Timer Plus Sumo

In this section the system of Figure 1 is modeled. The structure of the timer is very similar version of the timer compared to the construct of iGEM TU Delft team 2009. Here the input is changed to a T7 promoter and the output to Ulp-1. Furthermore, the Ulp-1 cleaves off� the SUMO from the peptide combined with the SUMO.

The most important aspect here is the modeling of the ‘time delay’: What is the difference in time between inducing and the output. In our case we have two outputs, a small delay must also be present between those two.

At the moment only one output is currently modeled, the ULP-1 protease. Results of this simulation are shown in Figure 1, here the time delay is about … minutes. Next to add is the interaction with the peptide tagged with SUMO and the ULP-1 protease. The second output will be added later (the kill switch cassette).

The peptide is expressed with a SUMO tag added to it, expressing the protease will cleave of the SUMO. It is important to accurately know this cleavage rate in order to describe the rate of ‘free’ peptide production.

Kill switch

The kill switch contains two important parts the expression of holin and the antiholin. These two are important tuning parameters to make the system stable and robust. It will be essential to make sure that the ‘free’ peptide reaches a steady state before lysis of the cell. Making sure of this yields the most efficient system, lysis before the cleavage is done yields less active peptides. The tuning parameters will be the respective RBS and the const. promoter, a thorough sensitivity analysis will be performed on this model to derive the optimal conditions and robustness.