Week 1: Principles and Practices

source: wikimedia commons

Genetically Engineered Diatoms to Bind with Building Rubble/Waste

Building materials like cement and brick are difficult to reuse and natural weathering or active demolition leaves behind tons of waste material that remains under-recycled. In a previous project my team from graduate school developed a porous, bio receptive glass using glass waste and I would like to expand upon that research by bioengineering diatoms into a silica scaffold of cement and glass rubble/frits to fuse these waste materials into a new architectural material. Diatoms are an exciting prospect for architecture for their silica frustules, inherent translucency, and their lacy pore structure. I am curious to see if there would be a way to pattern their silica deposits for enhanced carbon sink and particle processing in urban spaces. It would also be beautiful to see the formation of silica deposits depending on sun patterns on site, filling in the rubble scaffold where there is more direct light. It would also be interesting to potentially engineer the directional strength of a diatom-rubble piece and the lace pattern, playing with the idea of directional bias in architecture more theoretically but also for building methods.

Goals:

• When speculating such panels, it is important to keep policy which mitigates the impact of the material creation and installation on the surrounding soil and water biodiversity through run off material and resources used to grow the diatom structures. Diatoms can overpower other microorganisms and limits on the volume of diatom production could help reduce the chances of local ecological harm and biological impact.

• Another goal is to secure a non-stress inducing method of cultivating and sourcing the diatoms for this scale of application.

• This material should be lessening the burden of the construction industry.

Concerns:

• Who would have access to this material?

• How does this material impact the local environment?

• Will users receive this material positively and use it? Or will it be demolished or underutilized?

• Will this create excess burden on access to an organism for this scale? What are the political realities of sourcing this material?

• How do diatoms react with materials like cement, brick, and glass? Are there any reactions between materials that can cause issues?

Actions:

• When casting the rubble diatom mixture, create reusable casts/equipment workflows when possible.

• Work directly with ecologists to determine the site for harvesting and to determine if the site for installation is appropriate in case of run-off or other biological interactions.

• One method of mitigating the environmental impact of sourcing and using diatoms at this large scale could be to focus on cultivation from local areas with diatom overgrowth so that this helps cut down on the environmental impact of current unhealthy ecology.

• Lab testing, especially longevity testing, would help clear up uncertainty regarding the impact of the material on site as it weathers and does through periods of high and low growth. This includes the potential toxicity of the rubble, the biological interactions of the diatoms, and how the ,odified diatoms may change over time through mutations and biomass buildup.

• Community workshops/exhibits as user studies to understand how people would interact with the material (biosecurity of sensory interaction), understand whether they would accept it in their built environment, and educate them on the material itself.

Homework Slide Responses

Professor Jacobson:

1. The error rate for polymerase is 1:10^6. Compared to the length of the human genome of around 3.2Gbp (slide 10), the error rate is minute, occurring perhaps once in one genome. Biology deals with this discrepancy by proofreading the action before it is coded into the DNA.
2. There are 20 AA and 61 codons that specify amino acids (NHGRI), leading to different ways to code for an average human protein. Some reasons that all of these different codes don’t work for the protein of interest could be due to the physical structure of the protein, and the efficiency of each pathway. (not totally sure of this answer)

Dr.Leproust

1. The most common method for oligo synthesis is currently a chip based gene synthesis that couples the nucleotide with phosphonamidite. Historically, this has been a solid phase synthesis.
2. It’s difficult to make oligos longer than 200nt vir direct synthesis because of the surface and primarily the inefficiency of the current state of the art phosphonamidite method. Source: Yin Y, Arneson R, Yuan Y, Fang S. Long oligos: direct chemical synthesis of genes with up to 1728 nucleotides. Chem Sci. 2024 Dec 18;16(4):1966-1973. doi: 10.1039/d4sc06958g. PMID: 39759933; PMCID: PMC11694485.
3. You can’t make a 2000bp gene with direct olio synthesis because of the increased error rate in PCR. The new gene pool solution offers a 1 in 3,000 bp error rate which would allow for greater gene lengths to be processed. (slide 39)

