Enhanced Eicosapentaenoic Acid Production via Synthetic Biological Strategy in Nannochloropsis oceanica.

The rational dietary ratio of docosahexaenoic acid (DHA) to eicosapentaenoic acid (EPA) can exert neurotrophic and cardiotrophic effects on the human body. The marine microalga Nannochloropsis oceanica produces EPA yet no DHA, and thus, it is considered an ideal EPA-only model to pursue a rational DHA/EPA ratio. In this study, synthetic biological strategy was applied to improve EPA production in N. oceanica. Firstly, to identify promoters and terminators, fifteen genes from N. oceanica were isolated using a transcriptomic approach. Compared to α-tubulin, NO08G03500, NO03G03480 and NO22G01450 exhibited 1.2~1.3-fold increases in transcription levels. Secondly, to identify EPA-synthesizing modules, putative desaturases (NoFADs) and elongases (NoFAEs) were overexpressed by the NO08G03500 and NO03G03480 promoters/terminators in N. oceanica. Compared to the wild type (WT), NoFAD1770 and NoFAE0510 overexpression resulted in 47.7% and 40.6% increases in EPA yields, respectively. Thirdly, to store EPA in triacylglycerol (TAG), NoDGAT2K was overexpressed using the NO22G01450 promoter/terminator, along with NoFAD1770-NoFAE0510 stacking, forming transgenic line XS521. Compared to WT, TAG-EPA content increased by 154.8% in XS521. Finally, to inhibit TAG-EPA degradation, a TAG lipase-encoding gene NoTGL1990 was knocked out in XS521, leading to a 49.2-65.3% increase in TAG-EPA content. Our work expands upon EPA-enhancing approaches through synthetic biology in microalgae and potentially crops.