Free dihomo-γ-linolenic acid (DGLA), a polyunsaturated free fatty acid (FFA), is a precursor of the eicosanoid prostaglandin E1 and is expected to be a source material for artificial production. We previously constructed the Aspergillus oryzae mutant strain ARA1 that produced free DGLA from the disruptant of faaA, an acyl-CoA synthetase gene, where FFA productivity increased by 9.2-fold compared with that of the wild-type strain. Here, we aimed to achieve enhancement of free DGLA productivity. Because saturated FFAs, such as palmitic acid and stearic acid, accounted for about 45% and 25% of total FFAs produced by ARA1, respectively, we used a strategy to facilitate elongation and desaturation of these FFAs to oleic acid and linoleic acid by overexpressing genes encoding elongase, Δ9-desaturase, and Δ12-desaturase originally expressed in A. oryzae. Ten genes were predicted to encode desaturases, and their overexpression DNA constructs were introduced into ARA1. AO090001000224 and AO090011000488 facilitated Δ12-desaturation and Δ9-desaturation most, respectively, following overexpression. Next, ARA1 strain was modified to DGLA1cre strain for producing free DGLA as a final product, and co-overexpression of these two desaturase genes was then introduced to DGLA1cre. Moreover, single overexpression of two genes predicted to encode elongases was additionally introduced, and only AO090003000572 facilitated elongation. Consequently, in the co-overexpression mutant of AO090001000224, AO090011000488, and AO090003000572, free DGLA content ratio increased by 1.8-fold from ARA1 to 14.5%, and the productivity also increased by 1.8-fold to 0.086 mmol/g-dry-cell-weight. The yield was 284 mg/L. These findings provided insights into strategies for improving microbial production of polyunsaturated FFAs.

游离二高-γ-亚麻酸（DGLA）是一种多不饱和游离脂肪酸（FFA），是类二十烷酸前列腺素E1的前体，有望成为人工生产的原料。我们先前构建了米曲霉突变株ARA1，该突变株从faaA的破坏者中产生了游离的DGLA。，是一种酰基辅酶A合成酶基因，与野生型菌株相比，其FFA生产率提高了9.2倍。在这里，我们旨在实现免费DGLA生产率的提高。因为棕榈油和硬脂酸等饱和FFA分别占ARA1产生的FFA总数的45％和25％，所以我们采用了一种策略，通过过度表达基因来促进这些FFA向油酸和亚油酸的延伸和去饱和。编码最初在米曲霉中表达的延伸酶，Δ9-去饱和酶和Δ12-去饱和酶。预测有十个基因编码去饱和酶，并将它们的过表达DNA构建体引入ARA1。过度表达后，AO090001000224和AO090011000488分别最大程度地促进了Δ12去饱和和Δ9去饱和。接下来，将ARA1菌株修饰为DGLA1cre菌株，以产生游离的DGLA作为最终产物，然后将这两个去饱和酶基因的共过表达引入DGLA1cre。此外，另外引入了两个预计编码延伸酶的基因的单一过表达，并且仅AO090003000572促进了延伸。因此，在AO090001000224，AO090011000488和AO090003000572的共过量表达突变体中，游离DGLA含量比从ARA1增加了1.8倍，达到了14.5％，生产力也提高了1.8倍，达到0.086 mmol / g干细胞重量。产量为284mg / L。