Difference between revisions of "Part:BBa K1423006"
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The arsenic inducible banana odor generator consists of the arsenic inducible promoter, J33201, and the composite part J45199, which is made up of a ribosome binding site, the ATF1 gene, and a double terminator. E. coli transformed with this part can convert isoamyl alcohol to isoamyl acetate in the presence of heavy metals such as arsenic and cadmium. The generator may express false results, as the generator will produce isoamyl acetate without the presence of arsenic or other heavy metals. But expression appears to rise until a concentration of 5uM arsenite is injected. Any concentrations larger than this appear to be toxic to E.Coli cells, as the production of isoamyl alcohol drops significantly.
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The arsenic inducible banana odor generator consists of the arsenic inducible promoter, J33201, and the composite part J45199, which is made up of a ribosome binding site, the ATF1 gene, and a double terminator. The ATF1 enzyme converts isoamyl alcohol to isoamyl acetate. E. coli transformed with this part can convert isoamyl alcohol to isoamyl acetate in the presence of heavy metals such as arsenic and cadmium.  
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To test the efficiency of the ATF1 enzyme when regulated by the arsenic inducible promoter, we grew a 50 mL  culture of E. coli transformed with this Biobrick overnight with 50 μL of chloramphenicol, then diluted the culture to OD 0.2 the following day. 2 mL of the diluted liquid culture were then added to 24 test tubes to create smaller liquid cultures. Varying concentrations of isoamyl alcohol and sodium arsenite were added to the 2mL cultures. The contents of each test tube are outlined below.
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1 a-c. 2mL E. coli transformed with BBa_K1423006
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2 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol
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3 a-c. 2mL E. coli transformed with BBa_K1423006
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4 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol
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5 a-c. 2mL E. coli transformed with BBa_K1423006 + 100 μM sodium arsenite
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6 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 0.5 μM sodium arsenite
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7 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 5 μM sodium arsenite
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8 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 10 μM sodium arsenite
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9 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 50 μM sodium arsenite
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10 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 100 μM sodium arsenite
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11. 2mL LB + 5mM Isoamyl Alcohol + 5 mM Isoamyl acetate (standard)
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12. 2mL 2x dilution of the standard
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13. 2mL 10x dilution of the standard
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14. 2mL 25x dilution of the standard
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15. 2mL 50x dilution of the standard
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[https://static.igem.org/mediawiki/2014/b/b6/GCMSresults%282%29.png]
 
[https://static.igem.org/mediawiki/2014/b/b6/GCMSresults%282%29.png]

Revision as of 15:25, 27 October 2014

Arsenic Inducible Banana Odor Generator

WPI_ATF1MetalPromoter.png


The arsenic inducible banana odor generator consists of the arsenic inducible promoter, J33201, and the composite part J45199, which is made up of a ribosome binding site, the ATF1 gene, and a double terminator. The ATF1 enzyme converts isoamyl alcohol to isoamyl acetate. E. coli transformed with this part can convert isoamyl alcohol to isoamyl acetate in the presence of heavy metals such as arsenic and cadmium.

To test the efficiency of the ATF1 enzyme when regulated by the arsenic inducible promoter, we grew a 50 mL culture of E. coli transformed with this Biobrick overnight with 50 μL of chloramphenicol, then diluted the culture to OD 0.2 the following day. 2 mL of the diluted liquid culture were then added to 24 test tubes to create smaller liquid cultures. Varying concentrations of isoamyl alcohol and sodium arsenite were added to the 2mL cultures. The contents of each test tube are outlined below.

1 a-c. 2mL E. coli transformed with BBa_K1423006 2 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol 3 a-c. 2mL E. coli transformed with BBa_K1423006 4 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol 5 a-c. 2mL E. coli transformed with BBa_K1423006 + 100 μM sodium arsenite 6 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 0.5 μM sodium arsenite 7 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 5 μM sodium arsenite 8 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 10 μM sodium arsenite 9 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 50 μM sodium arsenite 10 a-c. 2mL E. coli transformed with BBa_K1423006 + 5mM Isoamyl Alcohol + 100 μM sodium arsenite 11. 2mL LB + 5mM Isoamyl Alcohol + 5 mM Isoamyl acetate (standard) 12. 2mL 2x dilution of the standard 13. 2mL 10x dilution of the standard 14. 2mL 25x dilution of the standard 15. 2mL 50x dilution of the standard



[1]

The link above leads to a quantification of the production of isoamyl acetate normalized to the same construct with the high strength constitutive promoter J23100.



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 255
    Illegal BglII site found at 646
    Illegal BamHI site found at 1973
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 1073
    Illegal SapI.rc site found at 2081