Professor George Church

1. According to the NIH, the nine essential amino acids are histidine, isoleucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine. This means that the Lysine Contingency is really already a contingency that all animals have upon receiving all nine of these AAs from their diet, not just lysine. (Fictional) dinosaurs are just like you and me, minus 8 more essential amino acids. Source: Lopez MJ, Mohiuddin SS. Biochemistry, Essential Amino Acids. Updated 2024 Apr 30. In: StatPearls Internet. Treasure Island (FL): StatPearls Publishing; 2025 Jan-. Available from: https://www.ncbi.nlm.nih.gov/books/NBK557845/

Week 2: DNA Read, Write, and Edit

Benchling Gel Art

DNA Design Challenge

When looking into proteins to explore further, I chose to focus on proteins related to the structure of diatom silica walls. These proteins would be exciting to understand to get a better picture of how diatoms would form the lacy micropatterns for the rubble-diatom material proposal for the final project. While there are a few different proteins key to the biosilica formation, and silaffins drive the lacy patterns and micropore structure.

In the NCBI database chose: silaffin protein (Thalassiosira pseudonana CCMP1335)

Silaffin from Uniprot

MKVTTSIITLLFASCGAADVQRVLEDVTEPAVTTPAATPAPITPEPATPAPTICEGRNFY RDDDTGKCSNEATGGIYGTLIECCVAISGSDSCPYVDICNTLQPSPSPETNEPSAKPITA APISSAPVSAAPVTSAPVAAPVETTSMTGPTTIVASIVSTNAPSSTNAPSSSLEAVVTRI PVETTNTASPTTTAASIVSTNAPSSSPEAVVTPRPTFRPSPKGTESNTFPASIASDVMFD PARSEPTFTPTSSSQPSSSEPTLSPSVSKEPTRYPTSSPSHSPTKSPSKSPSSSPTTSPS ASPTETPTETPTESPTELPTLSPTEFPSLSPTLSPTWSPTGYPTLAPSPSPSISSAPSVS SAPSSSPSISSAPSVSSAPSKNFGFLPGRNEMPTISPTEDHYFFGKSHKSHKSKATKTLK VSKSGKSSKSSKSSGSRPLFGVSQLSEGIAAGYAKSSGRSSQQAVGSWMPVAAACILGAL SFFLN

Reverse Translate Protein to DNA sequence

atgaaagtgaccaccagcattattaccctgctgtttgcgagctgcggcgcggcggatgtgcagcgcgtgctggaagatgtgaccgaaccggcggtgaccaccccggcggcgaccccggcgccgattaccccggaaccggcgaccccggcgccgaccatttgcgaaggccgcaacttttatcgcgatgatgataccggcaaatgcagcaacgaagcgaccggcggcatttatggcaccctgattgaatgctgcgtggcgattagcggcagcgatagctgcccgtatgtggatatttgcaacaccctgcagccgagcccgagcccggaaaccaacgaaccgagcgcgaaaccgattaccgcggcgccgattagcagcgcgccggtgagcgcggcgccggtgaccagcgcgccggtggcggcgccggtggaaaccaccagcatgaccggcccgaccaccattgtggcgagcattgtgagcaccaacgcgccgagcagcaccaacgcgccgagcagcagcctggaagcggtggtgacccgcattccggtggaaaccaccaacaccgcgagcccgaccaccaccgcggcgagcattgtgagcaccaacgcgccgagcagcagcccggaagcggtggtgaccccgcgcccgacctttcgcccgagcccgaaaggcaccgaaagcaacacctttccggcgagcattgcgagcgatgtgatgtttgatccggcgcgcagcgaaccgacctttaccccgaccagcagcagccagccgagcagcagcgaaccgaccctgagcccgagcgtgagcaaagaaccgacccgctatccgaccagcagcccgagccatagcccgaccaaaagcccgagcaaaagcccgagcagcagcccgaccaccagcccgagcgcgagcccgaccgaaaccccgaccgaaaccccgaccgaaagcccgaccgaactgccgaccctgagcccgaccgaatttccgagcctgagcccgaccctgagcccgacctggagcccgaccggctatccgaccctggcgccgagcccgagcccgagcattagcagcgcgccgagcgtgagcagcgcgccgagcagcagcccgagcattagcagcgcgccgagcgtgagcagcgcgccgagcaaaaactttggctttctgccgggccgcaacgaaatgccgaccattagcccgaccgaagatcattatttttttggcaaaagccataaaagccataaaagcaaagcgaccaaaaccctgaaagtgagcaaaagcggcaaaagcagcaaaagcagcaaaagcagcggcagccgcccgctgtttggcgtgagccagctgagcgaaggcattgcggcgggctatgcgaaaagcagcggccgcagcagccagcaggcggtgggcagctggatgccggtggcggcggcgtgcattctgggcgcgctgagcttttttctgaac

reverse translation into a base sequence of most likely codons from https://www.bioinformatics.org/sms2/rev_trans.html

