This page describes characterisation for part BBa K325909, the lux operon from Vibrio fischeri.

• Description
• Arabinose to light
• H-NS mutants

Description

E.Coli (Invitrogen TOP 10) cells transformed with BBa K325909 (blue light bulb) and BBa 325219 (red light bulb)

This page described the lux operon from Vibrio fischeri. To relieve LuxR control we placed Lux C, D, A, B, E under the pBad promoter.

Figure 1 - E.coli cells (Invitrogen TOP 10) transformed with BBa K325909 in a 96 well plate.

Arabinose to light

This page describes the relationship between Arabinose concentration in the medium with light output. We used a FLUOstar OPTIMA microplate reader to quantify the light output. Protocol and plate reader settings used are given below.

Data

Figure 1 - Light output of <partinfo>K325909</partinfo> as a function of Arabinose concentration in the media. The values correspond to the peak intensity within 5 hours of adding Arabinose to the media. Data points and error bars correspond to the mean and the standard deviation of 3 time repeats.

Figure 2 - Evolution of luminescence with time at different Arabinose concentrations for <partinfo>K325909</partinfo>. Measurements were taken every 30 minutes. Data points and error bars correspond to the mean and standard deviation of 3 time repeats.

H-NS mutants

It has been shown that the expression of the Vibrio fischeri lux operon when cloned into E. coli was repressed. This repression was linked to the nucleoid protein H-NS. To investigate this effect we cloned the operon into mutant E.coli cells in which the expression of the H-NS protein had been modified. We used a FLUOstar OPTIMA microplate reader to quantify the light output. Protocols and plate reader settings used are given below.

Data

Figure 1 - Peak light output from <partinfo>K325909</partinfo> cloned into H-NS mutant JM 230 H-NS -205::tn10. The data points and the error bar are the mean and standard deviation obtained by 3 tim repeats.

Figure 2 - Evolution of light output from <partinfo>K325909</partinfo> cloned into H-NS mutant JM 230 H-NS -205::tn10 with time at different Arabinose concentrations. The data points and the error bar are the mean and standard deviation obtained by 3 time repeats. Measurements were taken every 30 minutes.

Figure 3 - Figure 3 - Peak light output from <partinfo>K325909</partinfo> cloned into H-NS mutant BW25113 DELTA H-NS::kan. The data points and the error bar are the mean and standard deviation obtained by 3 tim repeats.

Figure 4 - Evolution of light output from <partinfo>K325909</partinfo> cloned into H-NS mutant BW25113 DELTA H-NS::kan with time at different Arabinose concentrations. The data points and the error bar are the mean and standard deviation obtained by 3 time repeats. Measurements were taken every 30 minutes.

Compatibility

Chassis: Device has been shown to work in Top 10 (Invitrogen) Plasmids: Device has been shown to work on <partinfo>pSB1C3</partinfo>

References

[1]: S.M. Marques and J.C.G. Esteves da Silva, (2009) Firefly Bioluminescence: A Mechanistic Approach of Luciferase Catalyzed Reactions,Life 61, 6-17.

[2]: T. Nakatsu et al. (2006) Structural Basis for the spectral difference in luciferase bioluminescence, Nature 440(16), 372-376.

[3]: K. Gomi and N. Kajiyama, (2001) Oxyluciferin, a Luminescence Product of Firefly Luciferase, Is Enzymatically Regenerated into Luciferin, The Journal of Biological Chemistry, 276(39), 36508-36513.

[4]: S. Ulitzur, (1998) H-NS controls the transcription of three promoters of Vibrio fischeri lux cloned in Escherichia coli,Journal of Bioluminescence and Chemiluminescence, 13(4), 185-188.

[5]: R.T. Dame et al., (2006) Bacterial chromatin organization by H-NS protein unravelled using dual DNA manipulation,Nature, 444, 387-390.