Team:LMU-Munich/Data/Constitutive

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The constitutive promoters '''P<sub>''liaG''</sub>''' and '''P<sub>''lepA''</sub>''' were evaluated in the reporter vector pSB<sub>Bs</sub>3C-<i>luxABCDE</i>, which contains the ''lux'' operon [[File:Lux operon.png|100px]]. The promoter activity results in gene expression and production of the protein luciferase. The luminescence produced by this protein can be measured with the plate reader ''Synergy2'' ([http://www.biotek.com/ BioTek]) ('''Fig.1'''). </p>
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The constitutive promoters '''P<sub>''liaG''</sub>''' and '''P<sub>''lepA''</sub>''' were evaluated in the reporter vector pSB<sub>Bs</sub>3C-<i>luxABCDE</i>, which contains the ''lux'' operon [[File:Lux operon.png|100px]]. The promoter activity results in gene expression and production of the protein luciferase. The luminescence produced by this protein can be measured with the plate reader ''Synergy2'' ([http://www.biotek.com/ BioTek]) (Fig. 1). </p>
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<font color="#000000"; size="2"><p align="justify">'''Fig. 1: Luminescence measurement of the constitutive ''Bacillus'' promoters P<sub>''liaG''</sub> and P<sub>''lepA''</sub> in the reporter vector pSB<sub>''Bs''</sub>3C-''luxABCDE'''''. OD<sub>600</sub> (up), LUMI (middle) and LUMI per OD<sub>''600''</sub> (down) depending on the time (h) are shown for two different clones (green/blue). Data derive from three independent experiments, the graphs show the mean fo the three experiments and the standard deviation. Curves were fitted over each other (t=0, OD<sub>''600''</sub>=0,3) and smoothed by taking the average of three neighboring values.</p></font>
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<font color="#000000"; size="2"><p align="justify">'''Fig. 1: Luminescence measurement of the constitutive ''Bacillus'' promoters P<sub>''liaG''</sub> and P<sub>''lepA''</sub> in the reporter vector pSB<sub>''Bs''</sub>3C-''luxABCDE'''''. OD<sub>600</sub> (up), LUMI (middle) and LUMI per OD<sub>600</sub> (down) depending on the time (h) are shown for two different clones (green/blue). Data derive from three independent experiments, the graphs show the mean fo the three experiments and the standard deviation. Curves were fitted over each other (t=0, OD<sub>600</sub>=0.3) and smoothed by taking the average of three neighboring values.</p></font>
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===β-galactosidase assays===
===β-galactosidase assays===
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<p align="justify"> The two constitutive promoters '''P<sub>''liaG''</sub>''' and '''P<sub>''veg''</sub>''' were evaluated with the reporter vector pSB<sub>''Bs''</sub>1C-''lacZ'', which contains the ''lacZ'' reporter gene [[File:LacZ.png|50px]] ('''Fig.2''').</p>
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<p align="justify"> The two constitutive promoters '''P<sub>''liaG''</sub>''' and '''P<sub>''veg''</sub>''' were evaluated with the reporter vector pSB<sub>''Bs''</sub>1C-''lacZ'', which contains the ''lacZ'' reporter gene [[File:LacZ.png|50px]] (Fig. 2).</p>
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<p align="justify">Promoter activity results in β-galactosidase expression. The β-galactosidase assays of P<sub>''veg''</sub> and P<sub>''liaG''</sub> were repeated three times. The graph shows data of one representative experiment. In the beginning of the growth curve, both promoters show only low activity before it increases to a maximum value at three hours. It then decreases steadily until reaching initial level after about seven hours (data not shown). Summing up, the time course of activity is very similar for both promoters. The highest β-galactosidase activity (P<sub>''veg''</sub>= Miller units, P<sub>''liaG''</sub>= 12 Miller Units) can be found during the transition from logarithmic to stationary phase.
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<p align="justify">Promoter activity results in β-galactosidase expression. The β-galactosidase assays of P<sub>''veg''</sub> and P<sub>''liaG''</sub> were repeated three times. The graph shows data of one representative experiment. In the beginning of the growth curve, both promoters show only low activity before it increases to a maximum value at three hours. It then decreases steadily until reaching initial level after about seven hours (data not shown). Summing up, the time course of activity is very similar for both promoters. The highest β-galactosidase activity (P<sub>''veg''</sub>= 65 Miller Units, P<sub>''liaG''</sub>= 12 Miller Units) can be found during the transition from logarithmic to stationary phase.
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