Team:Beijing 4ELEVEN/Model
During our experiment, we need to construct a co-expression system of tyrosinase and adhesive/cohesive proteins. Here we introduced a new EilR-PJExD induction system, and this system was used to control the expression of tyrosinase, while adhesive/cohesive proteins’ expression was controlled by commonly used T7-Lac I system. However, in the course of experiments, the production of these two proteins weren’t smoothly due to high metabolic pressure of the co-expression system. Thus, the addition time of the inducer needs to be precisely regulated to get a better effect.
The T7-Lac I systems has been studied thoroughly previously, while there are still relatively few related researches on our new system. So we need more information about the relationship between induction time and protein expression for this system to improve our project.
We monitored the relationship between CV inducer concentration and protein expression in previous experiments, and determined the optimal inducer concentration of 1μM. Here, we need to detect the relationship between the protein expression level and time under the inducer concentration. We experimentally monitored the OD600 value and the expression of sfGFP (in terms of fluorescence intensity) every two hours after induction from 0 h, and continued until 12 h, with 7 points in total.
Then we fitted the above experimental data through the model, and found that the maximum expression level can be reached within a short time after adding the inducer, which proves that the CV induction effect is outstanding, and we can try to postpone the addition time of CV inducer in subsequent experiments to control the expression of modifying tyrosinase and obtain more adhesives production.
Figure 1. Fitted curve of EilR-PJExD induction system.
The guidance obtained from the above data simulation is still limited for our project. We need the data fitted by the T7-LacI system and our new system at the same time, combined with the growth curve of the cells and the quantitative monitoring of the two proteins to achieve more sophisticated regulation of the co-expression system.
