Week 14: Moving forward with research

As part of our last class to develop ideas, we worked amongst our groups to prepare for the final presentation at BUGS taking place the following Friday.

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We scanned an oyster shell for the convenience of being able to model over it.

We also talked about modeling our cage design after the following structure.  When originally talking about our designs we were considering the structures be made of Calcium Carbonate.

In addition to this, while at the ‘Holy Spat’ talk, we realized that the current cage design is simply an inconvenience and what if that entirely was redesigned to accommodate the culture and cultivation of oyster forming and oyster reef rebuilding.  A similar structure to the jack toys was considered as our design model. As a structure that reaches from the inside to the outside, it allows wheel build up to maximize its volume and grow off of each other.

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A Week 12: Perspective on Culture as Medium exhibition

 

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Recently, I had the pleasure of attending Culture as Medium, curated by Margaret MacDonald(pictured above), during the opening reception at Motor House on April 1st, 2016.
Culture as Medium is a show utilizing a double-entendre for culture, the show operates at the intersect between the study of microbes, and artistic application. I found the show to be a successful attempt at identifying, and ultimately dissolving a cultural boundary between canonically scientific content, and artistic practice. The show is being hosted at both Motor House, a show-space, and Baltimore Under Ground Science Space (BUGGS), serving to further dissolve the societal construct between art and science. Each of the works showcased at Motor House had varying levels of community engagement, and were produced by artists who utilize biology as inspiration.
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Tal Danino, a scientist-turned-artist offered an installation playing with microbes as a visual medium. Working with various bacteria, the installation consisted of 3 monitors running visuals derived from manipulated bacteria, operating in conjunction with a rotating slide projection of curated bacterial samples from varying species, as well as a grouping of more slides, giving viewers the opportunity to engage with the slides and serving to further highlight the variety in scale between the projections, the actual slides, and the monitors. It also allowed for viewers to see the actual specimens and the beautiful diversity of color in the work. This work is particularly successful at removing the constraints between what is art and what is science, the practice required to develop this imagery is canonically rooted in science, but the results are visible, and palatable; this is taste that allows for a familiarity between a casual viewer and bacteria that extends beyond the stasis of a textbook, and the sterility of microscopy.

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Ryan Hammond, unlike Tal Danino is an artist-turned scientist. Hammond’s work consisted of a wooden box, housing a setup including a monitor, a joystick and living paramecium. Offering an opportunity for viewer participation, the work functioned utilizing a natural quality of the paramecium. Paramecium naturally move in response to electric stimuli, and within the paramecium housing unit the artist had set quadrants that correspond with the access of the joystick; as viewers moved the joystick the paramecium moved in response, allowing for the viewer to control the paramecium. Of particular interest to me were the conversations that surrounded the work. I had the pleasure of witnessing a debate between a viewer who was also a physicist, and Ryan Hammond, the debate beginning with the ethical question, do we have a right to essentially play games with a life form. The debate then opened up to a larger discussion first questioning anthropocentrism, and later discussing the concept of free-will, ultimately coalesqueing into the question on how to define sentience, and the value of life itself. As a side note, this work also helped facilitate a conversation with Nick, a child at the show, who claimed that “Germs are bad, they make people sick.” Because Nick had played with these “Germs” and felt a familiarity with them he and I were able to have a discussion with him about bacteria and his ethical point of view in a way that hopefully wasn’t too boring for him. The ability of this work to be available and palatable to someone not professionally engaged with artistic or scientific spheres of influence is particularly impressive to me.

