We propose post-integration conditions, rationally simplify the complex situation, and split the whole process into a series of chemical reactions. Assuming that intervals between two reactions obey exponential distribution, we use the Gillespie algorithm to calculate the changes in various substances in the system with reference to the Dynamics of the Brusselator . The ideas and methods of this model have strong promotion prospects and adaptability. Our model demonstrates the necessity of using the recombinase(rec) system，the improvement effect of the system after adding the pathways expressing RDF-inhibitor and rec-inhibitor in turn, and the robustness of the model .The experimental team verified some assumptions and results of the model and selected materials according to the parameters of the model.
For judging handbookWhat kind of modeling is being done and what information it will provide?
We use the Gillespie algorithm to calculate the changes in various substances in the systemWhat assumptions were made and why?
One of our assumptions is that the length of the interval between consecutive reactions obeys an exponential distribution so that we can use the Gillespie algorithm  to calculate the changes in various substances in the system.What kind of data was used to build/assess the model
(a) Expression rate of each gene (production rate of related proteins, consumption rate of each protein.
(b) The computing coefficient of each reaction’s rate.
Our model shows the improvement effect of the system after adding the pathways expressing RDF-inhibitor and rec-inhibitor in turn, so as to provide guidance to the experimental team.1. How impressive is the modeling?
Our model has successfully done a proof of concept and played a key role in guiding experimental direction and path design.2. Did the model help the team understand a part, device, or system?
Of course we did3. Did the team use measurements of a part, device, or system to develop the model?
The experimental group verifies the feasibility of the path step by step and roughly determines the parameter dimensions of the model.4. Does the modeling approach provide a good example for others?
The ideas and methods of this model have strong promotion prospects and adaptability. The codes are showed on our wiki , you can copy and run in you Matlab convenient.
|Grec||Gene of recombinase|
|synNotch||Active synNotch on endomembrane system. In our model synNotch is synNotch-TEV|
|GTetR||Gene of operon TetO’s repressor proteins|
|TetR||TetO operon’s repressor proteins|
|tetO||Operon which can be repressed by TetR|
|TetOR||Binary complex of operon TetO and repressor proteins TetR|
|TEV||synNotch’s intracellular domain which is falling off by shearing, an enzyme can divide TetOR binary complex while separate TetR and degrade it.|
|RDF||Reverse recombination factor, which can inverse DNA sequence between sites and make it back to the morphology not affected by rec.|
|Gx||Gene of a protein x|
Tips: Click the button named Aim.
1. Normal protein expression is a reaction which satisfies the post-integration conditions. All reactions which satisfy post-integration conditions can be viewed as one single chemical reaction. part1 . Aim>>To simplify calculation
2. Different reactions in cells occur independently. Aim>>To determine when the next reaction occurs and which reaction occurs.
3. The length of the interval between consecutive reactions obeys an exponential distribution. Aim>>To determine when the next reaction occurs and which reaction occurs.
4. The degradation of protein can be viewed as linear degradation. Aim>>To simplified calculation，this model is also compatible with other methods of calculating degradation for example Michaelis-Menten equation.
5. The repressor effect of the protein can be described by the Hill equation. Aim>>To calculate the expression of rec.
|c21=5;||% TetR Gene expression|
|c22=0.001;||% TetR degradation|
|c23=200;||% generate dipolymer|
|c24=0.003;||% TEV causes decomposition of dipolymer|
|c25=0.000005;||% self decomposition of dipolymer|
|NTetO=5;||% Hill coefficient|
|Swichpoint=2000;||% parameter to the hill equation|
|c31=8;||% rec's Maximum gene expression rate|
|c32=12;||% RDF-inhibitor's Maximum gene expression rate|
|c33=0.01;||% rec degradation|
|c34=0.01;||% rec-RDF-inhibitor degradation|
|c35=2;||% rec-inhibitor Gene expression（No rec-inhibitor can be set to 0）|
|c36=0.1;||% rec-inhibitor in combination with rec|
|c37=200;||% The noise reduction reaction of rec|
|c41=0.0000006;||% rec reverse reaction|
|c51=7;||% Turn on GENE expression downstream|
|c52=0.01;||% RDF degradation|
|c53=200;||% rec-inhibitor in combination with rec|
|c61=0.0000005;||% rec-RDF reverse reaction|
Guide for Experiment
 Gillespie, D. T. Exact Stochastic Simulation of couple chemical reactions. J. Phys. Chem. 81, 2340–2361 (1977).
 Ault, S. & Holmgreen, E. Dynamics of the Brusselator. Math 715 Proj. (Autumn 2002) 1–17 (2003). doi:10.1103/PhysRevE.61.2361
 Stark WM. 2014. The serine recombinase. MicrobiolSpectrum2 (6):MDNA3-0046-2014.
 Morsut, L. et al. Engineering Customized Cell Sensing and Response Behaviors Using Synthetic Notch Receptors. Cell 164, 780–791 (2016).
 Ramos, J. L. et al. The TetR Family of Transcriptional Repressors The TetR Family of Transcriptional Repressors. Microbiol. Mol. Biol. Rev. 69, 326–356 (2005).
 Phan, J. et al. Structural basis for the substrate specificity of tobacco etch virus protease. J. Biol. Chem. 277, 50564–50572 (2002).
 Olorunniji, F. J. et al. Control of serine integrase recombination directionality by fusion with the directionality factor. Nucleic Acids Res. 45, 8635–8645 (2017).