Week 3: Lab Automation

Opentrons Art

When creating this GUI art I created an image of cherries with a checkered background to see how common features in traditional drawing would translate to bacterial image creation including shading, regular patterning, thinner and thicker lines, as well as curved and straight forms. I downloaded the python script for the PCR plate system from this simulator.

created through Ronan’s Opentrons plate art simulator

Python Script for Opentrons

colab work: importing python script and error management

In the colab script I debugged until it was suggesting a bug that could fundamentally change the function for the code to properly converse with the opentrons system. This was about the “set_offset” condition. This error code is not present in the script cell, indicating that the issue is associated with the visualization cell.

Opentrons Plate Art

this lab required lots of troubleshooting, with this plate created after the lab time by the BUGSS TAs Amanda and Joel. Previous plates were unsuccessful due to various trouble shooting issues from the python script, the point size for the asperated droplet, and a potential air bubble in the well.

Opentrons Research and Potential Use of Automated Tools in Final Project

For the final project, if I were to pursue the diatom project idea, would be to use automated systems for cell seeding by scaning contruction rubble and detect the best spots to introduce diatoms into the rubble panel in response to the rubble placement, load bearing support, or areas that make easy bridges between rubble.

Salido et.al used automated technology for diatom identification:

Salido, J.; Sánchez, C.; Ruiz-Santaquiteria, J.; Cristóbal, G.; Blanco, S.; Bueno, G. A Low-Cost Automated Digital Microscopy Platform for Automatic Identification of Diatoms. Appl. Sci. 2020, 10, 6033. https://doi.org/10.3390/app10176033

Diatoms are traditionally identified and viewed using LM and SEM imaging, since they are invisible to the naked eye. Different species of diatoms have also remained unidentified due to these limitations. Automated technology and deep neural networks have assisted in the detection, classification, and counting which can be tricky due to the variety of shapes that diatoms may have.

Crit 1 Final Project Ideas

For the final project I would like to look closer at architectural materials and how they can respond to environmental stressors. Some of these ideas are a new perspective on previous ideas of mine, playing with how one concept can be explored through multiple fields.

Project 1: Diatoms and Rubble

This proposal looks into using diatoms to create a new aggregate material out of construction waste (ie cement) and diatoms to create a bioreceptive panel system. It plays with the way that diatoms create silica cell walls, leaving opalescent lacy micropatterns that could bring a new use to materials that are very hard to repurpose. By using diatoms with rubble, this ‘new’ material would create beautiful translucent seams that could potentially help host other organisms or itself help with nutrient filtration depending on the design application.

Project 2: Biopigment Stained Glass with Air Quality Indication

Stained glass has a history of storytelling for the masses, usually in churches and other religious spaces. It would be interesting to see if biopigments could:

1. stain glass without chemical inclusions
2. react to pollution levels and serve as a visual indicator for air quality in the area. This research would need to be further refined with the intensity of the color, how sensitive the reaction could be, and how the biofilm pigment could impact its environment in terms of microenvironments, both positive and negative.

Project 3: Brick Extremophiles

Looking back at a previous project where I studied brick walls and reimagined their structure, it would be interesting to see if there are certain prevalent extremophiles present in bricks especially in iron-rich clay areas. These extremophiles could be further designed to embody carbon and support environmental remediation.

Project 4: Bruising Homes

What if buildings bruised?

This is an idea that I pursued in a filmmaking studio in the past that I would like to unpack through alternative means. A touch-reactive biopigment could become a paint or biofilm that responds to the amount of pressure put onto the surface. Relating back to organisms like touch-me-not mimosa plants that are very sensitive to their environment, a paint like this could become an exploration into the permanence of human action, and document our relationship to our homes. The paint/biofilm would begin as a uniform color, and darken in reaction to pressure.

