by Lauren Kim (V), Maya Khan (V) Agrobacterium tumefaciens induces tumors in plants including duckweed by transferring segments of T-DNA into plant host genomes (Lee et al.). While some types of bacteria can enter plants through roots, A. tumefaciens typically enters plants through wounds and causes subsequent tumor growth. These tumors can stunt plant growth by blocking the flow of water and nutrients and leave plants more susceptible to harsh environmental conditions (University of Minnesota). This experiment investigated the effects of A. tumefaciens (administered via root system) on duckweed growth (measures using ImageJ) and potential tumor induction (this experiment was intended to determine whether agrobacterium can be taken up through duckweed roots (as opposed to wounds) and still induce tumors). The results of this experiment

demonstrate that the A. tumefaciens has no effect on the growth of the duckweed. The absence of tumors on the duckweed after three weeks of exposure to A. tumefaciens also suggests that A. tumefaciens cannot be uptaken through roots and therefore growing duckweed in agrobacterium-treated water does not cause tumor growth in the duckweed.

Effect of Polystyrene on Tenebrio molitor Development and Mortality

by Sarah Kloss (VI), Cole Morriello (VI), Olivia Telemaque (VI), Ethan Werbel To find better methods of degrading waste that pollutes the environment, researchers have investigated the ability of mealworms to consume plastic.

In our experiment, we tested how diets of differing concentrations of plastic affect the development and mortality of mealworms. We placed ten mealworms on four diets consisting of different ratios of grain to plastic measured by volume (tsp) [ 0:1; 1/3 : 2/3 ; 2/3 : 1/3 ; 1:0 ] and observed them for several weeks. We hypothesized that a decrease in weight and increase in mortality of mealworms on highly concentrated diets of plastic would indicate that mealworms are not suitable organisms to combat plastic pollution. Our results suggest that the ratio of grain to plastic does not affect the development

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of the mealworms. We did not find any statistically significant differences between the weight and mortality of the worms and the concentration of polystyrene in each diet. Therefore, in the larvae stage, mealworms are suitable to perform plastic degradation to reduce environmental pollution.

Effect of Different Saline Concentrations on Lemna minor Growth

by Katherine Xie (VI), Nicolas Zarbin (VI), John Grissinger (V) Lemna minor is a species of duckweed that has been largely investigated for its applications in wastewater purification, animal feedstock, and biofuel production. High plant yield is critical for L. minor’s industrial applications; thus, it is crucial to determine what conditions optimize L. minor growth. We observed varying saline concentrations’ impact on L. minor growth. For two weeks, L. minor from the same stock source was cultivated in wells. Growth was analyzed at three different time intervals by measuring the area of the duckweed mass. Image analysis demonstrated that the duckweed typically experienced more growth in water at specific salt concentrations than regular tap water. However, we found exceptions to this pattern at 10 and 35 mmol/L.

Our findings provide insight into the optimal conditions for efficient growth of L. minor.

Efficacy of Venturia inaqeualis on Malus domestica

by Allie Colella (V), Julia Covello (V) Prior genome research conducted on Venturia inaequalis, the causal agent of apple scab in orchards throughout the world, concluded that several varieties of apples contain a resistance gene that can withstand the spread of the disease. A consequent loss of host resistance has also been reported, indicating the fruit is further unable to resist the growth of the disease.

This research reveals that identifying a resistance gene in a particular variety of Malus domestica will not indicate whether or not the type is resistant to V. inaequalis. To determine the ranging susceptibilities of 6 different apple cultivars, we bored holes into each apple variety, inserted fungus-infected agar isolated from previously diseased apples, and calculated the spread of the disease on the fruit’s surface. Our results reveal

that Pink Lady, Gala, Fuji, and Red Delicious apples are more susceptible to the disease than Honey Crisp and Granny Smith apples, which we found are resistant to the spread of apple scab.

These results provide insight to farmers intending to maintain a healthy apple orchard without the need for costly and ineffective pesticides.

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Methods in

Molecular Biology Research.

Summary of Research Class

by Aanya Patel (VI), Belinda Poh (VI), Evan Berger (VI), Kenan Mushayandebvu (VI), Kristin Osika (VI), Meghana Pentayala (VI), Margaret Zachary (VI), Ryan Arrazcaeta (VI), Ulysses Smith III (VI), Molly Parker (VI), Luca Pizzale (VI), Diana Severineanu (VI), Maya Khan (V), Maddie Humphreys (V), Courtney Caputo (V), Adam Bauhs (V)

The BASIL (Biochemistry Authentic Scientific Inquiry) Biochemistry Curriculum allows students to conduct novel research on proteins with known structures but unknown functions using computational and laboratory techniques. The Pingry School’s Methods in Molecular Biology Research class uses the BASIL curriculum and allows students to design experiments to characterize proteins that have predicted enzymatic properties. Students may also enroll for a second year of the class to conduct an independent research project with instructor approval.

Figure 1: The structures of the proteins under investigation and their PDB ID code. These are some of the proteins are being studied by students in the Research 1 and Research 2 classes.

(From top left to right): 3B7F by Adam Bauhs, 4DIU by Ulysses Smith and Kenan Mushayandebvu, 2O14

by Margaret Zachary and Meghana Pentayala, 2QRU by Belinda Poh, Ryan Arrazcaeta, and Evan Berger

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Dnase1L3 Mutations Provide Novel Insight into the Catalytic Mechanism of Dnase1 Family Endonucleases

by Kristin Osika (VI) DNA degradation is a key post-cell death process driven by the Dnase1 family of endonucleases.