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Francois Lapointe was also featured in the Motor House exhibit. His work operating as both performance, and scientific experimentation, Lapointe presented viewers at the opening with two works. The first work visible in the show being several boards with documentation of an ongoing performance he has been doing in which he measures the changes in his micro-biome after engaging in 1000 Handshakes. He has performed this several times, including a performance walking around Baltimore, which he performed April 2nd, the day after the opening at Motor House. The other work Lapointe presented was a live performance/experiment. Donning a lab-coat, and with the help of his lovely and competent assistant, Lapointe performed a Kimchi Eating Performance, in which he ate only Kimchi for an extended period of time, periodically swabbing samples of his tongue in order to track the effect the kimchi has on his micro-biome. The performance was particularly entertaining considering his visible distaste for kimchi, allowing him to fully activate the idea of sacrifice in the name of art and science. While Lapointe has a performative practice rooted in scientific ritual, there is something to enjoy about how approachable he was, viewers were encouraged to speak with him during the act, offering a friendly face, and casual banter. This was my favorite part of his work considering it allowed me to connect the concept of the micro-biome and the potential effects of environment in real time on the micro-biome, as well as offering an experiment packaged as a person with which I can sympathize, ultimately fostering a period of self-reflection upon my own micro-biome and habits.

All of the works in the Motor House opening of Culture as Medium were, in my humble opinion, very compelling, and well curated given the theme of the show. Unfortunately I have yet to see Anna Dimitriu’s work at BUGGS, however if it is anything like the work on her website (http://annadumitriu.tumblr.com), then I expect to be similarly wowed. While I have focused mainly on my motor house experience, there is much more history and influence that was requisite for the show to be as successful as it was. If you are so inclined feel free to watch Margaret Macdonald’s talk (https://www.youtube.com/watch?v=Rk6NpBmpK90)  on how she thinks and her process of producing an exhibition as an experiment.

 

Week 11: Group Projects and Culture as Medium

 

In the morning of  April 5th,  our class had a follow up discussion on Margaret MacDonald’s exhibition Culture as Medium. At the opening in Motor House, many of the students from Biofabrication participated in assisting performance and initiating provocative conversation

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The exhibition Culture as Medium blurs the boundaries between science and art, artists and scientists. As François-Joseph Lapointe states, “For me, experimental science and experimental art are two branches of the same tree, identical twins separated at birth.” Tal Danino’s series of Microscopic Bacterial Videos (on view at Motor House) imparts the haunting beauty of the synchronized dynamic behavior of bacteria while revealing cutting edge research in synthetic biology. To produce Sequence Dress (on view at BUGSS), Anna Dumitriu immersed herself in whole genome sequencing research of clinically important bacteria in order to, as Dumitriu writes, “artistically explore the emerging technology of whole genome sequencing of bacteria and consider what it means to us personally, culturally and socially.” With his performance experiment, 1000 Handshakes (to be performed in Baltimore on April 2, 2016), Lapointe creates what he describes as “metagenomic self-portraits” or “microbiome selfies” that illustrate the metamorphosis of his bacterial self as he shakes hands with those around him.

http://www.cultureasmedium.com/about/

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In the afternoon, the class broke into three research groups(Body as site of manufacturing, Machine as site of manufacturing and Outdoors as site of manufacturing) to further discuss the idea and research as our research papers and presentations are due the next class meeting.

Week 10: Group Projects and Visitor Thomas Burkett

In class, on March 29th, we began by breaking up into our groups to continue discussing the ideas and research collected over the past few weeks.  Ryan Hoover greeted each group to review our ideas for project references and material possibilities.

 

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Research Ideas:

Group 1: [The Body as Site] Working with skin.

-largest organ on the body, filtration

-personal micro bioms

-cosmetics

Group 2: [The Machine as Site] Working with oysters

-harvesting oysters

-promoting oyster growth in the Bay due to excellent water filtration abilities

-possible material resource for glue production

Group 3: [The Outdoors as Site] Material Exploration for the production of PPE equipment and manufacturing process

-PPE respirator, bio-material for filtration

-Bio-plastic, Silk fibrin, and mycelium material

-Modular manufacturing system

 

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During the second half of the class, we had the pleasure of meeting and speaking with Thomas Burkett, one of the founders of BUGSS. BUGSS, Baltimore Underground Science Space, was created for anyone interested in Biotechnology, to learn, collaborate and hang out with people equally excited by the potential of this field.  This was an excellent opportunity to ask questions about how he opened up the non-profit space as well as share our project ideas for valuable feedback.

http://www.bugssonline.org/

Week 8: Presentations // Establishing Group Projects

 

Sarah Whelton's Presentation
Sarah Whelton’s Presentation

In class on 3/8, we each gave presentations answering the following 5 questions, in order to highlight common themes and interests, and help us establish research groups.

