Team:ZJUT-China/Model

Team:ZJUT-China - 2018.igem.org

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Team:ZJUT-China

Abstract

In order to make sure that our system could be common used by the researchers who want to clean the ARG after experiments, we developed a model to Calculate the point when all ARG are cleaned and the bacteria suicide. We got test data of our parts and then built rate equations. And next we used those rate equations to simulate how our system work and tried to find a better ratio of the vector’s copy number.

System modeling Var1.0

d[express-cas]/dt=[vector]*vm/(1+[light_signal])
d[cut off]/dt=kf*[sg]*[cas9]*[ARG]/(km+[ARG])
Our ODE model of the light-controlled ARG cutting system ver1.0 . We assumed that the expression rate is proportional to the copy number of the vectors, and vm means the mixture rate of one vector express the cas9, [light signal] we use is a dimensionless parameter here, and the cut-off rate of cas9 conform the Michaelis-Menten equation.

Result

This is the result we got by the ODE model of the light-controlled ARG cutting system ver1.0

System modeling Ver3.0

This presents an issue for researchers and factors who wish to make use of our system of the ARG killing parts in a more complex environment. In order to address this issue, we decided to develop a mathematical model of how light signal effects influence the time of cutting off all ARGs and bacteria's suicide induced by our ARG killing system. In the light-controlled ARG cutting system ver3.0 we want to add 3 repressors, one more sgRNA, and a lysin gene to make our cells can cut off all the ARGs and then suicide in chronological order which is designed.

d[express_r1]/t=kf*[vector_1]/(vm+[light signal])
d[degrade_r1]/t=vm*[repressor_1]/([repressor_1]+km)
d[express_r2]/t=kf*[vector_2]
d[degrade_r2]/t=vm*[repressor_2]/([repressor_2]+km)
d[express_r3]/t=kf*[vector_1]
d[degrade_r3]/t=vm*[repressor_3]/([repressor_3]+km)
d[express_cas]/t=kf*[vector_1]*(vm-[repressor_1])
d[express_sg1]/t=0
d[express_sg2]/t=kf*[vector_2]*(vm-[repressor_2])
d[express_ly]/t=kf*[chromosome]*(vm-[repressor_3])
d[cut off_1]/t=kf*[ARG_1]*[sg_1]*[cas9]/(km+[ARG_1])
d[cut off_2]/t=kf*[ARG_2]*[sg_2]*[cas9]/(km+[ARG_2])
Our ODE model of the light-controlled ARG cutting system ver3.0. In this model we considered the expression and degrading rate of the repressor and the effect of the copy numbers of two vectors.

Through changing the copy numbers of two vectors in our system model, we could get a ratio of the copy numbers letting our system do nothing before we give the light signal, while having a higher reacting rate when we want it to work.

Result

Figure 1:when Time>=1t the strength of light signal=0.1,it use 12.5t to cut off all antibiotic resistance gene.
Figure 2:when Time>=1t the strength of light signal=1,it use 12t to cut off all antibiotic resistance gene.
Figure 3:when Time>=1t the strength of light signal=1, and shutdown when Time=2, it use 12t to cut off.

Discussion

Our modeling and analysis focused on the aim of achieving a better theoretical ground to forecast how our system work after giving the light signal. From the test of ODE model of the light-controlled ARG cutting system ver3.0, we found that giving a short pulse light signal can make almost same effects as we constant light signal, so we find a plan to just give a short pulse light signal but make our system still response as fast as it was before, as well as saving energy used to give the light signal.

[1]Quantitative approaches to the study of bistability in the lac operon of Escherichia coli J R Soc Interface. 2008 Aug 6; 5(Suppl 1): S29–S39. Published online 2008 Apr 15. doi:  10.1098/rsif.2008.0086.focus [2]Combinatorial transcriptional control of the lactose operon of Escherichia coli Thomas Kuhlman, Zhongge Zhang, Milton H. Saier, Jr., Terence Hwa Proc Natl Acad Sci U S A. 2007 Apr 3; 104(14): 6043–6048. Published online 2007 Mar 21. doi: 10.1073/pnas.0606717104