Team:Heidelberg LSL/Project Summary

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Summary & Outlook

Lying in the sun, we now have time to recover and think about the past weeks…

Summary

After a long phase of planning and development, we sucessfully developed a new biological system in E.coli that reacts quantitatively to UV radiation in both artificial, and natural doses of radiation.

All of our promoters, precA, precB and psulA, which are linked with the SOS-response, reacted reliably to UV-induced damages and started expression of our reporter genes GFP and LacZ. All combinations such as precA_LacZ, precA_GFP, precB_LacZ, psulA_LacZ and psulA_GFP worked regarding quantitative UV-detection, which is an unexpected success. Outdoor tests with our lacZ-reporter constructs in sunlight and in shade indicated a light-dependent color change which is visible even to the naked eye. The color of the exposed bacterial suspension changed in the course of irradiation time from light-yellow to a dark greenish blue. This color change is a result of the UV-induced expression of the enzyme beta-galactosidase which catalyzes the reaction of X-Gal to a blue product. With our test series, we furthermore discovered significant differences in the strength of UV detection between rec- and sulA-promotors. precA_LacZ constructs for instance showed less visible color change after 8 minutes of sun exposure than psulA_LacZ constructs.
These successful real-life tests emphasize the possibility to use our biological systems in a real applicational context such as our iGEM-jewelry collection.

After these successful constructions and tests we began to carefully characterize our new biological systems and submitted four new entries to the parts registry. To make this important contribution to the synthetic biology community and to the partsregistry, we adjusted our constructs to the official requirements of the partsregistry such as cloning it into the pSB1C3 plasmid backbone with the required pre- and suffix. Our new three parts are precA_LacZ, precB_LacZ and psulA_LacZ. Additionally, we looked up, ordered, tested and registered the sequence of the recB- promotor, which was not yet available in the registry. First tests showed that UV-induced activity of precB can be compared with those of precA.

With this unpredictable success, we are able to embed this new and innovative UV detection system in a fancy application: iGEMS. iGEMS are small tubes filled with bacterial suspension and integrated into trendy necklaces and bracelets. We developed this fictitious product in order to raise the public awareness of the “invisible danger”, to encourage curiosity. With the establishment of a virtual ‘Online Shop’ we tried to put genetically modified organisms in a modern context and give people a better understanding of synthetic biology.
For us, personally, the project broadened our scientific horizon and gave us the opportunity to learn how to work scientifically. We had a great time together and look forward to present our work to you at the Jamboree 2012.

Outlook

In the past few months, the project was brought up to a great level. We have constructed different BioBricks which were sent to the partsregistry, tested our parts and designed a jewelry collection. But, nevertheless, we have encountered difficulties that could be optimized in the future. We have worked with enthusiasm on the bacterial tool kit which can detect UV radiation and have made a lot of progress, therefore we want to continue working on it.

In the future we want to create a clearer characterization of different members of the same gene family as precA. Our assumption is that DNA damage leads to the successive activation of the different rec genes are activated dependent on the UV radiation dose. By implementing also these genes we want to generate a more gradual UV-response. The successive induction of the different rec promoters could help to determine the exact amount of radiation. This is important to provide the bacterial tool kit with the properties of a dosimeter. We started testing precB and preC. PreB lead to a positive result, whereas induction of preC did not result in the generation of the reporter. Cloning of this part will be repeated.

For the iGEM competition we only tested our constructs by exposing the samples to UV light. We have not analyzed whether the construct would also work with radioactive radiation. Radioactive radiation causes DNA damage different from UV-induced DNA damage, therefore it is not for sure whether precA is activated in the same way as with UV radiation. In principle, we assume that the induction by radioactive radiation would work because the recA promoter is known to be activated by the induced damage. We could not test our constructs with radioactive radiation, because we did not have the right equipment.

We also want to include the constructs with a GFP reporter instead of LacZ in our iGEMS collection. GFP cannot be seen by a color change of the bacterial suspension and normally you need further detection systems for GFP, because it has to be excited by a specific wavelength. To overcome this challenge, we want to combine the iGEMS with LED lamps. LEDs emit light of a certain wavelength, e.g. blue light with 460 nm which is the excitation wavelength of GFP. Therefore, we can make GFP visible using blue LEDs. Furthermore, we could also develop a construct with dtTomato, a very light, red-orange fluorescent protein, as another reporter. By using different fluorescent reporters we could generate a color range from green to orange to red with just one single blue LED.

The system still has to be optimized for real life application. Therefore, it has to be adapted to the sensitivity of mammalian cells towards UV-induced DNA damage. Among other things this would include optimization of the bacterial container which might absorb a certain amount of UV radiation.
Our project comprises a novel idea and opens up new perspectives, especially in Human Practices. The integration of the bacteria into jewelry represents a completely new and consumer-friendly solution to give the public an understanding of synthetic biology. We found an easy way for everyone to protect themselves from sunburn. The method to use dosimeters is old-fashioned, complicated and expensive. Lying on the beach, nobody uses dosimeters because they are too big and uncomfortable. Not only do our fancy iGEMS warn people from too intense UV radiation but also are trendy accessories!

We are looking forward to the iGem HS Jamboree 2012 and to presenting our work to the other teams from all over the world. We hope to get into contact with other pupils interested in synthetic biology.

Achievements

Three different UV-Sensor parts constructed and submitted to the registry
All parts were characterized carefully and show a UV-dose-dependent color-production
Outdoor-Test shows our parts to be working in a real application context
Human Practices - consumer product development for raising the public awareness of the “invisible danger” and presenting our project in an easy-to-understand way
Finally, we learned a lot about synthetic biology, laboratory methods and had a great time working together on our project, and are now looking forward to meeting the other teams at the iGEM HS Jamboree!