Team:Paris Bettencourt/Model

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@iGEM_Paris

MODELLING

OPTIC MODEL

your text

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders,
which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

SECOND MODEL

your text

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

RNA is a light cost nucleotide material in the cell, We aim to recreate RNA agglomerations as formed in mammalian cells with triple repeat disorders, which show liquid phase separation, forming a organelle-like vesicle, where local concentrations of enzymes can be created.

Logic circuit modeling

Recent work on transcription elements showed that assembling insulated synthetic operator upstream and downstream of a insulated T7 promoter core allowed for a more diverse control of gene expression and a more specific response time (Zong et al., 2017).
More importantly, the expression of a gene regulated by such repressible promoters can be well-described by a simple equation:

α, β , η_A, K_A are respectively the maximal and basal promoter activity, the Hill coefficient and the dissociation constant of the transcriptional activator-promoter core pair. η_R and Κ_R represent the Hill coefficient and dissociation constant of the binding of a repressor to its cognate operator. δ_R represents the relaxation time, the expected time in which an operator is not bound to any repressor.

Making the assumption that the elements are insulated, we can easily combine them to create not single but dually repressible promoters, and predict their performance by generalising equation 1. In Equation (2), the fact that more than one repressor type binding to the promoter was taken into account and changes to relaxation time and the number of total microstates in the equilibrium were made accordingly.

Figure 1:

Using the experimental data acquired from single repressible promoters, we were able to simulate the behavior of corresponding dually repressible promoters. The experimental data was obtained by testing the impact of different configurations of operators (Figure 1) on gene expression. Results from the data obtained in configuration A and B were combined in silico to create a large number of promoters. Indeed, we worked with 11 different repressors, leading to a total of 110 promoters. The results obtained for the entirety of this library are available here. We will focus for now on three promoters with interesting behaviors.

Three representative candidates were chosen for further testing depending on parameter values: the dissociation constant of repressor-operator binding had to be reasonably low and the Hill coefficient η needed to allow for a cooperative behavior.Each promoter was designed to contain two out of the three chosen repressors - TetR, P22c2, HKcI. For each couple of repressors, four different arrangements of the operators were characterized experimentally under different input combinations. More information on the experimental part of this project can be found here.

References

Zong, Y., Zhang, H., Lyu, C., Ji, X., Hou, J., Guo, X., Ouyang, Q. and Lou, C. (2017). Insulated transcriptional elements enable precise design of genetic circuits. Nature Communications, 8(1).

Centre for Research and Interdisciplinarity (CRI)
Faculty of Medicine Cochin Port-Royal, South wing, 2nd floor
Paris Descartes University
24, rue du Faubourg Saint Jacques
75014 Paris, France
bettencourt.igem2017@gmail.com