Homework
Weekly homework submissions:
Week 1 HW: Principles and Practices
First, describe a biological engineering application or tool you want to develop and why. This could be inspired by an idea for your HTGAA class project and/or something for which you are already doing in your research, or something you are just curious about. A biological engineering project I have been passionate about to bring into the world intersects art conservation, biology and design. My goal for the final project portion would be to create a conservation treatment for cultural heritage objects specifically that of ceramics and textiles utilizing synthetic biology.
Week 2 HW: DNA Read, Write and Edit
My initial confusion was trouble shooting how to cut at a specific segments of DNA. On my benchling, I copied a segment of DNA between the desired cut sites and pasted it into a “New DNA sequence” file. This was purely for artistic purposes, however, in lab, I asked our instructor how we would select for these specific sites for future gel art. There were two options: Purchase or design primers to select at the specific site OR
The design above was made using the platform graciously programmed by Ronan. The design is inspired by golden pieces made by Colombian craftsman. I was in Colombia at the time and had the privilege of seeing El Museo del Oro (The Gold Museum) and aimed to replicate some of the features here. I input the coordinates from Ronan’s program into the opentrons file as seen above. There was quite a bit of trouble shooting as the equipment stated in the code was different from BUGSS supply. That caused some coordination issues within the opentrons that was eventually solved. During the lab, the majority of the time was dedicated to trouble shooting these minute bugs and I wasn’t able to run my script. However, Amanda and Joel (our instructor and TA! ((Thank you guys so much)) ran the script after lab hours which yielded this result:
Answer any NINE of the following questions from Shuguang Zhang: (i.e. you can select two to skip) How many molecules of amino acids do you take with a piece of 500 grams of meat? (on average an amino acid is ~100 Daltons) In a 500g piece of meat, there is approximately 7.8x1023 amino acids. In researching for this question, I was unable to find further resources other than previous HTGAA pages.
Superoxide dismutase 1 (SOD1) is a cytosolic antioxidant enzyme that converts superoxide radicals into hydrogen peroxide and oxygen. In its native state, it forms a stable homodimer and binds copper and zinc. Mutations in SOD1 cause familial Amyotrophic Lateral Sclerosis (ALS). Among them, the A4V mutation (Alanine → Valine at residue 4) leads to one of the most aggressive forms of the disease. The mutation subtly destabilizes the N-terminus, perturbs folding energetics, and promotes toxic aggregation.
What are some components in the Phusion High-Fidelity PCR Master Mix and what is their purpose? “Phusion High-Fidelity PCR Master Mix with HF Buffer is a 2X master mix consisting of Phusion DNA Polymerase, deoxynucleotides and reaction buffer that has been optimized and includes MgCl2. All that is required is the addition of template, primers and water.” - NEB
What advantages do IANNs have over traditional genetic circuits, whose input/output behaviors are Boolean functions? IANNs are more “organic” in that though it is still a structured system, it does not operate on a binary code as a genetic circuit does. This makes IANNs more attuned for detecting and accounting for fluctuating systems such as varied metabolic activity and hormone changes. IANNs can also support multiple functions at once while genetic circuits, in order to scale, need be highly specific. An IANNs system is responsive while a genetic circuit is decisive is how I think of it. However, though this sounds great, IANNs is more complex to build.
The highlighted sections are questions I am unsure how to answer, but made the educated assumptions from the lecture and research. Explain the main advantages of cell-free protein synthesis over traditional in vivo methods, specifically in terms of flexibility and control over experimental variables. Name at least two cases where cell free expression is more beneficial than cell production. Mutations are still a phenomenon being understood in a synthetic cell system, however, in a cell FREE system, your risk for mutational error is lessened without an entire cell incorporated. Therapies in the environment and on people without the elevated risk of cellular contact The system is overall less time intensive allowing for reactions to happen in a day rather than a multi-day to multi-week turn around time. So far, the cell free system is projected to be less expensive than cell dependent systems as the labor, components and reaction times are less “problematic”. My assumption is this also allows increases the iterative aspects to experimentation. If the system takes less time with less resources and costs, researchers can directly build upon their experiments quicker. Describe the main components of a cell-free expression system and explain the role of each component.
Week 10 HW: Imaging and Measurment
Please identify at least one (ideally many) aspect(s) of your project that you will measure. It could be the mass or sequence of a protein, the presence, absence, or quantity of a biomarker, etc // Please describe all of the elements you would like to measure, and furthermore describe how you will perform these measurements. What are the technologies you will use (e.g., gel electrophoresis, DNA sequencing, mass spectrometry, etc.)? Describe in detail.
Referencing the cell-free protein synthesis reaction composition (the middle box outlined in yellow on the image above, also listed below), provide a 1-2 sentence description of what each component’s role is in the cell-free reaction. E. coli Lysate BL21 (DE3) Star Lysate (includes T7 RNA Polymerase) - The strain of E.coli used to lysate and extract all of the needed cellular components like polymerases for transcription and ribosomes for translation. There is reduced chromosomal degradation. Salts/Buffer