Difference between revisions of "Part:BBa K4343079"
(→‎Characterization)
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===Characterization===
 
===Characterization===
 
We construct the plasmid containing promoter TEF, PaDes12, terminator lip2t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then integrated into the Intc site of Po1f. PaDes12 gene was expressed under the control of TEF promoter, which can catalyze C18:1 to C18:2. Then, the activity of ∆-12 desaturase was tested by gas chromatography.
 
We construct the plasmid containing promoter TEF, PaDes12, terminator lip2t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then integrated into the Intc site of Po1f. PaDes12 gene was expressed under the control of TEF promoter, which can catalyze C18:1 to C18:2. Then, the activity of ∆-12 desaturase was tested by gas chromatography.
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<center>https://static.igem.wiki/teams/4343/wiki/puc-huh-intc-pades12-map.png</center>
  
 
===Result===
 
===Result===

Revision as of 11:52, 9 October 2022


pUC-HUH-IntC-PaDes12

PaDes12 gene encodes a ∆-12 desaturase of from Parietiestrium sp. This enzyme can catalyze C18:1 to C18:2. This gene is expressed in Y. lipolytica.

Characterization

We construct the plasmid containing promoter TEF, PaDes12, terminator lip2t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then integrated into the Intc site of Po1f. PaDes12 gene was expressed under the control of TEF promoter, which can catalyze C18:1 to C18:2. Then, the activity of ∆-12 desaturase was tested by gas chromatography.

puc-huh-intc-pades12-map.png

Result

PaDes12 was the least efficient catalyst, with a conversion rate of only 37%. Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 3344
    Illegal XhoI site found at 3377
    Illegal XhoI site found at 8005
    Illegal XhoI site found at 8921
    Illegal XhoI site found at 9212
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 3532
    Illegal AgeI site found at 1422
    Illegal AgeI site found at 1521
    Illegal AgeI site found at 1882
    Illegal AgeI site found at 4585
    Illegal AgeI site found at 4946
    Illegal AgeI site found at 6088
    Illegal AgeI site found at 7310
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1
    Illegal BsaI site found at 1425
    Illegal BsaI site found at 2062
    Illegal BsaI site found at 5126
    Illegal BsaI site found at 5663
    Illegal BsaI site found at 6091
    Illegal BsaI site found at 7313
    Illegal BsaI.rc site found at 1419
    Illegal BsaI.rc site found at 6085
    Illegal BsaI.rc site found at 7307
    Illegal BsaI.rc site found at 7807
    Illegal BsaI.rc site found at 7817
    Illegal BsaI.rc site found at 9235
    Illegal SapI site found at 8733
    Illegal SapI.rc site found at 1787
    Illegal SapI.rc site found at 4851