One such enzyme, Dnase1L3, functions primarily in the liver. While its activity is known to prevent the onset of Systemic Lupus Erythematosus (SLE), the catalytic mechanism of Dnase1L3 remains disputed. Prior research has implicated two catalytic histidines as catalytically necessary, but mutations have not been tested to date. By transforming, inducing, purifying, and conducting an activity assay, here I plan to test the impact of a proposed catalytic site mutation, H155A, on the impact of Dnase1L3 function. Assay results should illustrate that while wild type (WT) Dnase1L3 effectively degrades DNA, H155A mutants experience loss of function in vitro. Understanding Dnase1L3 catalytic site mutations contributes to a growing body of research aimed at understanding not only Dnase1L3 activity, but also the onset of SLE and other autoimmune conditions in humans.

Figure 1: A model created by Kristin Osika testing the impact of catalytic site mutation, Hr55A on the impact of Dnase1L3 function.

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Bioinformatic Characterization of 3L1W

by Diana Severineanu (VI)

3L1W is one such protein selected from the Protein Data Bank (PDB) with an unknown function, but a known structure predicted to be a hydrolase. Using the BLAST and Pfam programs, which compare proteins based on amino acid sequences, the top results were all shown to be part of the endonuclease/exonuclease/phosphatase enzyme class, giving 3L1W a predicted EC number of 3.60.10.10. This EC number would later be used in the program ProMol to compare possible active sites. The DALI program, which compares proteins independent of sequence and based on 3D structure, was also used and returned similar results to BLAST and Pfam. In vitro, BL21(DE3) Competent E. coli cells were successfully trans-

formed with the 3L1W plasmid, and the protein was expressed using OvernightExpression autoinduction medium. Cells were lysed and protein was purified using buffers of our design, and results were visualized using SDS-PAGE gels.

Due to the disruption of the 2020-2021 school year due to the COVID-19 pandemic, I could not fully confirm our proteins’ predicted functions.

This year, I aim to continue to determine the exact function of 3L1W by using various assays to test substrate affinity, rate of activity, and more, as well as model the reaction computationally and gain a deeper understanding of protein structure and its strong correlation with function.

Figure 1: A model of protein 3L,W from the Protein Data Bank

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MAPS Team.

The Role of Ideonella sakaiensis PETase in the Degradation of PET Plastics: a Structural Comparison of the Wild Type and S238F/W159H Double Mutant

by Britney Alfieri (IV), Madeline Alfieri (IV), Miles Kelly (IV), Siyara Kilcoyne (IV), Lauren Poprik (IV), Ananya Sanyal (IV), Ally Smith (IV), Mehr Takkar (IV), Rohan Variankaval (IV) Advisor: Dr. Pousont Polyethylene terephthalate (PET), a synthetic polymer, is one of the most commonly used plastics today and can be found in over 97% of packaging materials. PET’s durability as a plastic also makes it resistant to biodegradation. According to ​​the National Association for PET Container Resources, only 29.1% of PET packaging waste was recycled in 2018, leaving 27 million tons of plastic in landfills. These alarming statistics indicate new technology to process discarded PET is needed. A recently discovered bacterium, Ideonella sakaiensis 201-F6, expresses the enzyme PETase, which binds and hydrolyzes PET. The structure of PETase closely resembles enzymes in the cutinase and lipase families. Similar to cutinases, PETase has an a/beta hydrolase fold, but with a more open active-site cleft. The catalytic residues in PETase’s active site hydrolyze PET plastics into intermediate molecules which are then further broken down by the enzyme MHETase. Wild type PETase breaks down crystalline PET inefficiently, due to the rigid and dense structure of PET. In order to increase PETase efficiency, scientists altered the active site of PETase. As a result, they discovered that the S238F/W159H double mutation narrowed the binding cleft, making the enzyme more active and therefore more efficient. The Pingry School’s

MSOE Center for BioMolecular Modeling MAPS Team used 3-D modeling and printing technology to examine structure-function relationships of both the PETase wild-type and S238F/W159H double mutant. These models give an in-depth view of the structure of PETase and illustrate how it can be utilized to degrade the highly prevalent PET plastic that litters our planet. Furthermore, comparison of the wild-type and double mutant structures provides molecular level details about structural changes that can enhance the performance of this enzyme even further.

This perspective and knowledge could lead to further advances in biotechnology solutions to the degradation of PET and related polymers.

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Figure 1: The PETase protein model from Ideonella sakaiensis without ligand

HIRT.

Humanities Independent Research Teams

American Modernism

by Martine Bigos (VI), Nicole Moncada (VI), Mirika Jambudi (V), James Thomas (V), Julia Eng (IV), Sriya Tallapragada (III) Advisor: Mr. Burkhart “That was when I learned that words are no good; that words don’t ever fit even what they are trying to say at.” Or so says Addie Bundren in William Faulkner’s As I Lay Dying, as she grapples with the relationship between language and human experience. The research of the American Modernism HIRT has similar concerns. In what way does fiction position itself as a form of knowledge?

Where does it fail to match experience and where does it generate its own? What does fiction actually represent, if anything? What tensions exist between fiction and historical and artistic records?