Week 4: Protein Design Part 1

Conceptual Questions

1. Why do humans eat beef but do not become a cow, eat fish but do not become fish?

   As we eat other materials, the proteins and molecules are broken down through our digestive system, leaving us with building blocks to support our cell replication processes.

2. Why are there only 20 natural amino acids?

   These 20 amino acids were created as life was forming, and after the ‘frozen accident’, the proteins that developed at this time seem to have standardized these 20 amino acids. There were more and are more amino acids, however this connection between protein and AA during early evolution created this set of 20.

3. Can you make other non-natural amino acids? Design some new amino acids.

   Amino acids are primarily composed of hydrogen, oxygen, carbon, and nitrogen and as proven from the beginnings of RNA, there are many non-standard amino acids that could be made in fact many non-natural AAs already exist. In terms of function, I would design an AA that could create florescent tags for injury sites, helping identify issues that are less visible like endometreosis or even to find a bug bite from tics. It could help speed up wound identification and reduce time lost in exploritory tests.

4. Where did amino acids come from before enzymes that make them, and before life started?

   Many natural events have created the primodial soup that led to life as we know it, and the ingredients of AAs seem to have formed in a similar way.

5. If you make an α-helix using D-amino acids, what handedness (right or left) would you expect?

   I would expect righthandedness because ‘D’ typicially refers to right handed molecules and ‘L’ refers to left.

6. Can you discover additional helices in proteins?

Probably! Given that there is a high percentage of proteins that are still to be discovered and studies, there are probably proteins with additional helices.

11. Why are most molecular helices right-handed?

    DNA is right-handed, but also many aspects of the natural world have a handedness, like the right hand test in physics as well. This seems to be connected back to thermodynamics and create helices that are more stable.

12. Why do β-sheets tend to aggregate? What is the driving force for β-sheet aggregation?

Their structure is flat, making it easy to pack, and the edges are designed with hydrogen backbones that help form hydrogen bonds with other beta sheets.

11. Why do many amyloid diseases form β-sheets? Can you use amyloid β-sheets as materials?

    Many amyloid diseases happen when proteins misfold, and they form beta sheets because of their thermodynamic stability and their straight hydrogen bond structure allows for tighter folds. They can probably be used to support material tensile strength and rigidity, but I don’t think they classify as materials themselves.

Protein Analysis and Visualization

Fro this analysis, I am focusing on silaffin once again, following through from week 2. Silaffin is the protein responsible for the lacy biosilica patterning that diatom frustules form. I am interested in modifying this protein because tuning that pattern could allow for more targeted and intentional interactions between cement rubble and diatom placement for the final project.

silaffin sequence (from marine diatom: Thalassiosira pseudonana)

MKVTTSIITLLFASCGAADVQRVLEDVTEPAVTTPAATPAPITPEPATPAPTICEGRNFYRDDDTGKCSNEATGGIYGTLIECCVAISGSDSCPYVDICNTLQPSPSPETNEPSAKPITAAPISSAPVSAAPVTSAPVAAPVETTSMTGPTTIVASIVSTNAPSSTNAPSSSLEAVVTRIPVETTNTASPTTTAASIVSTNAPSSSPEAVVTPRPTFRPSPKGTESNTFPASIASDVMFDPARSEPTFTPTSSSQPSSSEPTLSPSVSKEPTRYPTSSPSHSPTKSPSKSPSSSPTTSPSASPTETPTETPTESPTELPTLSPTEFPSLSPTLSPTWSPTGYPTLAPSPSPSISSAPSVSSAPSSSPSISSAPSVSSAPSKNFGFLPGRNEMPTISPTEDHYFFGKSHKSHKSKATKTLKVSKSGKSSKSSKSSGSRPLFGVSQLSEGIAAGYAKSSGRSSQQAVGSWMPVAAACILGALSFFLN

silaffin has 485 amino acids, with S appearing most frequently occuring 99 times. It doesn’t belong to any protein families, which could be because this protein has a very low confidence score and is more rare, leading to less information on its structure and protein family.

screenshot from uniprot

screenshot from alphafold

With a less documented protein, there is also more likelihood for error. While some areas of the protein with better confidence have a defined beta sheets and alpha helices, most of the protein is modeled with very low confidence.