Prompt:

  1. What is the most interesting thing you’ve learned in class so far?

2. What is a project from another artist/designer/researcher that uses biology in a way  you find inspiring?

3. What is an approach to manufacturing (loosely interpreted) that you find compelling?

4. Describe an intersection of biology and another discipline that you find exciting.

5. What do you want to continue to explore?

 

After the presentations, we discussed some of the recurring themes from our presentations, including the role of synthetic biology in art and the role of art in synthetic bio, open source (in general), Open Source Gendercodes, sustainability, etc…

We then divided into three groups under the general titles:

  1. Body as site of manufacturing (formerly the “visual” team) [C.B., A.B., S.K., B.K.]
  2. Machine as site of manufacturing [A.F., M.R., E.K., S.C.]
  3. Outdoors as site of manufacturing [J.O., G.B., E.W., S.W.]

 

“What’s Up Daniel Grushkin!”

We were lucky enough to have one of the Biodesign Challenge’s top ‘team’ members in our midst for the entirety of class. He spearheaded a lot of the discussion and helped push in the right directions w/ regard to group formation. He came equipped with inspiration…here’s his bio:

 

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Daniel Grushkin is a former fellow at the Woodrow Wilson International Center for Scholars, where he researched the field of synthetic biology. He is an Emerging Leader in Biosecurity at the UPMC Center of Health Security. He cofounded Genspace, the world’s first community laboratory. Fast Company ranked Genspace among the top 10 most innovative education companies in the world. As a journalist, he reports on the intersection of biotech, culture, and business for publications including Businessweek, Fast Company, Scientific American, and Popular Science.”

 

Week 6: Biomachines

Notes from Dr. Rebecca Schulman Lecture on Bioengineering and Nanotechnology

  • Note on artificial intelligence: High level of difficulty replicating the design and function of simple organisms such as ants, more so in many cases than replicating aspects of high level human intelligence?
  • That said, can we create:
    • self-assembled structure across scales?
    • a cell?
    • self replicating materials?
    • self healing materials?
    • Shape-changing materials or materials with controlled dynamics?
  • Biology points at fundamental challenges in material design.  We should be able to change what materials are doing, have them react for directly to their environment for example.
  • Could we imagine a “matter computer”: A way to automate the building of structures on a nano-scale to be able to design at a more feasible human scale.
  • Nanostructures can be made by weaving DNA, which is comparable to weaving with an asymmetrical slinky, meaning that we have limited control of structures designed on a nanoscale.
  • Overview of DNA origami: how to construct nanostructures by folding pieces of DNA.  
    • How do we use our knowledge of DNA structuring on a nanoscale to make material do what we want?  
    • Directly scaling up nanostructures is not feasible since DNA easily gets deformed.  
    • With the amount of DNA that would be involved in just extending simple DNA origami structures, the failure rate is about 100%.  
    • Instead, how can we get molecules to do more complex jobs?  
    • Could we imagine building complex objects by replicating and evolving genetic code?
  • Overview of the self-replication of clay through crystal growth, no enzymes or other additional substances involved.
  • By switching the growth “rules” we can build libraries of complex patterns from a small number of substrates.

 

William Shih lecturing on DNA origami
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Images of DNA weavings on a nano scale

 


After the lecture by Rebecca Schulman, we broke out into groups for a brainstorming session on applications of nanotechnology.  Ideas do not have to be feasible or “good”.

Group 1 Prompt: Redesign a tool you use commonly as a nano scale device.

Ideas: A crane that would use the technique discussed by Dr. Rebecca Schulman of programmable molecules that grow fibers and move about randomly in the process of diffusion until it meets its specified partner molecule and sticks to it.  The fiber would then swing the object and move it to the desired location.

Group 2 Prompt: Design a material that could self-replicate to form a structure on a larger scale.

Ideas: Self-replicating wall pigment that would cover a wall with a single drop of pigment with a built-in kill switch that would be activated when an organic surface was detected.