In past years of the project, we used selected works by modernist American author William Faulkner (As I Lay Dying, The Sound and The Fury), along with selected philosophical works, to develop a framework for thinking about literature. We particularly looked at how Faulkner’s prose encourages, and perhaps demands, an experience that mirrors how we experience the world: full of (re)processing memories, shifting perspectives, andelements ambiguity.

We are currently researching the philosophical and literary inspirations for Modernism, particularly the novel Moby Dick and selected works of German philosopher Friedrich Nietzsche. We are completing short fiction writing prompts that reflect these works in hopes of creating a collection of modernist stories.

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Cultural Racism and Its Imperceptible Impacts

by Emily Shen (VI), Leila Elayan (VI), Lailah Berry (VI), Fara Odunlami (VI), Meher Khan (V), Victoria Ramos (V), Laura Liu (IV) Advisor: Ms. Dunbar When examining racial justice in the 21st century of American society, numerous questions may arise. What marked the beginning of racial inequalities in this country? What were the forces created by the psychological desire to create a racial hierarchy? How was the said desire to create a racially unjust society conveyed to individuals, and how were those individuals’ views transformed due to the resulting societal pressure? This project aims to discuss the fundamental ideas of cultural racism by analyzing the power dynamics between the rulers and the ruled, between the elite or hegemonic class and

the subaltern. In our main discussion, we’ll take a closer look at postcolonial theories of Orientalism and cultural hegemony proposed by Edward Said and Antonio Gramsci, as well as modern-day society in the light of centuries of imperialist influence. Then, members are encouraged to explore and collaborate with each other to present the impact that the colonial empires had on shaping our present-day world views. Through methods such as art analysis, members will then present their work through different media.

Creativity During Covid-19

by Maddie Humphreys (V), Caroline Rewey (V), Mary Claire Morgan (V), Courtney Caputo (V), Lulu Field (V), Anjola Olawoye (V), Chelsea Peng (IV), Krish Patel (III) Advisor: Dr. Dickerson It is impossible not to wonder how writers will represent the Covid-19 pandemic in poetry, fiction, theater, and film in the years to come. This project seeks to discover how the pandemic affects artists’ and writers’ creative processes and material. Have these artists written like never before, or has the pandemic disrupted their creative flow? Are they attempting to capture the pandemic experience in their work, and if so, how? Has the pandemic altered their view of humanity and their experiences in the world? In addition to artists in Pingry’s art department, we interviewed the prize-winning author of Everything Asian, Sung Woo, and

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several visual artists, including painter Laurinda Stockwell and abstract artists Reagan and Danny Geschardt. Last year, we made a flipbook to showcase our work. As the pandemic continues, we have extended our research by interviewing printmaker and former Pingry art teacher Jennifer Mack-Watkins, Pingry’s 3D art teacher, clay working/woodshop artist Seth Goodwin, interior designer Beth Field, and Abby Park, current FIT illustration major.

IRT.

Independent Research Teams

Accelerated Molecular Dynamics for Complex Physical Systems

by Jack Angell (V), Asher Matthias (V), Laura Liu (IV), Hansen Zhang (IV), Dr. Chu Molecular dynamics (MD) simulation is a versatile technique for analyzing physical systems. However, runtime for MD simulations increases dramatically when the analyzed systems become more complex. This also makes running MD simulations using supercomputing facilities cost-prohibitive.

Thus this research seeks to develop an accelerated MD scheme on personal computers through the use of graphics processing units (GPU). The accelerated MD scheme will also be employed for analyzing various complex physical systems such as crystal lattices and quantum mechanical objects.

Figure 1: Crystal Lattice

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Artificial Collagen as a Biomaterial

by James Houghton (VI), Michelle Lee (VI), Alexandra Drovetsky (V), Kirsten Lytle (V), Annabelle Shilling (IV), Theodore Strelecky (IV), Dr. Haven Collagen is the most abundant protein in the human body and plays an essential structural and functional role in many physiological and pathogenic processes. At the molecular level, collagen consists of three polypeptide chains that form a secondary triple helical structure.

These subunits then self-assemble in a lateral staggered association to create fibrils with a unique 67 nm gap-overlap repeat known as a D-period. The unique conformity and behavior of the collagen molecule arise from the repeating Glycine-X-Y pattern in its primary structure.

The current goal of our project is to maximize the yield of collagen mimetic peptides that model the physical and chemical properties of natural type I collagen. We are currently utilizing a pre-existing peptide, named V-F877, which is 222 amino acids

long and has a V-domain, a trimerization domain found in bacteria that is known to be integral for optimizing artificial collagen production. We will compare the yield of the V-F877 sequence with and without the V-domain to understand better the V-domain’s role in improving collagen yield.

Currently, collagen is most commonly purified from animals for the medical field. This process is expensive and often results in a high degree of variation. Generating collagen mimetic peptides that successfully replicate human collagen could provide a safer, pathogen-free, and cost-effective alternative. We plan to provide critical research on these foundational steps to deepen knowledge of collagen’s structure and determine the best methods of creating viable artificial collagen that can be used as a biomaterial.

