Team:NCTU Formosa/CI data

High Temperature Induced System – cI Promoter & cI repressor

Data

To verify that this circuit can work as expectation, we use GFP (Green Fluorescence Protein, BBa_E0040) as reporter protein. Part BBa_K098988 is also designed by Team Harvard 2008. In this circuit BBa_K098988, we can regard GFP (Green Fluorescence Protein, BBa_E0040) as reporter protein to tell us that the gene after previous circuit BBa_K098995 can be induced or not(Figure 3).

First day, we incubated E.coli which contains this circuit (BBa_K098988) with GFP (Green Fluorescence Protein, BBa_E0040) at 37℃ overnight. The next day, we transferred them to new three tubes with M9 medium. When the O.D. reached 0.08, we incubated them to 25℃, 37℃, and 42℃, respectively. We collected the samples once an hour, diluting them 200x for accurate GFP (Green Fluorescence Protein, BBa_E0040) measurements. Here is our experiment raw data (Table 1). The mature data will be obtained by unit-conversion according to beads control data (Table 2). We can use Table2 data to draw line chart data, which is easy for us to compare the mean fluorescence at different temperature, 25℃, 37℃, and 42℃. (Figure4) The vertical axis is mean fluorescence, and the horizontal axis is time (unit: an hour).

The following diagrams, Figure5-1.~Figure5-3., are from the Flow cytometer experiment we did. The green fluorescence intensity of E.coli incubated at 25℃ (Figure5-1) and 37℃(Figure5-2) for 9 hours is lower than incubated at 42℃(Figure5-3). Also can refer to the data from Table 1. In Figure5-2., the peak of green fluorescence intensity of E.coli incubated at 37℃ for 9 hours moves right a little bit than incubated at 25℃ for 9 hours. Last, in Figure5-3., incubated at 42℃ for 9 hours, the peak of green fluorescence intensity is significantly stronger than incubated at 25℃ and 37℃ for 9 hours.

Discussion

Form the experimental data, we find that MEFL increases steeply at 42℃ in first two hours, which means the expression of GFP is high. Although after the first two hours its performance decreases and maintains at certain volume, it still expresses a lot more than the other two samples in different temperatures. We can regard GFP as our target protein. This means that our hypothesis has been confirmed that any protein placed after this circuit (BBa_K098995) can be regulated by elevating or lowering temperature. At higher temperature(42℃), our target protein will perform significantly. On the contrary, at lower temperature(25℃), it will express slightly.