Projects

These were the projects developed in the Summer of 2018:

pGLO It and Grow It

by Rupin Mittal, Aidan Zhang, Zack Chang

The pGLO plasmid allows E. coli bacteria to glow under UV light. Arabinose allows for the production of Green Fluorescent Protein (GFP) and LB/agar allows the bacteria to grow. In our experiment, we changed Arabinose and LB/agar levels to see if the production of GFP could be changed. This can be observed by the bacteria glowing different amounts. Since the human is limited to qualitative observation, we used computer software called Image J to quantitatively assign values, on a 0-255 scale, to the brightness of the bacteria's glowing. By optimizing our setups, we were able to create mathematical models to model the relationships between GFP production and Arabinose and LB/agar. Using this data we can predict at what conditions will ideally produce the most GFP. We also used TinkerCell to create a digital model of the biological systems. We were also able to create physical logic circuits to model the biological systems. We hope our experiment in optimizing GFP production will lead to more experiments in optimizing protein production.

pGLOgurt

by Julia Picker, Lea Twicken, Hallie Seay, and Nathan Glonek

The idea of this project is to use the pGLO plasmid and parts the procedure for the transformation of E. coli in order to made lactic acid bacteria present in yogurt (Streptococcus thermophilus and Lactobacillus bulgaricus) that floresces. We hoped also to use that bacteria to make yogurt that glows under UV light. We grew bacteria from the yogurt starter bacteria in milk, which we then centrifuged in order to try and isolate the bacteria so that we could transform it. After being heat shocked, the bacteria accepted the plasmid and were producing Green Florescent Protein by the next day! (To our surprise and amazement) Our procedure took parts of BioRad’s pGLO procedure, and combined it with parts of processes we found online for growing lactic acid bacteria in milk for an entirely new procedure that is unique to this project.

Binary-man vs Gradual-man: pDawn of Justice

Testing the effect of different blue light brightness levels on RFP production in pDawn transformed E. coli

by Miles Lee, Ravi Patel, and Kevin Ju

Light-regulated gene expression functions in modified organisms through a system of light-receptors and transcription factors. The pDawn plasmid in transformed E. coli bacteria reacts to blue light and as a result synthesizes red fluorescent proteins. Here, we attempt to determine whether pDawn gene expression is a binary or spectrum by shining different amplitudes of blue light on transformed bacterial cultures and using a pixel imaging program to identify the degree of gene expression (i.e. the brightness of the red glow from red protein). After two trials of light for around 16 hours each, our results unexpectedly showed that the brightest blue lights caused the least amount of red protein production and vice versa. Furthermore, we found that pDawn expression is mostly a binary system, with a small spectrum. Nevertheless, the test results were inconclusive due to limited time and testing apparatus -- more testing is needed.

What a Colorful World

by Nikko Smith and Koko Saadiq

Exploring the impact on different genetic chassis on identical genetic sequences and the study of bacterial growth on plate design.

Grow and Glow

by Airol Anne Ubas and Jenna Rotheram

Our goal is to test the accuracy and efficiency of CRISPR-Cas9. In the presence of streptomycin, E. coli dies because streptomycin inhibits proper ribosomal function. Using CRISPR-Cas9, we induced a point mutation in the essential ribosomal subunit rpsl of E. coli, allowing it to survive in the presence of streptomycin. Then, we tested for possible genetic damage by comparing the transformation efficiency of bacteria with the point mutation and without. CRISPR+ bacteria grew in all of our LB plates with streptomycin and they also had a greater transformation efficiency with the pGLO plasmid.

iTune Device

by Ammerica Barraza, Tannya Tang, Ellen Wang

The production of a protein includes transcription, which is heavily affected by the promoter, and translation, which is heavily affected by the ribosomal binding site. To understand how to control the production of a protein, we tested 9 strains of E. coli with different promoter and ribosomal binding site strengths. IPTG was used as an inducer to cause E. coli to produce beta-galactosidase, an enzyme which acts upon allolactose. SDS was used to dissolve the cell membrane and release this enzyme and put it in proximity to substrate. Instead of allolactose, ONPG was used as the substrate because its products include nitrophenol, which is visibly yellow. A more yellow culture was thus indicative of a higher rate of protein/enzyme production. We discovered that while the strongest promoter and strongest RBS understandably produced the most product and the weakest promoter and weakest RBS produced the least, the relationship between component strength and production was not direct. For example, having a medium RBS and any strength promoter produced more than any strong RBS for the most part. We concluded that having a stronger component made it stickier. While stickiness would ensure binding and transcription/translation, RBS being stickier would increase the time that mRNA binds to a ribosome, causing some lag. The strength of the promoter seemed to be less affected by this behavior because it lies at the beginning of protein production as opposed to in the middle.