Figure 1. Hierarchical Organization of Collagen Structure (A. Collagen Fibrils under an Electron Microscope, B. Staggered and Intertwined arrangement of triple helices within a fibril, C. Different stages of Fibrillogenesis from the Primary Structure (Bottom))

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Drover: Drone-Rover Communication for Pathfinding

by Samuel Henriques (VI), Diego Pasini (V), Charles Jiang (V), Ayush Basu (V), Shaan Lehal (V), Nicholas Meng (IV), Dr. Jolly The integration of an unmanned aerial vehicle (UAV, or drone) and an unmanned ground vehicle (UGV, or rover) into a single autonomous system (a drover) is an enticing prospect. The use of drovers is currently being explored for cold trailing, garbage collection, search and rescue, and replenishing post-wildfire terrain. This project is concerned with building a drover that is cheap, effective, and capable of pathfinding in unfamiliar environments. The drone’s global view complements the rover’s local view. The drone component has a clear view of the entire path, enabling it to calculate the shortest, most efficient path around long-term obstacles. The rover must communicate and follow these instructions while making short term decisions and dealing with obstacles at a closer view than the drone.

The primary challenge in the development of this pathfinding system is the implementation of a multi-robot simultaneous localization and mapping (SLAM) protocol which can integrate the perspectives of the drone and the rover into a global map in real time. This project is currently developing a solution implementing an Extended Kalman Filter SLAM scheme which works with 2D images captured by a downward-facing camera on the drone and the states of both the drone and rover based on GPS and other sensor readings. In addition to using SLAM, the drover must be able to effectively maneuver through unknown environments. As such, this project proposes a cooperative pathfinding protocol called Longterm Direction, Short-term Correction (LDSC), wherein the drone draws and dictates long-term paths for the rover, and the rover identifies smaller objects in its path and dodges them in the shortterm. The drone’s path-planning ability involves turning images into node maps that are scanned from a starting point to an end point using the A* search algorithm. In the process of calculating

the path based on a drone image, the Long-term Direction pixel path must be converted into GPS coordinates for the rover to follow. In developing the drone end of the LDSC algorithm, it was found that weighing the nodes by color, rather than predetermining the path viability of each pixel on an absolute basis, proved more effective for pre-planning paths on roads. In order to minimize camera distortion error, the checkerboard camera calibration algorithm is used to generate distortion coefficients for the drone’s camera. To convert the pixel path into a GPS path, the drone calculates the real-world distance between each pixel coordinate of the path and the center of the image. The Vincenty formula is used to convert the distance measurements into GPS coordinates, yielding results that are accurate to the millimeter. Ultimately, the LDSC algorithm will be able to generate these paths on continuous footage, translate them into motion instructions for the rover, and allow the rover to alter these motion instructions efficiently in the short term.

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Drone

Rover

A*

Distance Calculation data

Effect of Dietary Restrictions on the Lifespan of Drosophila melanogaster

by Bella Vieser (VI), Kimberly Wang (VI), Mirika Jambudi (V), Lauren Kim (V), Kate Marine (IV), Dr. D’Ausilio, Dr. Pousont Previous research has proven that a calorie-restricted diet increases the lifespan of Drosophila melanogaster. Our experiment aims to test the proportionality between the diet-induced lifespan elongation and the flies’ cognitive skills.

Our experiment consists of two groups of flies: a control group fed molasses-based fly food (control) and an experimental group fed diet food (dilution of the regular food with an agar solution). After breeding flies on the two diets, we initially planned to test the memory of both groups using an odor T-maze.

Last year, we ran a trial to test the lifespan of flies fed either the control diet or the calorie-restricted diet. Ultimately, the data sup-

ports our hypothesis that a calorie-restrictive diet correlates with an increase in the lifespan of drosophila melanogaster (Figure 1).

After our initial hypothesis has been confirmed, we planned to continue to experiment with the odor T-maze to determine how cognitive and memory abilities correlate with this proven extended lifespan. However, we could not follow through with this plan due to limited materials.

Therefore, this year, we continued to keep our fly stocks running as we planned to test the effect of new dietary supplements on the flies: including additional phosphate, nitrate, and sulfate.

Figure 1: The results are a composition of data from 9 different vials: 4 control vials and 5 calorie-restricted vials from the lifespan experiment. The total number of flies remaining in control and calorie-restricted vials were counted over time, and the data standardized to show the percentage of flies remaining in each group over a period of 100 days. Over time, the difference between the number of flies alive in the control and calorie-restricted vials increased. As the number of control flies approached zero, the calorie-restricted diet flies still had 1/3 of their total population alive. The data indicates that a calorie-restricted diet keeps the flies alive for longer than the control diet does.

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Expression and Analysis of Fluorescent Fish Proteins in Vitro

by Shannen Gallagher (VI), Rachel Zhang (V), Morgan McDonald (V), Ethan Boroditsky (IV), Annika Shekdar (IV), Dr. Fried There is an ever-growing need for improved fluorescent tags in the medical field. The discovery of the Japanese freshwater fluorescent eel (Anguilla japonica) in 2013 is the first evidence of fluorescent protein in vertebrates—one step closer to more efficient fluorescent applications. Since then, several fluorescent marine organisms have been identified, including lizardfish, scorpionfish, seahorses, and even turtles. We currently aim to purify red fluorescent fatty-acid binding proteins from scorpionfish (Scorpaenidae) and lizardfish (Synodontidae). To fluoresce, these proteins require the cofactor bilirubin. However, researchers are currently unable to bind biliru-

bin in vitro and achieve fluorescence. So far, we have been able to express the lizardfish protein with high purity, so the next step for our team is determining how to bind bilirubin to the protein and attempting to induce fluorescence.