Group 3 Prompt:  Redesign a non-living object as a pseudo-living machine. Redesign a living organism as a pseudo-living machine. 

Ideas: Instead of a tooth brush and tooth paste, a genetically-modified organism that would inhabit the mouth and feed off of plaque and other corrosive bacteria.
A robotic pollinator that could perform the roles of living pollinators such as bees.


 

The remainder of the day was spent reviewing readings “Applications of Designed Biological Systems” from Synthetic Biology Primer and Chapter 1 & 2 of Synthetic Aesthetics.

This opened up a discussion of ethics in relation to Bioengineering and the even bigger question of what constitutes life and intelligence.

We also discussed some of the applications of bioengineering laid out in Chapter 7 of Synthetic Biology: a Primer.

Week 5: Bioprinting

In Week 5 we started our introduction to bioprinting by manually writing gcode to draw our intitials with a pen using a Prusa 3d printer. We then worked with Ryan Hoover’s Xylinus tools for Grasshopper (a visual scripting plug-in for Rhino) where we learned to write a script the could produce gcode directly in grasshopper, which also allows much more manual control than programs like Slic3r and Cura – something that will be useful when creating gcode for bioprinting. In the afternoon we got an overview on using a pneumatically driven syringe for bioprinting which we will be using in the coming weeks.

 

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The basic gcode commands we worked with.
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We drew on an x,y grid then translated it into gcode.
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Sarah’s handwritten gcode which she then transcribed into a text editor.

 

 

 

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Gcode drawing previewed and then sent to the Prusa printer in Pronterface.
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Finished drawing from the Prusa.

 

video of the Prusa drawing the manually written gcode.

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Ryan’s script using Xylinus tools in Grasshoppper to produce gcode from geometry directly in Rhino.

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Emma testing the air pressure on the pneumatically driven syringe on Ryan’s bioprinter.
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Testing out some water in the syringe to get started.

 

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The syringe with the fine needle tip inserted into it’s mount on the printer.

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Week 4: Programming and Making with Proteins

Morning(9:00am-12:00pm)

Stenciling & Protein Visualizations in Grasshopper

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Today in class we started by cutting stencils for our petri dishes out of oven bags and aluminum foil. The stencil serves as an attempt to control the growth of the GFP that we saw expressed here ——->IMG_5827IMG_5831

After identifying these fluorescent colonies, we sampled choice colonies that had good protein expression and glowed brightest under UV exposure.  Mixing these colonies with growth media we prepared a 1:5 mixture of e.coli bacteria and growth media.

Before laying the stencils on our agar plates, we put the whole lot of them into the autoclave for sterilization.

Meanwhile, we moved to the computer lab to re-visit our digital files of the digestion/ligation process.

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Taking the GFP backbone and the LAC promoter, we can visualize through Alba plugins for Grasshopper what the digested plasmid looks like. Taking this a step further we used a component that spits out an amino acid sequence for us to read and translate into the corresponding condones for protein synthesis. The goal is to take the amino acid sequence and send it through a third-party protein library to get a visualization of the protein. Below is a rendering of the GFP protein.

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Afternoon (1:00pm-2:30pm)

Lecture from visiting scientist Marc Ostermeier

Ostermeier began his lecture discussing the way we look at proteins and how they can be engineered to function within systems. Breaking down the stages of inquiry into protein design, cell design and evolutionary design algorithms (his niche). Screen Shot 2016-02-15 at 8.37.32 PM

IMG_5853The process of protein design is best illustrated in Ostermeier’s graphic of the maltose bind protein and beta-lactamase. Beta-lactamase produces ampicillin. The maltose binding protein binds to maltose. These two proteins each have their own individual function. When the DNA of the proteins is spliced together, the resulting combination changes the function of both proteins. Dr. Ostermeier’s lab screened a high volume of spliced proteins for maltose dependent ampicillin production. Thus, when maltose is not present, no ampicillin is produced. When maltose is present, ampicillin is produced. This resulting interdependency is called a protein switch i.e. maltose dependent beta-lactamase. (above) A protein switch is a versatile part of cell design and can be applied to many different scenarios. In the lab, we use ampicillin to screen for antibiotic resistance. When our antibiotic resistant e.coli bacteria were introduced to our ampicillin treated petri dishes, only those cells that expressed the antibiotic resistance would survive, assuring only desirable colonies to grow.

