Abstract
Thraustochytrid is a promising algal oil resource with the potential to meet the demand for docosahexaenoic acid (DHA). However, oils with high DHA content produced by genetic modified thraustochytrids are not accepted by the food and pharmaceutical industries in many countries. Therefore, in order to obtain non-transgenic strains with high DHA content, a two-stage adaptive laboratory evolution (ALE) strategy was applied to the thraustochytrid Aurantiochytrium sp. Heavy-ion irradiation technique was first used before the ALE to increase the genetic diversity of strains, and then two-step ALE: low temperature based ALE and ACCase inhibitor quizalofop-p-ethyl based ALE were employed in enhancing the DHA production. Using this strategy, the end-point strain E-81 with a DHA content 51% higher than that of the parental strain was obtained. The performance of E-81 strain was further analyzed by component analysis and quantitative real-time PCR. The results showed that the enhanced in lipid content was due to the up-regulated expression of key enzymes in lipid accumulation, while the increase in DHA content was due to the increased transcriptional levels of polyunsaturated fatty acid synthase. This study demonstrated a non-genetic approach to enhance lipid and DHA content in non-model industrial oleaginous strains.
Highlights
Very long-chain polyunsaturated fatty acids (ω-3) (ω-3 PUFAs), such as eicosapentaenoic acid (EPA, C20:5) and docosahexaenoic acid (DHA, C22:6), are considered as the essential fatty acids in human nutrition and health [1,2,3]
Deep-sea fish oil is the traditional source of PUFAs, while it is insufficient to meet the global demand for PUFAs [5,6,7]
Heavy-ion irradiation was applied before adaptive laboratory evolution (ALE) to increase the genetic diversity of starting strains
Summary
Very long-chain polyunsaturated fatty acids (ω-3) (ω-3 PUFAs), such as eicosapentaenoic acid (EPA, C20:5) and docosahexaenoic acid (DHA, C22:6), are considered as the essential fatty acids in human nutrition and health [1,2,3]. Thraustochytrid Aurantiochytrium, a heterotrophic non-photosynthetic protist, is well-known for its capacity to accumulate DHA [8]. Thraustochytrids can use glucose, glycerol, and molasses, etc., as a carbon source for fermentation, while some species have the xylose utilization capacity [11,12,13]. The broad substrate utilization capacity increases the potential of thraustochytrids as microbial cell factories for lipid biosynthesis. Several genetic engineering strategies have been successfully performed to optimize the lipidaccumulating capacity of Aurantiochytrium [15,16,17,18]. An adaptive laboratory evolution (ALE) strategy has been developed in order to obtain non-transgenic strains with high DHA content
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