Anguilla japonica

Metagenomic Analysis of a Compost Microbiome

by Ryan Arrazcaeta (V), Ainsley Ellison (V), Milenka Men (IV), Max Watzky (IV), Dr. Sparrow

Metagenomics is the study of genetic material taken from a population of organisms. The composition of microorganism populations change in response to factors in the environment such as temperature and pH level. By performing metagenomic analyses of genetic material from the Pingry composter, we hope to understand how the microbial community of the composter changes in response to these conditions. From the 2019-20 school year through the present, the team has harvested nearly three dozen compost samples and successfully isolated DNA, submitted DNA for sequencing, and analyzed the data for useful or interesting information on the composter’s complex microbial community.

During the 2021-22 school year, the team collected compost samples approximately once per month, both at the distal end (finished compost) as well as 10 feet in from the distal end. To further characterize the composter environment, we used HOBO data loggers to measure the internal temperature of the composter. The data loggers

naturally move through the composter over the course of 100 days, taking the temperature once every hour. The team added a second data logger to the composter during the 2021-2022 school year at a 30-day offset from the original one, allowing the team a more complete look at internal temperature at different locations within the composter. Further characterization of the composter material includes measurement of pH on fresh samples. After collection, samples are dried in a dessicator for several days, and then DNA is extracted using a Qiagen power soil pro kit. Sample purity is tested using agarose gel electrophoresis and spectroscopy, and samples unfit for NGS are discarded. The rest of the samples are sent to GeneWiz for next-generation sequencing, which returns several different kinds of data, which we then analyze using statistical significance tests and trend analysis. Our research will provide a natural history of the composter and generate practical knowledge for future operation of the composter.

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PETase - Plastic Degrading Enzymes

by Tyler Headley (VI), Maya Khan (V), Juliana Amorosi (IV), Sebastian Talarek (IV), Dr. D’Ausilio, Dr. Pousont The United States of America only recycles about 35% of its total waste, sending nearly 150 million tons of trash to landfills each year. There, it takes up to 450 years to fully biodegrade. Often, it ends up invading oceans or forests, where it poses a serious threat to native species. Polyethylene terephthalate (PET) is one of the most widely used plastics in the United States, but only 31% of the 3.1 million tons of PET produced in America annually are recycled. It is necessary to find an efficient way to recycle plastic in order to mitigate the damage done to our planet.

Our team works with PET depolymerase (PETase), an enzyme with the unique ability to break PET polymers into monomers, which can then be reassembled into new plastic. This nuanced recycling method is termed “biochemical recycling” and has become an increasing focus of research—however, it requires optimization before it can be implemented commercially.

Because the severity and pervasiveness of plastic-related damage to the environment is growing, the scientific community is eagerly seeking to improve PETase through mutation–specifically, its efficiency and thermostability. Our goal is to identify and investigate various mutations, and assess the change in efficiency and thermostability that each mutation offers. After researching mutations that demonstrate promise for accomplishing these two goals through scientific literature, we obtained various PETase mutations through our partnership with Deerfield Academy. Deerfield created these mutations using site-directed mutagenesis, a common method to induce mutations, utilizing a primer to confer a mutation in double-stranded plasmid DNA. To replicate the mutated plasmid, we transform it into NEB 5-alpha cells and grow colonies on LB-AGAR plates with ampicillin. We harvest the colonies using inoculation loops before miniprepping: lysing the cells and discarding all contents besides the mutated plasmid.

Then, we transform the mutated DNA into BL21 cells and induce protein expression with autoinduction to create copies of the mutated PETase enzyme. Finally, we perform cell lysis and purification to obtain the pure enzyme. To determine the efficiency of this mutated PETase enzyme, we plan to develop an assay. This assay will reveal whether, and how efficiently, our PETase mutants are capable of biodegrading PET. In the coming months, we hope to make more progress researching and documenting the effects of different mutations on PETase, which will further advance the scientific community’s knowledge of the enzyme, and contribute to the global effort to make recycling more efficient and widely used.

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PETase Structure

Protein Purification of Shigella dysenteriae Transcription Factor VirF

by Brian Li (VI), Arjun Sen (VI), Maddie Humphreys (V), Daniela Karnaugh (V), Dr. Pousont, Dr. D’Ausilio Abstract

Methods

Shigella is a pathogen that is a leading cause of bacterial foodborne illness worldwide. The virulence cascade of shigellosis requires one of the AraC superfamily transcription factors, the protein VirF. Other members of this superfamily, such as ToxT, which is involved in the virulence cascade of Vibrio cholerae, show inhibition of DNA by binding a small fatty acid. We wish to determine whether or not VirF can be inhibited in a similar manner, and are therefore working to express, purify, and ultimately solve the crystal structures of VirF. Solving the structures will help to reveal the mechanism of transcriptional regulation in S. dysenteriae and could lead to development of novel treatments for Shigella related illnesses.

Our team will be working at the laboratory at the Pingry School under the guidance of Dr. Jennifer Pousant and Dr. Morgan D’Ausilio. We received the VirF plasmid dried on filter paper from Dartmouth’s Kull Laboratory in January 2021. We rehydrated the plasmid in distilled water, and proceeded to conduct bacterial transformation to replicate the plasmid. Then, through a miniprep, we isolated the plasmid from the bacteria and sent the plasmid to a lab for sequencing.

We then analyzed the sequencing/plasmid map to confirm there are no mutations in our sample.