Screen Shot 2016-02-15 at 8.59.52 PMThe implementation of these protein swithes can be seen through their usage of gene expression. A simple gene circuit like this example allows a specific gene to be expressed when lactose is present. This specific scenario serves as a model for gene therapies and other applications. For example, cancer cells are attracted to cells showing hypoxia factors or low levels of oxygen. By designing a protein that searches for these high levels and bonding to the corresponding cancer cells their may be a way to safely use protein switches identify areas of cancer cell concentration.

Ostermeier’s lab concentrates on a particular way of going about the biology part of this theoretical framework. He introduced the fitness landscape paradigm for understanding evolutionary design and it’s success. The success of the evolution is dependent on the variable being measured or really what you’re looking to achieve. Screen Shot 2016-02-15 at 9.29.12 PM

 

Post Lecture Brainstorming ~ 30 minutes exercise

We split up in to teams of four to five students and were given this set of rules. —->

 Rules.

  • No idea is too preposterous.
  • Technical realities can provide a foothold. When they are an obstacle, they may be handwaved.
  • In this context, neither ownership nor responsibility over ideas is a concern.
  • Bad ideas may run freely.
  • Communal participation is a communal responsibility.
  • Riffing and clashing are equally acceptable forms of exchange.

Each Team then got their prompts and 25 minutes to pass around ideas.

Team Prompts

Team One: Redesign a game as a collection of genes.

Team Two:Pick current two presidential candidates. Design a single-celled organism mimics an aspect of their personalities or policy.

Team Three: In the lab last week the fine powdered ice was a perfect solution to a technical need. Outside of the lab we saw our city come to a halt as it was covered in a massive amount of fine powdered ice. Consider the dramatic shifts that happen with volume and context. Design an organism that has one sort of function in the lab, then another when it exits the lab and operates at a greater scale.

The class met again to discuss each groups findings and how we could implement something as simple as a fluorescent protein.

Night (7:00pm-7:30pm)

After an artist talk on project OpenSourceGenderCodes a portion of students went back to the lab to finish applying stencils and the bacteria/growth media mixture. The stencil was laid down flat on the petri dish. Then the bacteria/growth media mixture was spread, brushed and spritzed onto the surface of the agar plate and over the stencil. Below is the before stencil and that same stencil 72 hours later after incubation.

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Week 3: Synthetic Biology, in the Lab (Digestion, Ligation and Things That Glow)

I. An overview of the Bio Lab 

 pipet protocol

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  • Pipette Etiquette
  • USE A LOT OF TIPS. WHEN IN DOUBT CHANGE THE TIP.
  • do not leave the lid of pipet tip holder open after grabbing new tip.
  • units we used in the lab: mainly micro-liters
  • Microliter – µ1 -.000001
  • Liter – L – 1

 

 

 

 

 

 hot plates 

  • rapid heating, heat shocking – (thermometer must be in water)

 incubator

  • an enclosed apparatus providing a controlled environment for the growing of microorganisms under controlled conditions

 autoclave 

  • performs sterilization through steaming/heat

*IMPORTANT: keep thing sterile in the lab, gloves – don’t touch face, mouth etc./pipet etiquette/do not drop things on the floor 

centrifuge

  • machine with a rapidly rotating container that applies centrifugal force to its contents, to combine or settle 

 


II. Overview of our first project in the lab

In this lab, we take DNA sequences for GFP (Green Fluorescent Protein) from fluorescent jellyfish, and catalyst its production; then combine with E Coli. cells – in theory, producing colonies that would soon fluoresce under black light.

