Part:BBa_K4343089

pUC-HUH-Scp2-rElo2

PaElo16 gene encodes a C16/18 elongase from rat, which can catalyze C16:0 to C18:0. This gene is expressed in Y. lipolytica.

Characterisation

We constructed a plasmid containing promoter TEF, rElo2, terminator Cyc1t, and homologous arm through the T4 ligase. The plasmid was linearized by enzyme cleavage and then was integrated into the Scp2 site of strain Po1f-3. MaElo16 gene was expressed under the control of TEF promoter, which can catalyze the elongation of C16:0 to C18:0, it also has catalytic activity for C18:1. Then the activity of rElo2 was tested by gas chromatography.

Result

The C16:0 content reduced to 12.4% of TFAs and the percentage of C18:2 they produced were almost doubled. 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 1431
  Illegal XhoI site found at 3465
  Illegal XhoI site found at 3498
  Illegal XhoI site found at 7829
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 3653
  Illegal NgoMIV site found at 6778
  Illegal AgeI site found at 889
  Illegal AgeI site found at 1543
  Illegal AgeI site found at 1642
  Illegal AgeI site found at 2003
  Illegal AgeI site found at 4706
  Illegal AgeI site found at 5067
  Illegal AgeI site found at 6209
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1
  Illegal BsaI site found at 1546
  Illegal BsaI site found at 2183
  Illegal BsaI site found at 5247
  Illegal BsaI site found at 5784
  Illegal BsaI site found at 6212
  Illegal BsaI site found at 6499
  Illegal BsaI.rc site found at 401
  Illegal BsaI.rc site found at 1540
  Illegal BsaI.rc site found at 6206
  Illegal BsaI.rc site found at 7032
  Illegal BsaI.rc site found at 7302
  Illegal BsaI.rc site found at 8464
  Illegal BsaI.rc site found at 8848
  Illegal SapI site found at 6960
  Illegal SapI.rc site found at 1908
  Illegal SapI.rc site found at 4972