Difference between revisions of "Part:BBa J45199:Experience"

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Applications of BBa_J45199

Banana odor based reporter of transcriptional signal.

User Reviews

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Characterization

Smell test of banana odor generator

The banana odor generator (BBa_J45200) includes the ATF1 enzyme generator (BBa_J45199) under the control of a constitutive promoter (BBa_R0040).

Smell tests of cultures of wintergreen and banana odor generators (BBa_J45120 and BBa_J45200, respectively)
At the 2006 iGEM Jamboree, we conducted blind smell tests in which iGEM participants had to distinguish between cell cultures producing wintergreen odor, banana odor and the natural fecal odor of E. coli. Of the 116 respondents, 64% were able to correctly identify the culture producing methyl salicylate through its wintergreen bouquet, 87% were able to correctly identify the culture producing isoamyl acetate through its banana bouquet, and 86% were able to correctly identify the laboratory E. coli strain TOP10 through its stink. The error bars are the specific margin of error at 95% confidence. For the smell tests, we used the odor-free chassis (E. coli strain YYC912) for the wintergreen and banana odor generators.

Gas chromatography analysis of banana odor generator

The banana odor generator (BBa_J45200) includes the ATF1 enzyme generator (BBa_J45199) under the control of a constitutive promoter (BBa_R0040).

Verification of odor production by banana odor generator
We successfully designed, constructed and tested biosynthetic devices for banana odor production from exogenously supplied precursor. To confirm that the banana odor generator produced the correct smell compound, we analyzed cultures with the device supplemented with 5mM isoamyl alcohol by gas chromatography. The banana odor generator (BBa_J45200) produced high levels of isoamyl acetate when we added the precursor isoamyl alcohol to the culture medium (A). The cellular chassis alone (E. coli strain TOP10) did not produce isoamyl acetate although we added isoamyl alcohol to the culture medium (B). The retention time of the isoamyl acetate peak from BBa_J45200 is identical to that of the pure isoamyl acetate standard (C). Most E. coli strains produce indole. Octyl acetate was used as an internal standard for all samples containing isoamyl acetate.

Use of ATF1 generator in banana odor generator

The ATF1 enzyme generator (BBa_J45199) is used in both banana odor generators (BBa_J45200 and BBa_J45250).

Growth dependent banana odor production
To demonstrate growth phase dependent banana odor production, we compared the behavior of constitutive and stationary phase dependent banana odor generators (A and B, respectively). We measured isoamyl acetate concentrations of cultures of the constitutive and stationary phase banana odor generators at different culture cell densities (OD600nm) (C). As expected, the stationary phase banana odor generator produced very little isoamyl acetate at low cell densities but its isoamyl acetate production increased with cell density. By comparison, the constitutive banana odor generator produced more isoamyl acetate at lower cell densities than the stationary phase banana odor generator. To visually aid comparison of the two odor generators, an empirical fit to the data for each device is shown.