We have successfully constructed seven main composite parts that are important to our project, including six multi gene combined expression plasmids by combining β-oxidation pathway genes (ylPOT1, ylMFE1, and ylPOX1-6) together on plasmid pYLXP’, and one assembled Δ6-fatty acid desaturase gene M-Δ6-D into the hp4d-based expression vector p0. The experimental results showed that combined overexpression of genes ylPOT1, ylMFE1, and ylPOX4 or ylPOX5 significantly improve the cell growth of Y. lipolytica with using the edible oil as the substrate. Next, we introduced the Δ6-fatty acid desaturase encoding gene M-Δ6-D into the engineered strains with above improved cell growth by the genomic integration method. As a result, we obtained a titer of 59.3 mg/L γ-linolenic acid from the engineered strain with using the gutter oil as substrate. The result set up a new stage of engineering Y. lipolytica as a sustainable biorefinery chassis strain for the synthesis of γ-linolenic acid with the gutter oil as substrate. Please go to our DESIGN and PARTS page for details.