Part:BBa_K4343078

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.

Result

The conversion rate of CcDes12 from C18:1 to C18:2 is 62.7%. (Fig: Polf-5)

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