Docosahexaenoic acid (DHA, C22: 6), which can be synthesized by the heterotrophic microalga Crypthecodinium cohnii, has received significant interest recently, due to its important roles in human health. However, the high production cost caused by low productivity and yield is still a major hurdle for industrial production of DHA using C. cohnii. The reduction of starch content has been found to be a promising mean to drive more carbon source to lipid biosynthesis in C. cohnii; however, it usually affects cell growth negatively, which limits its practical application. In this study, atmospheric and room temperature plasma mutagenesis associated with high-throughput screening based on iodine vapour was developed to identify a starch-deficient C. cohnii mutant 16D. The mutant 16D accumulated more biomass, lipid and DHA, with less starch and extracellular polysaccharide. The productivity and the yield of DHA were elevated 1.7 and 1.3 fold, reaching levels of 57.7 mg/L-h and 52.0 mg/g, respectively, in fed-batch fermentation, which are the highest reported so far for DHA production using C. cohnii. LC- and GC–MS based metabolomic analyses revealed multiple possible mechanisms relevant to increased growth and lipid content in mutant 16D, including strengthened Embden-Meyerhof, Krebs cycle and pentose phosphate pathways to provide more CoA, ATP and NADPH for supporting growth and lipid accumulation; and a significant increase in the intracellular abundance of tagatose, which was demonstrated to be involved in lipid content enhancement possibly by activating the triacylglycerol biosynthesis pathway in mutant 16D. The study presents a new strain improvement strategy for DHA-producing C. cohnii and new insights for switching metabolism from starch to growth and lipid in C. cohnii. In addition, the study also demonstrates the inductive effects of tagatose on lipogenesis in heterotrophic microalgae.

可以由异养微藻Crypthecodinium cohnii合成的二十二碳六烯酸（DHA，C22：6）由于其在人类健康中的重要作用而受到了广泛的关注。然而，由低生产率和低产率引起的高生产成本仍然是使用C工业生产DHA的主要障碍。甲藻。淀粉含量的降低已被发现是一种有前途的平均来吸引更多的碳源至脂质生物合成中Ç。科尼; 但是，它通常会对细胞生长产生负面影响，从而限制了其实际应用。在这项研究中，用高通量筛选基于碘蒸气被开发相关联的大气和室温下等离子体诱变以鉴定淀粉缺陷Ç。cohnii突变体16D。突变体16D积累了更多的生物量，脂质和DHA，淀粉和细胞外多糖更少。分批补料发酵的DHA生产率和产量分别提高了1.7和1.3倍，分别达到57.7 mg / Lh和52.0 mg / g的水平，这是迄今为止使用C生产DHA的最高报道。科尼。基于LC-GC和GC-MS的代谢组学分析揭示了与突变体16D中生长和脂质含量增加相关的多种可能机制，包括增强的Embden-Meyerhof，Krebs循环和戊糖磷酸途径，以提供更多的CoA，ATP和NADPH以支持生长和脂质积累; 以及塔格糖的细胞内丰度的显着增加，这可能与激活突变体16D中的三酰基甘油生物合成途径有关，参与了脂质含量的提高。该研究提出了一种用于产生DHA的C的新菌株改良策略。甲藻和新的见解对淀粉代谢转换经济增长和脂质Ç。科尼。此外，该研究还证明了塔格糖对异养微藻中脂肪生成的诱导作用。