Introduction

Future Steps

The Shigella bacterium causes sh igellosis, specifically mammalian gastroenteritis. An estimated 80-165 million cases of gastroenteritis due to Shigella species occur globally each year. Symptoms vary from inflammation of the intestines causing diarrhea to nausea and vomiting. Shigella also kills over 600,000 people annually. Infection begins with the ingestion of contaminated food or water. Many of the genes required for intestinal penetration, invasion of host cells, and intestinal and diarrheal disease are carried out by the major regulatory protein VirF located on the PINV as part of the regulatory cascade. This member of the AraC protein family is a DNA-binding protein that activates toxicity factors icsA and virB genes. These genes are then able to activate other downstream effectors involving invasion of the cell wall and host infection. Thus, inhibiting VirF would inhibit the whole virulence cascade of shigellosis.

Our goal is to isolate the VirF proteins which will help to understand the mechanism of transcriptional regulation in S. dysenteriae. Understanding how to possibly inhibit these structures could lead to development of novel treatments for Shigella related illnesses. Our next step is to run an autoinduction to express our target protein. We will then purify our protein through affinity chromatography.

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Quantifying the Effect of Structure on Neural Network Training

by Emmet Houghton (VI), Katherine Xie (VI), Olivia Taylor (V), Evan Xie (IV), Divya Subramanian (IV) Dr. Jolly As artificial neural networks become increasingly ubiquitous in emerging technologies across a vast array of industries, the efficiency and accuracy of these machine learning tools are of utmost importance. It is widely accepted that the structure of neural networks has an impact on their ability to interpret data, but the nuanced implications of network topology are largely not understood. The ShallowMind project seeks to answer this central question: how, and to what extent, does the structure of a neural net affect the network’s efficiency and accuracy?

This year, we have been exploring this question by building evolutionary networks and comparing their efficacy to that of traditional neural networks. While traditional networks have a fixed structure and alter their weights and biases throughout training, evolutionary networks undergo a rigorous process similar to that of biological evolution that not only enhances weights and

biases, but also network structure. The genetic algorithm dictating this optimization, “NeuroEvolution of Augmenting Topologies” (NEAT), reproduces, mutates, and trains a population of unique networks until a network performs sufficiently on validation data. By comparing the performance of traditional networks to that of these final evolved networks, we will determine which tasks evolutionary networks are more suitable for solving.

Additionally, we hope to gain insights into the effects of network structure through examining the resultant structures of the evolved networks.

In the future, we will be experimenting with tuning the algorithm’s hyperparameters to further boost the network’s performance and developing a series of systematic methods for comparing performance between traditional and evolutionary networks. We also aim to enter data science competitions to quantitatively compare our networks to those of the broader research community.

Quantifying the Phototactic Memory of Chlamydomonas reinhardtii

by Christine Guo (VI), Sarah Kloss (VI), Maximus Liu (V), Sejal Patel (V), Leo Xu (V), Dr. Samadani Phototaxis, the behavior describing the movement of organisms towards or away from light, plays a crucial role in cell survival, allowing some organisms to find ideal locations for photosynthesis (1). Chlamydomonas reinhardtii, a unicellular green algae that phototaxes towards green light, serves as a popular model organism for phototaxis. C. reinhardtii has been studied at the molecular level to examine its use of Rhodopsin proteins for phototaxis as well as the population level to examine collective motility. We have designed an algorithm to track cells in two and three dimensions, which has allowed us to quantify the be-

havior of C. reinhardtii. We quantify this behavior through the use of a phototaxis index, a value ranging from -1 to 1 that describes the correlation between the cell’s movement and the location of the light source. We aim to determine if C. reinhardtii has phototactic memory, that is, if it continues to move in the direction of a light source after the light source has been turned off. The existence of a phototactic memory would prompt further research into the mechanism allowing for this memory as well as the potential impact of this memory for other functions beyond motility.

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Smart Heart: Deep Learning-Based Analysis of 12-Lead Electrocardiograms for Cardiovascular Disease Risk Prediction

by J.P. Salvatore (V), Evan Wen (V), Alan Zhong (IV), Dr. Jolly

Heart disease is the leading cause of death worldwide, accounting for one in every four deaths in the United States. Electrocardiograms (ECG) are the most common cardiovascular diagnostic procedure with over 100 million samples obtained every year in the U.S. alone. While previous works have developed deep learning systems for accurate diagnoses of ECGs, there has been insufficient study on predicting future diagnoses of heart disease. Our work is focused on developing a novel deep learning architecture for highly-accurate ECG classification which can be adapted to the untapped problem of predicting diagnoses. The model’s architecture consists of learning image representations of each ECG before utilizing various image classification pipelines. After applying multi-domain image-to-image translation on image representations of the ECGs, we compare feature maps between healthy and abnormal ECGs. We evaluate our work using data from the PhysioNet/Computing in Cardiology 2020 challenge and a set of printed ECGs from a local cardiologist. Since these are in printed format, we also developed MATLAB scripts to transform scans into processable data for our models. Our work is the first to estimate the likelihood for later diagnosis of heart disease, giving patients an incentive to take action for their heart’s future health.

Figure 1: An overview of the proposed network architecture for 12-lead ECG classification.

