Difference between revisions of "Part:BBa I13015"

Latest revision as of 17:43, 17 October 2018

3OC₆HSL Sender controlled by pBad

The pBad promoter controlling a LuxI sender device. The protein is an untagged LuxI enzyme which produces an AHL (3OC₆HSL).

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 1205
21
INCOMPATIBLE WITH RFC[21]
Illegal BglII site found at 1873
Illegal BamHI site found at 1144
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 979
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI site found at 961

Activity compared to synthetic 3OC6 HSL

Team USP-Brazil (2018) characterized this part by using it in a quorum sensing system along with LuxR and promoter pLux (see BBa_K2771030), in an assay comparing pBad at full activity (2.5mM arabinose) and a quantity known to activate well pLux (10^-4 mM HSL). Arabinose induction showed an equilibrium about 7x higher than with just synthetic HSL, and a bigger EC50, due to the necessity of transcription of two genes and signal production by LuxI for the reporter to be seeable:

Figure 2: Liquid culture assay with 2.5 mM arabinose or 10^-4 mM 3OC6 HSL induction at 0h. Arabinose induction shows clearly higher concentration of HSL than in the synthetic HSL curve, showing higher curve inclination and higher equilibrium point. Fluorescence measured as a ratio of a YFP reporter with the quorum sensing promoter, normalized by the value of a constitutive CFP reporter, proxy for cell conditions and environmental influence

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	Team USP-Brazil (2018) characterized this part by using it in a quorum sensing system along with LuxR and promoter pLux (see [https://parts.igem.org/Part:BBa_K2771030 BBa_K2771030]), in an assay comparing pBad at full activity (2.5mM arabinose) and a quantity known to activate well pLux (10<sup>-4</sup> mM HSL). Arabinose induction showed an equilibrium about 7x higher than with just synthetic HSL, and a bigger EC50, due to the necessity of transcription of two genes and signal production by LuxI for the reporter to be seeable:		Team USP-Brazil (2018) characterized this part by using it in a quorum sensing system along with LuxR and promoter pLux (see [https://parts.igem.org/Part:BBa_K2771030 BBa_K2771030]), in an assay comparing pBad at full activity (2.5mM arabinose) and a quantity known to activate well pLux (10<sup>-4</sup> mM HSL). Arabinose induction showed an equilibrium about 7x higher than with just synthetic HSL, and a bigger EC50, due to the necessity of transcription of two genes and signal production by LuxI for the reporter to be seeable:

−	[[File:T--USP-Brazil--ara HSL assay.png\|500px\|thumb\|none\|alt=experiment 1.\|Figure 2: Liquid culture assay with 2.5 mM arabinose or 10<sup>-4</sup> mM 3OC6 HSL induction at 0h. Arabinose induction shows clearly higher concentration of HSL than in the synthetic HSL curve, showing higher curve inclination and higher equilibrium point]]	+	[[File:T--USP-Brazil--ara HSL assay.png\|500px\|thumb\|none\|alt=experiment 1.\|Figure 2: Liquid culture assay with 2.5 mM arabinose or 10<sup>-4</sup> mM 3OC6 HSL induction at 0h. Arabinose induction shows clearly higher concentration of HSL than in the synthetic HSL curve, showing higher curve inclination and higher equilibrium point. Fluorescence measured as a ratio of a YFP reporter with the quorum sensing promoter, normalized by the value of a constitutive CFP reporter, proxy for cell conditions and environmental influence]]