Difference between revisions of "Part:BBa K4343066"

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We constructed a plasmid containing promoter TEF, EgElo9, and terminator Cyc1t through the T4 ligase. The plasmid was linearized by enzyme cleavage and then transformed into Yarrowia lipolytica and randomly integrated into its genome. EgElo9 gene was expressed under the action of promoter TEF, which can catalyze linoleic acid (LA, C18:2) to eicosadienoic acid (EDA; C20: 2n-6). Subsequently, the activity of EgElo9 was examined by gas chromatography.
 
We constructed a plasmid containing promoter TEF, EgElo9, and terminator Cyc1t through the T4 ligase. The plasmid was linearized by enzyme cleavage and then transformed into Yarrowia lipolytica and randomly integrated into its genome. EgElo9 gene was expressed under the action of promoter TEF, which can catalyze linoleic acid (LA, C18:2) to eicosadienoic acid (EDA; C20: 2n-6). Subsequently, the activity of EgElo9 was examined by gas chromatography.
 
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===Result===
 
===Result===

Revision as of 07:17, 9 October 2022


pUC-HUH-EgElo9

The EgElo9 gene from Euglena gracilis encodes a ∆-9 elongase that catalyzes the conversion of linoleic acid (LA, C18:2) to eicosadienoic acid (EDA; C20: 2n-6)

Usage and Biology

We constructed a plasmid containing promoter TEF, EgElo9, and terminator Cyc1t through the T4 ligase. The plasmid was linearized by enzyme cleavage and then transformed into Yarrowia lipolytica and randomly integrated into its genome. EgElo9 gene was expressed under the action of promoter TEF, which can catalyze linoleic acid (LA, C18:2) to eicosadienoic acid (EDA; C20: 2n-6). Subsequently, the activity of EgElo9 was examined by gas chromatography. testpng.png

50px

Result

We integrate one copy of codon-optimized EgElo9 gene from Euglena gracilis under the control of strong promoter PTEF into the genome to obtain the engineered strain Po1f-1 by non-homologous End Joining (NHEJ). The percentage of eicosadienoic acid (EDA; C20: 2n-6) were 31.8%


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NotI site found at 5470
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 1920
    Illegal XhoI site found at 1953
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 2108
    Illegal AgeI site found at 97
    Illegal AgeI site found at 458
    Illegal AgeI site found at 3161
    Illegal AgeI site found at 3522
    Illegal AgeI site found at 4664
    Illegal AgeI site found at 5673
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1
    Illegal BsaI site found at 638
    Illegal BsaI site found at 3702
    Illegal BsaI site found at 4239
    Illegal BsaI site found at 4667
    Illegal BsaI site found at 5194
    Illegal BsaI site found at 5676
    Illegal BsaI.rc site found at 4661
    Illegal BsaI.rc site found at 4723
    Illegal BsaI.rc site found at 5670
    Illegal BsaI.rc site found at 5940
    Illegal SapI.rc site found at 363
    Illegal SapI.rc site found at 3427