 

Alba the rabbit 

 

 


 

III. Protocol

 

1. Digestion

Refer to the Digestion+Ligation+Transformation Protocol worksheet 

*snowflake symbol means keep on ice

A. GFP PLASMID

  1. Begin with  DNA – the GFP (combined with the RBS and Terminator) on a plasmid with chloramphenicol resistance 
  2. Add CutSmart buffer to help the reaction
  3. Add Enzymes  (PstI & XbaI)

The restriction enzymes cut the DNA at the site of a specific sequence.
Repeat these steps in separate tube for each color of fluorescent protein (GFP, YFP, RFP)

B. LACI PLASMID

  1. Begin with DNA –  LacI Promoter on a plasmid with Ampicillin resistance
  2. Add CutSmart buffer to help the reaction
  3. Add + Enzymes (PstI & Spel)

Incubate tubes at room temperature for 1 hour

Heat using hotplate for 20 min @ 65°C (kills enzymes)

2. Ligation

Ligation is done on ice.

Repeat these steps in separate tube for each color of fluorescent protein (GFP, YFP, RFP)

Combine-

  1. digested DNA ( RBS + GFP + terminator)
  2. digest DNA – Lacl promotor (+ampicilin-resist)
  3. Ligase buffer (contains ingredients essential to ligase activity)
  4. ligase (enzyme used catalyze the joining of two molecules by forming a new chemical bond)

incubate @ 16°C overnight – or- @ room temp. 10min

heat inactivate, with hotplate, @ 65°C for 10min

chill on ice in prep for transformation (or in -20° freezer for storage)

 

 

 

 

3. Transformation

*Keep on ice

Prepare tube with – 

  1.  (dh5α competent) E. coli cells
  2.  assembled GFP plasmid 

Incubate on ice for 20min.
Heat shock @ 42°C for 45sec
Chill on ice 2+min

Combine –

  1. +transformed E. Coli cells
  2. +SOC Media (maximizes the transformation efficiency of competent cells)

 Incubate @ 37°C for 2 hours in shaker.
 Pipette 200μL onto petri plates with LB agar + Ampicillin
 Incubate overnight (14-18hr) @ 37°C
 Check for growth and protein production (colonies appear as dots)
 Transfer plates to 20°C refrigerator.

 

4. CULTURE SINGLE COLONY in liquid media 

in a glass test tube, combine

  1. LB liquid media
  2.  transformed E.Coli colony

Incubate overnight in shaker at 37 C


IV. Additional Resources

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~emma w

Week 2: Synthetic Biology, in silico

We began class on 2/2/16 with learning about the namesake of the primary Grasshopper Tool that our class will be primarily using through the Semester, Alba.

“Alba” is the name of a genetically modified rabbit created by artist Eduardo Kac, with help from French Geneticist Louis-Marie Houdebine in 2000. By using a Green Fluorescent Protein (GFP) ordinarily found in Jellyfish, Alba is able to glow entirely green under Blue Light.

 

Afterwards, we dove right into leaning Ryan’s Alba Grasshopper Suite. We are using the scripting component of Grasshopper combined with resources from BioBricks to fully understand DNA and the ways we can interact with it in this rapidly developing field.

For example,  we can use several visualizers to show us the base pairs, and amino acids of a particular Promoter.

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In the first Alba Grasshopper definition we created, we simply visualized the Amino acids and Base pairs of a Green Fluorescent Protein (E0040) into the Rhino Viewport.

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1st Alba Definition

 

In the Second Definition we worked with,  We visualized the Green Fluorescent Protein again into the Rhino Viewport, but we learned how to highlight the subsections in the DNA. Using the visualizer, we highlighted the promoter of the Green Florescent Protein.

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2nd Alba Definition Highlighting Promoter

 

In the third Definition we created, we practiced running Digestion ad Ligation within Alba. We essentially laid out what we were going to do in the lab later after lunch. We visualized the Digestion of the GFP Plasmid with the Pstl and the Xbal Enzymes, and the digestion of the Lacl plasmid with the Pstl and the Spel enzymes. Afterwards, we visualized the ligation of these two plasmids together.

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3rd Definition, Digestion and Ligation

You can see in the Rhino viewport how we selectively cut out certain sub sections of DNA to eventually Ligate together.

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The Two inner circles are the Digested Plasmid with the specific sub sections of DNA we Digest, with the outer circle being the final completed Ligation of the two Plasmids.