ML Design Tools

the mutation scan shows that amino acids that are most likely to mutate (more yellow in this diagram), are the L,R, and S amino acids. Position 27 is most likely to have a mutation as well.

latent space analysis shows concentrated areas. Some of the matches include proteins from organisms ranging from fungi to humans with seemingly low similarity in function. This could be due to the low confidence in the silaffin protein, so the comparative space could be drawing upon very different organisms since there is very little information to respond to.

Week 5: Protein Design Part 2

hello

Week 6: Genetic Circuits

DNA Assembly

1. What are some components in the Phusion High-Fidelity PCR Master Mix and what is their purpose?

   This Master Mix is used to increase the speed and yield of the PCR product, with accurate DNA sequences maintained for DNA replication for testing. The HF buffer variant contains DNA polymerase, deoxynucleotides, buffer, and magnesium dichloride.

   source: https://www.neb.com/en-us/products/m0531-phusion-high-fidelity-pcr-master-mix-with-hf-buffer?srsltid=AfmBOoq3xHVgYXqvCn6vETXyCMTKGo4MNvB1Zs6M25yaSdNrW9j4dHWK

2. What are some factors that determine primer annealing temperature during PCR?

   Annealing temperatures are determined by the melting temperature of the selected primers. Some factors that determine the melting point include the number of nucleotides present in the DNA oligo (the short single strand piece of DNA for this experiment).

   source: https://www.thermofisher.com/us/en/home/life-science/cloning/cloning-learning-center/invitrogen-school-of-molecular-biology/pcr-education/pcr-reagents-enzymes/pcr-cycling-considerations.html

3. There are two methods from this class that create linear fragments of DNA: PCR, and restriction enzyme digests. Compare and contrast these two methods, both in terms of protocol as well as when one may be preferable to use over the other.

4. How can you ensure that the DNA sequences that you have digested and PCR-ed will be appropriate for Gibson cloning?

   You can check that the product has the right DNA concentration for the Gibson assembly using the nanodrop after purification.

5. How does the plasmid DNA enter the E.coli cells during transformation?

   The E.coli cells are thermally shocked. First they are kept cold on ice, then heat shocked for a few minutes, before

6. Describe another assembly method in detail (such as Golden Gate Assembly)

   1. Explain the other method in 5 - 7 sentences plus diagrams (either handmade or online).

      Golden Gate Assembly works by

   2. Model this assembly method with Benchling or Asimov Kernel!

Week 7: Neomorphic Circuits

Intracellular Artificial Neural Networks

1. What advantages do IANNs have over traditional genetic circuits, whose input/output behaviors are Boolean functions?

   IANNs allow for less prescriptive circuit method, whereas traditional genetic circuits need to be specified at each step, making it a target specific circuit by finding signalling patterns across a vast dataset. Traditional genetic circuits are limited to simpler action/reaction function simulations to create an mRNA therapy.

2. Describe a useful application for an IANN; include a detailed description of input/output behavior, as well as any limitations an IANN might face to achieve your goal.

   IANNs can be very useful for biomaterials that depend on growth based on the specific conditions of each material batch. For example, if you wanted to make a stained glass using biopigments that stains relative to the colors around it, it could be useful to create a series of relative inputs (ex: high presence of red, medium presence of blue, and no presence of black) that could generate a very specific output (purple gene expression) without manually or preemtively determining color expression. The limitation of the IANN approach is that it has a ‘blackbox’ in reasoning, and doesn’t always present a clear reasoning, unlike traditional genetic circuits. When the reasoning is hidden, it could be making decisions from the wrong signals or not following a genetically safe order of operations, potentially leading to unforseen outcomes.

3. Below is a diagram depicting an intracellular single-layer perceptron where the X1 input is DNA encoding for the Csy4 endoribonuclease and the X2 input is DNA encoding for a fluorescent protein output whose mRNA is regulated by Csy4. Tx: transcription; Tl: translation. Draw a diagram for an intracellular multilayer perceptron where layer 1 outputs an endoribonuclease that regulates a fluorescent protein output in layer 2.

Fungal Materials

1. What are some examples of existing fungal materials and what are they used for? What are their advantages and disadvantages over traditional counterparts?

