From 2007.igem.org

Projects

Design: Wet Lab

Design: Printer

Design: Software

Testing

Construction: The Wetlab

Protocols

Final Result of E.co Lisa

The project we selected was to design and build a biomechanical printer; composed of a two dimensional plotter equipped with a red laser, software to translate computer images into instructions for the plotter, and E. coli cells engineered to respond to the laser light. Bacteria are spread in a solid lawn on the plate, or mixed in the media before pouring the plate. The response triggered by this biological circuit will produce beta agarase, an enzyme which degrades the agar polymer that the cells rest on.

The printer can then be used to "draw" high resolution images on the bacteria with the laser. The bacteria will then dissolve the agar where the laser was shone. This results in Bacterial Lithography, where the dissolved agar forms a picture. Cool eh.

We also chose a second project to include in our entry to the competition this year. That is an in Silico Biobrick Evolution system. The purpose of this project is to design a system that will accept user entered parametres and use them to search through the registry database. Using the given parametres the system will try to construct circuts (a series of biobricks) that will produce the desired product. More information on this project can be found in our evoGEM sections.

Our team investigated a number of potential projects before selecting our light sensing printer. This section offers a brief outline of some of the ideas we considered.

• Antibacterial Plasmid - A novel strategy for stopping difficult bacterial infections: give the patient an antibiotic and an engineered plasmid with resistance to that antibiotic. (Alternatively, give the bacteria any plasmid that would grant them a strong survival advantage in the patient.) Within a short time, most the infectious bacteria should be carrying the resistance plasmid... at which time a chemical signal is administered to the patient that initiates ccdB (control of cell death protein B) production on the plasmids, wiping out the infection.
• Hydrogen Disulfide Sensor System - A useful tool for the oilpatch might be a biological H2S sensor, in the form of bacteria that turn a different colour / glow in the presence of H2S gas. H2S is highly toxic; and the human nose cannot detect H2S at very low or very high (including potentially lethal) concentrations. This makes detectors invaluable for safety in H2S laboratories and in the oilpatch, where H2S is present in oil and especially in natural gas deposits. H2S sensors are not inexpensive however, a cheap biological sensor might be useful. Ironically, concerns about the safety of working with H2S in the lab precluded this idea =P.
• Addressable Bacterial Memory - This was a mostly unformed idea. The thought behind it was use previously developed bacterial memory to store something interesting or relevant to the bacteria.
• Phototaxic Bacteria - The idea behind this project was to couple a light sensing system to bacterial chemotaxis to create bacteria that woud swim towards the light
• Directed Evolution of the Light Sensor - The light sensor currently absorbs in a range around the 660 nm range: red light. Directed evolution could be used to modify this spectrum, possibly changing it to an entirely different light colour. Selective pressure would be applied by linking light sensor activation to antibiotic resistance gene expression, and gradually changing the emitted light frequency. As the frequency increases (or decreases), mutants with sensors sensitive to that frequency will produce more of the resistance gene product and tend to outcompete the rest in an antibiotic environment. Gradually changing the emitted frequency could allow us to 'pull' the actual absorbed frequency along.
• E.coli Reverse Chemotaxis - Another unformed idea. We were looking at setting up a system where E. coli would run away from nutrients
• Cross Species / Gram Negative to Gram Positive Signalling - One team last year (Rice U) attempted to do cross species signalling from a gram+ species of bacteria to a gram-, using a quorum sensed molecule. We beleive this experiment failed because gram+ bacteria have one membrane and gram- bacteria have two, and the particular gram+ signal was unable to penetrate to the engineered receiving system in the gram- strain. However, communication going the other way (gram- to gram+) shouldn't experience this complication.
• Use the Cell Specificity of Pathogenic Bacteria to Deliver Drugs - Certain pathogenic bacteria have evolved the ability to infect specific cell types in humans. If the toxin production in these bacteria was replaced with pharmaceutical production, the result would be a highly targeted drug delivery system. Probably some ethical issues with this one though...
• Isolate Bacterial Vesicles with Synthesized Drug Inside - The idea is to target a synthesized chemical / pharmaceutical to vesicles, small membrane sacs, within the cell. Then, the cells are lysed in a way that destroys the cell membrane while leaving the vesicles intact, allowing their easy recovery by centrifuge.
• Attenuated Cells: Iron Supply Limit - All cells require iron to live, and developping diverse iron acquisition systems is a particular passtime of bacteria. However, iron is used mainly in cofactors and enzymes: it is not consumed by the cell and is not excreted as a waste product. An engineered cell endowed with a large supply of iron but stripped of it's uptake mechanisms should be able to divide a finite number of times (each division halving the amount of cellular iron) before iron levels drop too low to sustain life. This might be good a mechanism for controlling genetically engineered species released into the wild, requiring them to die off after a set number of generations, thus limiting or negating their environmental impact. Ethical issues abound here too!