Difference between revisions of "Part:BBa K4343069"

Revision as of 09:09, 9 October 2022

pUC-HUH-PaDes17(LEU)

The PaDes17 gene encodes a ∆-17 elongase from Pythium aphanidermatum that catalyzes the conversion of arachidonic acid (ARA; C20:4 n-6) to Eicosapentaenoic acid (EPA), an ω 3 polyunsaturated fatty acid (PUFA).

Characterisation

We constructed a plasmid containing promoter TEF, PaDes17, 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-2 was obtained. The PaDes17 gene was expressed in the presence of the promoter TEF, which can catalyze the conversion of arachidonic acid (ARA; C20:4 n-6) to Eicosapentaenoic acid (EPA). The activity of PaDes17 was examined by gas chromatography.

Result

We integrate one copy of codon-optimized PaDes17 gene from Euglena gracilis under the control of strong promoter PTEF into the genome of engineered strain with one copy of EgDes5 to obtain the engineered strain Po1f-1 by non-homologous end joining (NHEJ) Fig. 1 and Fig. 2 showed that the engineered strain Po1f-2 was able to produce 16.3% of arachidonic acid (ARA; C20:4 n-6) and 2.4% of EPA in the TFAs, which enabled the synthesis of EPA in the Po1f. Sequence and Features