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The Effect of KIF11 Activity on YAP Localization

by Luca Pizzale (VI), Elodie Wardle (VI), Matthew Oatman (V), John Grissinger (V), Katia Krishtopa (IV), Dr. Sparrow

Our project is focused on the relationship be tween two proteins, KIF11 and YAP. KIF11 is a kinesin, otherwise known as the “tractor of the cell”, which is responsible for pulling chromosomes apart during mitosis. YAP, on the other hand, is a transcription factor that activates cell growth and sometimes plays a role in cancer. Inactive YAP has phosphate groups and is located in the cytoplasm, but when the Hippo pathway dephosphorylates YAP, it moves into the nucleus and becomes active. We’ve been in touch with Dr. Gao and his lab at Rutgers University, where they hypothesize that KIF11 and YAP are linked together via the interactions of other proteins. Our goal is to test this hypothesis by culturing SW480 cells, manipulating their KIF11 activity, and performing western blots to observe changes in YAP activation.

The discovery of a new way to manipulate YAP could prove helpful in targeted cancer therapies.

During the 2018-19 school year, we obtained SW480 cells from Dr. Gao’s lab and began to culture them. These are human colon cancer cells that, when properly maintained, are capable of multiplying indefinitely and are therefore very useful to our study. Once we had cultured enough cells, we stored some away for future use and used the rest in our first western blot.

This western was done to familiarize ourselves with the procedure, but also to confirm the presence of YAP and KIF11 in our cell samples.

as the concentration of Ispinesib increases, nuclear YAP decreases, suggesting that the inhibition of KIF11 prevents YAP from being activated.

In the 2021-2022 school year, we have cultured cells to increase our supply after previous samples were destroyed in the freezer. We have also run gels (to practice) and a western blot. Although the western blot from 2020 suggests that inactive KIF11 results in inactive YAP, we cannot confidently make that assertion based on only one experiment. Going forward we would like to repeat this experiment to see if we can reproduce the same results. And, after communicating with Dr. Gao about what we observed, he suggested that we also apply two other controls. The first would be to re-probe a blot with anti-histone antibodies.

Histones are proteins only found in the nucleus, so staining for them would tell us if we succeeded in separating the nucleus from the cytosol. Second, we would also want to probe a future blot with an antibody exclusive to phosphorylated YAP. This would let us know if manipulating KIF11 impacts the Hippo pathway’s dephosphorylation of YAP.

In the 2019-2020 school year, our main goal was to reculture our cells, inhibit KIF11, and then perform a second western blot. Prior to beginning the western blot, we titrated the cells with varying concentrations of a drug called Ispinesib in order to inhibit KIF11. And, considering that YAP’s location changes with its activation, we decided to split the cell’s nuclear components from the cytosol via centrifugation. We observed that,

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Club Highlights.

by Lauren Kim (V), Maria Loss (V)

Anatomy Club provides students with the opportunity to learn the anatomy of various organisms through hands-on dissections. So far, we have dissected fetal pigs, sharks, snakes, hearts, brains, and more. This club is dedicated to fostering interest in and preparing students for potential careers in the biomedical field. Dissections are meant to enhance students’ curiosity and knowledge of this fundamental aspect of biology. Anat-

omy Club is advised by Ms. Torres and led by student leaders Maria Loss (V) and Lauren Kim (V). Overall, through this club we hope to provide memorable learning experiences that instill, in students, a love for science and exploration.

by Maria Loss (V), Kristin Osika (VI) Ainsley Ellison (VI), Sam Wexler (VI), Maya Khan (V), Meher Khan (V), Grace Stowe (V)

FYI Sci is a student-run organization which aims to convey scientific research discoveries to the Pingry student body through the creation of blogs, presentations, videos, and more.

Even in light of the COVID-19 pandemic and Pingry’s transition out of the hybrid learning model, the 2021-2022 school year has been an incredible success for FYI Sci. The club’s newsly structured departments are overseen by President Kristin Osika (VI) and Vice President Maria Loss (V) by

organizing resources and creating forms, presentations, and emails, culminating in a cohesive plan for the year. In September, FYI Sci accepted over twenty new members into our three departments: blogs, videos, and podcasts and slideshows. Our members engaged in department-specific projects covering a broad variety of topics in addition to running an all school Trivia Competition.

More information can be found on our website: https://students.pingry.org/fyisci/social-media/.

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This school year has been the most prolific to date for our blogs department, which is run by Ainsley Ellison (VI) and Meher Khan (V). Members have written about topics ranging from “The Symbiotic Infection of Herpes” to “Discoveries in the SARS-CoV-2 Genome” throughout the year. Blogs can be complex and time-consuming projects, which makes the incredible success of the department all the more impressive!

Under the leadership of Grace Stowe (V), the videos department was also incredibly productive.

They released a video asking Pingry students why the sky is blue, joining science and fun to present to bring the community together. Members are working on a special video to promote research at Pingry in preparation for Pingry’s Research Week.

In the podcasts and slideshows department, led

cent projects from all departments are summarized and linked in a single publication for the Pingry community. At the heart of communications are Instagram posts, for which members find photos and write captions describing a topic of interest. This year, we posted about telomeres, chromotherapy, tectonic plates, and more.

Instagram also serves as a platform for FYI Sci to promote other projects, including podcasts, blogs, and upcoming events, and we recently added a Linktree to consolidate supplementary links and resources.

In addition to social media, FYI Sci also maintains its own website, which is frequently updated by our Head Web Editor, Sam Wexler (VI). Sam has posted new content, maintained website security, coded embedded forms, and ensured that our website looks wonderful.

