Stenciling & Protein Visualizations in Grasshopper
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 ——->
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.
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.
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).
The 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.
The 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.
Post Lecture Brainstorming ~ 30 minutes exercise
We split up in to teams of four to five students and were given this set of 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 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.
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.