   There are headphones designed by Aivan that use mycelium in an experimental synbio headphone design for the leather ear cushion covers. This product is a beautiful example of the multitude of characteristics that bio-based materials like mycelium, silk proteins, and PLA could embody instead of using plastics to make these parts of the headphones. The issue with this product at this time goes partly into user trust and long-term durability of each component. User’s may not feel comfortable having fungus pressed against their ears, and the materials are more susceptible to degredation when exposed to water, heat, sun, etc than a traditional plastic object would be. Headphones are also often tossed into a backpack, worn over earrings, or worn during exercise, leading to a whole range of user habits that could degrade each layer over time. Traditional plastics are also more easy to control and form at this time, when even simple glues may not act as expected with fungal materials over time.

   

   (source: https://www.dezeen.com/2019/05/24/korvaa-headphones-bioplastic-fungus-yeast-materials-aivan/)

2. What might you want to genetically engineer fungi to do and why? What are the advantages of doing synthetic biology in fungi as opposed to bacteria?

   Fungi feed on simple sugars very well, but this also means that it is easy for mycelium products (before they are fired) to get contaminated during the growth cycle of the fungal products. It would be very useful to have a mycelium product that grows using a specalized sugar source over simple glucose to mitigate the issues of contamination. However this may also create a dependency on a narrow stream of sugar sources that could be used reliably for mycelium fabrication, and potentially limit the accessibility of lo-fi mycelium explorations. Instead it would be cool to have a strain of mycelium with an anti-mold protein, preventing mold from growing in mycelium products whereever mycelium spores are present. Each option would need to be tested for any side effects such changes could have on the mycelium in use. One key advantage of fungi synthetic biology is the speed of growth and visibility of the fungal spores. They are easy to notice by the human eye (larger scale), and still have a high replication rate.

Week 9: Cell-Free Systems

General Homework Questions

1. 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.

   Because cell-free methods exist without the limitations of maintaining a living cell, they offer greater flexibility in exploring the details of protein synthesis without

2. Describe the main components of a cell-free expression system and explain the role of each component.

3. Why is energy provision regeneration critical in cell-free systems? Describe a method you could use to ensure continuous ATP supply in your cell-free experiment.

4. Compare prokaryotic versus eukaryotic cell-free expression systems. Choose a protein to produce in each system and explain why.

5. How would you design a cell-free experiment to optimize the expression of a membrane protein? Discuss the challenges and how you would address them in your setup.

6. Imagine you observe a low yield of your target protein in a cell-free system. Describe three possible reasons for this and suggest a troubleshooting strategy for each.

Homework Questions from Kate Adamala

Design an example of a useful synthetic minimal cell as follows:

1. Pick a function and describe it.

   a. What would your synthetic cell do? What is the input and what is the output?

   b. Could this function be realized by cell-free Tx/Tl alone, without encapsulation?

   c. Could this function be realized by genetically modified natural cell?

   d. Describe the desired outcome of your synthetic cell operation.

2. Design all components that would need to be part of your synthetic cell.

   a. What would be the membrane made of?

   b. What would you encapsulate inside? Enzymes, small molecules.

   c. Which organism your Tx/Tl system will come from? Is bacterial OK, or do you need a mammalian system for some reason? (hint: for example, if you want to use small molecule modulated promotors, like Tet-ON, you need mammalian)

   d. How will your synthetic cell communicate with the environment? (hint: are substrates permeable? or do you need to express the membrane channel?)

3. Experimental details

   a. List all lipids and genes. (bonus: find the specific genes; for example, instead of just saying “small molecule membrane channel” pick the actual gene.)

   b. How will you measure the function of your system?

Homework Questions from Peter Nguyen

Freeze-dried cell-free systems can be incorporated into all kinds of materials as biological sensors or as inducible enzymes to modify the material itself or the surrounding environment. Choose one application field — Architecture, Textiles/Fashion, or Robotics — and propose an application using cell-free systems that are functionally integrated into the material. Answer each of these key questions for your proposal pitch:

1. Write a one-sentence summary pitch sentence describing your concept.

   A ‘bruising’ paint that reacts to pressure, creating a lasting impression on a house as it is used more routinely.

