Part:BBa_K4343067
pUC-HUH-EgDes8
The EgDes8 gene encodes a ∆-8 desaturase from Euglena gracilis that catalyzes the conversion of eicosadienoic acid (EDA; C20: 2n-6) to dihomo-γ-linolenic acid (DGLA; C20: 3n-6).
Characterisation
We constructed a plasmid containing promoter TEF, EgDes8, and terminator Cyc1t through the T4 ligase. The plasmid was linearized by enzyme cleavage and then transformed into engineered Yarrowia lipolytica and randomly integrated into its genome. The engineered strain Po1f-1 was obtained, which can catalyze the conversion of eicosadienoic acid (EDA; C20: 2n-6) to dihomo-γ-linolenic acid (DGLA; C20: 3n-6) by expressing EgDes8 gene. And subsequently, the activity of EgDes8 was examined by gas chromatography.
Result
We integrate one copy of codon-optimized EgElo8 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) and DGLA were 20.8% and 8.5%, respectively(fig:po1f-1).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 4994
Illegal XhoI site found at 1920
Illegal XhoI site found at 1953
Illegal XhoI site found at 5315 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE 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 5955 - 1000INCOMPATIBLE 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 5958
Illegal BsaI.rc site found at 4661
Illegal BsaI.rc site found at 5952
Illegal BsaI.rc site found at 6222
Illegal SapI.rc site found at 363
Illegal SapI.rc site found at 3427
Illegal SapI.rc site found at 4939
| None |