Team:ETH Zurich/Interplay

From 2012.igem.org

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Contents

Interaction between the laboratory and the modelling

Lab and Modelling, or Modelling in the Lab?

To save time and work, laboratory work and in silico modelling should interact closely so that the model gives useful insights and the laboratory provides results from experimental set ups to refine the model. Here we want to outline how laboratory work and modelling interacted throughout our project.

Photoinduction

Given the four available light receptors available as BioBricks, LovTAP, Cph8, YcgE/F and Ccas, we wanted to know which one is best suitable for our purpose of detecting sun light. Thus we modeled the activating rates of the different receptors when exposed to sunlight, the photoinduction. This helped us to rule out YcgE/F and Ccas that did not give the desired activation under sunlight. We continued with LovTAP and Cph8 for our decoder.

The photoinduction model was also used to derive the activity of our new fusion protein, UVR8-TeTRDBD to get a grasp on its activation due to sunlight.

Circuit modeling

UVR8 Circuit

We modeled the direct UV-B detection by UVR8-TeTRDBD circuit with the downstream components: the PABA producing enzymes and the pigment to get an idea how much PABA the system can produce at steady state.

UVR8 Feedback

We were interested in how the absorption of UV-B radiation by our protection molecule PABA can affect the activation of our UVR8-TetRDBD fusion protein. The negative feedback reduces the output, but has no other adverse effects on the system.

Decoder Circuit LovTAP Cph8

To see how we can indirectly detect sunlight with blue and red light receptors, we came up with the idea and biological implementation of the decoder. Our model showed that the decoder can work in theory and resulted in constraints given the promotor strength in the system.

PABA and the Sun protection factor

Last but not least we were wondering how much PABA we need to produce to achieve a certain Sun Protection Factor (SPF). Or, vice versa, given the amount of PABA our bacteria produced, what is the achieved Sun Protection Factor. This helped to decide for a chorismate accumulating strain to produce more PABA to yield a higher SPF. It also revealed that we also need an UV-A absorbing molecule such as the PABA related [http://en.wikipedia.org/wiki/Padimate_O Padimate O] to reach meaningful SPFs since UV-A and UV-B radiation are likewise dangerous.

Trivia

The modellers produced a month stock of liquid LB media and LB agar, performed a Miniprep and loaded a gel. The lab guys became Photoshop and Illustrator experts for the poster and presentation.

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