Composite

Part:BBa_K1149009

Designed by: Margarita Kopniczky   Group: iGEM13_Imperial_College   (2013-09-13)

Cutinase-Like Enzyme

Encodes a lipase that hydrolyses compound polylactic acid.

Source organism: Cryptococcus sp. strain S-2

Characterisation

Our Cutinase-like enzyme (CLE) construct contains sfGFP within an operon and therefore fluorescence can be utilised to determine if expression is being induced by addition of Xylose.

450px-CLE.png

CLE growth assay. E. coli (MG1655) transformed with CLE BBa_K1149009 were grown with either 0% or 2% Xylose to induce CLE and sfGFP expression. Induction with xylose shows growth inhibition. Ideally you would normalise the fluorescence with OD600 to give fluorescence/cell in order to get the actual difference in induction expression. A two-tailed t-test shows that p = 0.4308 > 0.05, therefore there is no statistical difference between induced and uninduced. Growth was at 37°C in LB with shaking. Error bars are SEM, n=4. Figure made by Imperial College London 2013 iGEM.

450px-CLE_fluor.png

CLE induction assay. E. coli (MG1655) transformed with CLE BBa_K1149009 were grown with either 0% or 2% Xylose to induce CLE and sfGFP expression. CLE induction does not seem to increase the amount of fluorescence emitted by the GFP. Growth was at 37°C in LB with shaking. Error bars are SEM, n=4. Figure made by Imperial College London 2013 iGEM.

Conclusion: There is no growth inhibition caused by induction, nor is there any significant fluorescence.

References:

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 99
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 464
    Illegal AgeI site found at 1229
  • 1000
    COMPATIBLE WITH RFC[1000]


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