Difference between revisions of "Part:BBa K4343078"
(→‎Characterization)
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===Characterization===
 
===Characterization===
 
We construct the plasmid containing promoter TEF, CcDes12, terminator lip2t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then integrated to the Intc site of Po1f. CcDes12 gene was expressed under the control of TEF promoter, which can catalyze C18:1 to C18:2. Then, the activity of Cc12Des was tested by gas chromatography.
 
We construct the plasmid containing promoter TEF, CcDes12, terminator lip2t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then integrated to the Intc site of Po1f. CcDes12 gene was expressed under the control of TEF promoter, which can catalyze C18:1 to C18:2. Then, the activity of Cc12Des was tested by gas chromatography.
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<center>https://static.igem.wiki/teams/4343/wiki/puc-huh-intc-ccdes12-map.png<center>
  
 
===Result===
 
===Result===

Revision as of 12:12, 9 October 2022


pUC-HUH-IntC-CcDes12

CcDes12 gene encodes a ∆-12 desaturase from Coprinus cinereus, which can catalyze C18:1 to C18:2. This gene is expressed in Y. lipolytica.

Characterization

We construct the plasmid containing promoter TEF, CcDes12, terminator lip2t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then integrated to the Intc site of Po1f. CcDes12 gene was expressed under the control of TEF promoter, which can catalyze C18:1 to C18:2. Then, the activity of Cc12Des was tested by gas chromatography.

puc-huh-intc-ccdes12-map.png<center>

Result

The conversion rate of CcDes12 from C18:1 to C18:2 is 62.7%.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 6185
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 8581
    Illegal XhoI site found at 3344
    Illegal XhoI site found at 3377
    Illegal XhoI site found at 6321
  • 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 6405
  • 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 6393
    Illegal BsaI.rc site found at 1419
    Illegal BsaI.rc site found at 6085
    Illegal BsaI.rc site found at 7152
    Illegal BsaI.rc site found at 7433
    Illegal BsaI.rc site found at 7933
    Illegal BsaI.rc site found at 7943
    Illegal BsaI.rc site found at 9465
    Illegal SapI.rc site found at 1787
    Illegal SapI.rc site found at 4851