Part:BBa_K539691:Experience

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Experience by 2012 iGEM NCTU_FORMOSA

 Our team modified Part:BBa_K539691 from 2011 NCTU_ Formosa iGEM team by adding three kinds of zinc finger binding domain genes
preceded in three protein genes(alss, ilvC and ilvD) respectively. Using characteristic of zinc finger, we can improve the result
without adding zinc finger binding domain. More detail information, please refer to Part:BBa_K887000

Experience by 2011 iGEM NCTU_FORMOSA

Analysis of the isobutanol production in different temperature

Figure 1.

(A.) Mean fluorescence intensity (MEFL) at 30℃,37℃,42℃ The vertical axis is mean fluorescence(MEFL), and the horizontal axis is time. We can regard GFP as our target protein,Kivd. Under different temperature, a tendency shows that the production of GFP proteins will increase in lower incubation temperature. The result shows that our low-temperature released device can work at these three temperatures. In this way, we can do the experiment as the same, but changing the genes of our butanol circuit.

(B.) Production of isobutanol at 30℃,37℃,42℃ Cotransform BBa_K539691 and BBa_K539742 into DH5α, so the expression of alss, ilvC, ilvD and kivd are under the control of temperature. Coupling A. and B. diagram, the Kivd protein expression and the production of isobutanol are under temperature control, and both of their production will increase in lower incubation temperature. Therefore, we can conclude that the Kivd protein expression is related to the production of isobutanol.

GC (Gas Chromatography) graph

Figure 2. Cotransform two circuits, Part:BBa_K539691 and Part:BBa_K539742, into DH5α. Incubate DH5α at 37℃ until OD reaches 0.5. Before we switch the temperature to 30℃, we analyze the chemical composition of medium by GC.

Figure 3. Incubate DH5α at 37℃ until OD reaches 0.5, then we switch the temperature to 30℃ incubating for 24hr and analyze the chemical composition of medium by GC.

Comparison of isobutanol production under low-temperature released device or not

Figure 4. Control group (non-temperature controlled device): BBa_K539671 and BBa_K539742 Experimental group(low-temperature released device): BBa_K539691 and BBa_K539742 GC graph in 30℃, 37℃, 42℃

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