Finally, FYI Sci will be presenting at Research Week! Our team will lead multiple experiments, engaging students in hands-on activities such as Egg Drop Contest and Strawberry DNA Lab. During Research Week, we will also assuredly celebrate our members and their projects, as they are the core of FYI Sci. Building off of the progress made during this markedly strong year, FYI Sci hopes to continue to expand its reach for years to come, guided by our motto, “Bringing science to you!”

by Maya Khan (V), members write scripts, and record in groups for seven to ten minutes. Our latest podcast, “Gene Therapy,” and past podcast recordings can be found on our website and on Soundcloud. The team is additionally working to produce slideshows to promote the club around the school to further involve the community.

This year, Sam Wexler (VI), Ally Smith (IV), Coco Simon (IV), and Monroe Russell (IV), the Media Managers, work to amplify community outreach while maintaining a strong social media presence. Katia Krishtopa (IV) has established “fact of the week” on the Upper School announcements document, and has created and distributed a bimonthly newsletter, in which re-

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by Mirika Jambudi (V), Kristin Osika (VI), Sarah Gu (IV), Evan Xie (IV), Annabelle Shilling (IV) The Pingry Community Research (PCR) journal highlights student contributions to novel scientific research and promotes students’ interest in science, while emphasizing the importance of critical thinking and the diverse, interdisciplinary nature of scientific research fields. Though PCR only included AP Biology and IRT papers in prior years, our submission criteria have recently become more inclusive to reflect our contributors’ varied interests and their diversity in submission format; we now accept investigative articles, commentary, summer research, and more.

This year, Pingry Community Research produced this special journal issue, highlighting the work of students who are presenting at Research Week!

In the future, Pingry Community Research hopes to increase reader accessibility through the use of digital media. We recently began developing a website, which provides background information on Pingry Community Research and highlights current and past journal issues. In the future, we hope to increase our online presence further.

Girl Code

by Olivia Taylor (V), Zala Bhan (V) Girl Code is a club focused on empowering women at Pingry to explore and delve into computer science. We have held a wide range of events in the past focused on everything from teaching programming languages to hosting female speakers with experience in CS-related nonprofit or industry work. However, our main highlight is always bringing our club members to the annual FemmeHacks, an all-female hackathon at the University of Pennsylvania! While FemmeHacks has traditionally been held in-person at the Pennovation Center, the hackathon was remote this year due to the pandemic. Girl Code had a great time participating virtually, with two teams submitting projects. Ananya Sanyal (IV), Zala Bhan (V), Olivia Taylor (V), and Divya Subramanian (IV), coded an application called Charity Connect, which used Google Cloud and Android Studio. Through Charity Connect, they aimed to connect people with charity drives in their communities. Their project won second place for Best Use of Google Cloud, awarded by a team of Google engineers.

Laura Liu (IV), Annabelle Shilling (IV), Sia Ghatak (III), and Carolyn Zhou (III) created an interactive website that highlighted the achievements of LGBTQ+ historical figures. It recommended similar figures based off of the user’s selection of filters such as gender, time period, race, and sexual orientation.

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Robotics

by Dwayne Bazil (V), Olivia Taylor (V), Diego Pasini (V), Laura Liu (IV), Alan Zhong (IV), Keira Chen (IV), Zayed Ali (III), Carolyn Zhou (III), Matt Carrieri (III), Connor Francis (III), Sia Ghatak (III), James Kotsen (III) Do you like robots? Of course you do! Come see Pingry Robotics in action by interacting with our world-championship robots; they can do pullups, climb stairs, make half-court shots, raise discs ten feet in the air, and more! At our exhibit you will be able to drive some of our robots, see our robots that got to world championships, and ask questions to the brilliant team behind them.

The Pingry robotics team builds two robots to complete specific tasks in time-constrained competitions to win points. We compete in FTC (FIRST Tech Challenge) and FRC (FIRST Robotics Competition) along with mentoring the middle school team.

In this year’s FTC competition, points are scored by placing cubes and balls onto hubs for the respective teams. After going through our intake, a set of entraption stars, the cargo is transported up a linear slide and dumped out onto the different levels of the hubs. To turn the carousel—a disk on the corner of the field where rubber ducks are positioned—we attached a spinning wheel, so that the ducks fall into the field to score extra points.

The drivers use gamepads to control the robot.

In this year’s FRC competition, points are scored by launching balls into a central hub and climbing sets of raised rungs that have each rung positioned higher than the last. We use a hopper outtake system to shoot balls 15 feet into the air and have two sets of climbing arms to traverse the rungs. During the autonomous period of the match (the first 15 seconds), the robot drives itself using sensors and cameras which help it move and score, and during the tele-operated period (the other 145 seconds), our drivers use joysticks to control the robot.

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Throughout the past few years we have been using our skills and experience from FTC and FRC in order to help those around us. Despite the challenges brought on by COVID-19 and our graduating seniors, we have embraced Pingry’s values of intellectual engagement, discovery, and innovation to help us continue building amazing robots.

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Email: Mirika Jambudi (V): mjambudi2023@pingry.org Annabelle Shilling (IV): ashilling2024@pingry.org Evan Xie (IV): exie2024@pingry.org Sarah Gu (IV): sgu2024@pingry.org Mr. D. Maxwell: dmaxwell@pingry.org

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The Pingry School 131 Martinsville Road Basking Ridge, NJ 07920 908-647-5555