Composite

Part:BBa_K1893003

Designed by: Alyssa Henderson, Aditi Satija   Group: iGEM16_Imperial_College   (2016-10-14)
Revision as of 18:22, 22 October 2016 by ASatija (Talk | contribs)


Rhl receiver with GFP reporter (RhlR+pRhl+GFP)

The gene encoding the transcriptional activator RhlR is downstream a constitutive Anderson promoter, j23101. RhlR is activated by C4-HSL. Included in the construct is the RhlR-activated promoter pRhl upstream a reporter sequence which includes GFP


Usage and Biology

IC16 BBa K1893003 characterisation.png

Figure 1. Characterisation of the Rhl response device (BBa_K1893003). (A) Transfer function curve of normalised fluorescence against cognate inducer C4-AHL concentrations. (B) Heat map of normalised fluorescence of RhlR-GFP system over a range of AHL concentrations: (i) Binding of RhlR-GFP to its cognate AHL (C4 AHL). (ii) Binding of RhlR-GFP to 3 non-cognate AHLs (3O-C6 AHL, 3O-C12 AHL, 3OH-C14 AHL). (C) Transfer function curves of normalised fluorescence against non-cognate inducer AHL (C4 AHL) concentrations to investigate inducer AHL crosstalk: (i) C6-AHL (3O-C6 AHL) of the Lux system (ii) C12-AHL (3O-C12 AHL) of the Las system (iii) C14-AHL (3O-C14 AHL) of the Cin system. Experiments were performed in E. coli Top10 cell strain cultured at 37°C. Normalised fluorescence was calculated by dividing fluorescent signal by cell density (OD600). Fluorescence measurements were recorded at 180 minutes. Reported values represent the mean normalised fluorescence value from 3 technical repeats and error bars represent standard deviation of these.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 301
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 776
    Illegal BsaI.rc site found at 1658


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Categories
Parameters
None