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Team:IPN-UNAM-Mexico/Results
From 2009.igem.org
Theoretical Results
As we could see in the first simulations in Comsol Multiphysics, with our estimated diffusion coefficients and the classical activator inhibitor dynamic equations, we recovered a spotted pattern.
We believe the modelling part of our project is a very strong component of it. First of all, the system was designed to fulfill the requirements of a theoretical proposal by A. Turing of a reaction-diffusion model on morphogenesis, more specifically an activator-inhibitor system. These requirements were checked by calculating and using the diffusive properties of the components involved (AHLs) in our system on the Geirer and Meihnardt model of activator-inhibitor, successfully predicting the generation of patterns in our design.
Later, we wanted to recreate the generation of a pattern from its very biochemical basis to be able to predict under what conditions the pattern would form and how it would be, so we described all the biochemical steps involved in our network (form transcription to complexes formation) by constructing a kinetic model with parameters available in the literature. From this kinetic model we derived our ordinary differential equations and used it to make a spatial simulation of the behavior of a colony, predicting the formation of some kind of instability but not yet a spatiotemporal pattern.
Even when there was no explicit pattern formation on the detailed model, we did obvserve it in our simulation of the general model of activator-inhibitor; so we will explore in the future other approaches to figer out if the predictions of this model are conclusive or not, like stochastic approaches. And we will also have to compare these results with the experimental results that we will obtain with the fully implemented system.
Experimental Results
During this months we acomplished 11 ligations from already available BioBricks, this ligations resulted in the synthesis of 11 new BioBricks which were already sent to the iGEM HQ, to finish the entire system we have only 1 ligation left. Once we acomplish this we will be able to finally test our system as we originally concieved.
We also have preeliminary experiments confirming that the regulatory IPTG module coupled to the Activator module works as we expected, as can be seen in the Parts section
Final Conclusions
We were able to build most of our original design (1 biobrick ligation remaining) and to experimentally test the functioning of one of our regulatory modules coupled to the autocathalytic module.
Besides having one regulatory module working as expected we also have the support from our models on the formation at least of spatiotemporal instabilities, this sustents the validity of our design on its potential to emulate an activator-inhibitor system.
The fact that our designed system is able to behave in this way under certain conditions provided by the regulatory modules shows that the original assumptions on A. Turing’s model have the potential to naturally have emerged in living organisms during evolution, and to be the underlying mechanism by which some of the naturally founded patterns are formed during morphogenesis.
We also realized during the implementation of the project that there are some details, both on the modelling and in the design, we could improve to make a better approximation of the activator-inhibitor system.
We are very excited about the reachings that our project could have if we can successfully implement it, and our results show we are going on the right direction. This opens the potential of synthesizing compact genotipes that can display phenotypes beyond the explicit coding of their genomes, as actually happens in nature.
Further work
The first step after the Jamboree will be finish our last biobricks' ligation, test our system in different conditions and concentrations of IPTG and aTc and characterize the functioning of the different modules relative to the whole net. Then we can really see if our system really works in the way we thought it would. Besides we need different approaches to mathematically model the interactions that take part in our design in order to accurately predict its behavior.
As extensions of our project we are planning to work with this system in eukariotic tissues and replace the IPTG and aTc depedence with another kind of synthetic morphogens and the GFP E. colis with melanocytes to reproduce pigmentation patterns and fully demostrate Turing's work and make a great advance developmental biology.