2. How will the idea work, in more detail? Write 3-4 sentences or more.

   It would work by creating a biofilm ‘paint’ with pressure-reactive sequences from organisms like mimosa plants. Alternatively the biofilm could use thermoreactive organisms, marked with florescent protein to signal when the temperature is around the range of human skin.

3. What societal challenge or market need will this address?

   This starts a conversation around permanence in a space, and how buildings are seen as more temporary than the people as we move away from being able to afford to own our homes and instead move through rental properties, where our spaces are more defined through the objects than a sense of connection to the walls around us.

   In a healthcare sense, this could be useful to highlight surfaces that have been touched, potentially helping with sanitation for sterile rooms if we enter another large-scale health event.

4. How do you envision addressing the limitation of cell-free reactions (e.g., activation with water, stability, one-time use)?

   Walls are not often washed outside of bathrooms and kitchens perhaps, but our hands have moisture, but probably not enough to introduce water into the cell-free system. Pressure on the wall could compress and break micro-scale water cells that are suspended in the biofilm, causing a localized water exposure for the biopigment to react with.

Homework Questions from Ally Huang

Freeze-dried cell-free reactions have great potential in space, where resources are constrained. As described in my talk, the Genes in Space competition challenges students to consider how biotechnology, including cell-free reactions, can be used to solve biological problems encountered in space. While the competition is limited to only high school students, your assignment will be to develop your own mock Genes in Space proposal to practice thinking about biotech applications in space!

For this particular assignment, your proposal is required to incorporate the BioBits® cell-free protein expression system, but you may also use the other tools in the Genes in Space toolkit (the miniPCR® thermal cycler and the P51 Molecular Fluorescence Viewer)

1. Provide background information that describes the space biology question or challenge you propose to address. Explain why this topic is significant for humanity, relevant for space exploration, and scientifically interesting. (Maximum 100 words)

   There could be a cell-free system that reacts to

2. Name the molecular or genetic target that you propose to study. Examples of molecular targets include individual genes and proteins, DNA and RNA sequences, or broader -omics approaches. (Maximum 30 words)

3. Describe how your molecular or genetic target relates to the space biology question or challenge your proposal addresses. (Maximum 100 words) Clearly state your hypothesis or research goal and explain the reasoning behind it. (Maximum 150 words)

4. Outline your experimental plan - identify the sample(s) you will test in your experiment, including any necessary controls, the type of data or measurements that will be collected, etc. (Maximum 100 words)

Week 10: Advanced Imaging and Measurement Technology

Based on the older information:

• m/z(n) = 824.1148

• m/z(n+1) = 800.608

• n = 34.0172

MW = (n * m/z(n)) - n

MW = ((34.0172) (824.1148)) - 34.0172

MW = 28,000.06 daltons

Error Rate/Accuracy = ((28006.6 - 28000.06)/28006.6)1000000 = 0.0002341000000 = 233.52ppm

This value is much higher than the 50ppm threshold, indicating an issue with the tested product. Also, I think this calculation seems to actually be for inaccuracy or error rate.

The sequence has 20 K’s and 6 R’s for peptide segmentation sites

MVSKGEELFTG VVPILVELDG DVNGHKFSVS GEGEGDATYG KLTLKFICTT GKLPVPWPTL VTTLTYGVQC FSRYPDHMKQ HDFFKSAMPE GYVQERTIFF KDDGNYKTRA EVKFEGDTLV NRIELKGIDF KEDGNILGHK LEYNYNSHNV YIMADKQKNG IKVNFKIRHN IEDGSVQLAD HYQQNTPIGD GPVLLPDNHY LSTQSALSKD PNEKRDHMVL LEFVTAAGIT LGMDELYKLE HHHHHH

There are 21 labeled peaks, and one unlabeled peak in the chart between 0.5 seconds and 6 seconds.

Compared to the 19 rows for the peptide prediction, the 21 labeled peaks in the chart indicate that there is a slight discrepecy in the threshold between the two software. One observation is that it could down to the two double peaks within two of the larger peaks on the chart, which accounts for the difference of 2 between the rows and peaks. The double peaks could be referring to the same peptide, while each potential peptide in the table is unique.

• m/z(n) = peak 1 = 525.767

• m/z(n-1) = peak 2 = 526.259

delta between peaks = 0.492

z = 1/delta = 2+ charge