Genes encoding Delta6 desaturase, Delta6 fatty acid elongase, and Delta5 desaturase from the alga, Phaeodactylum tricornutum, were co-expressed in Pichia pastoris to produce arachidonic acid (ARA; 20:4 Delta(5, 8, 11, 14)) and eicosapentaenoic acid (EPA; 20:5 Delta(5, 8, 11, 14, 17)). A panel of Pichia clones carrying progressively increasing copies of the heterologous gene expression cassette was created using an in vitro multimerization approach. ARA and EPA accumulated up to 0.3 and 0.1% of total fatty acids, respectively, in the recombinant P. pastoris carrying with double copies of these three heterologous genes, as compared to 0.1 and 0.05%, respectively, in the recombinant P. pastoris with single copy.
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