Chen Yicun, Cui Qinqin, Xu Yongjie, Yang Susu, Gao Ming, Wang Yangdong
State Key Laboratory of Tree Genetics and Breeding, Chinese Academy of Forestry, Beijing, 100091, People's Republic of China,
Mol Genet Genomics. 2015 Aug;290(4):1605-13. doi: 10.1007/s00438-015-1011-0. Epub 2015 Mar 10.
Genetic engineering to produce valuable lipids containing unsaturated fatty acids (UFAs) holds great promise for food and industrial applications. Efforts to genetically modify plants to produce desirable UFAs with single enzymes, however, have had modest success. The key enzymes fatty acid desaturase (FAD) and diacylglycerol acyltransferase (DGAT) are responsible for UFA biosynthesis (a push process) and assembling fatty acids into lipids (a pull process) in plants, respectively. To examine their roles in UFA accumulation, VfFAD2 and VfDGAT2 genes cloned from Vernicia fordii (tung tree) oilseeds were conjugated and transformed into Rhodotorula glutinis and Arabidopsis thaliana via Agrobacterium tumefaciens. Real-time quantitative PCR revealed variable gene expression levels in the transformants, with a much higher level of VfDGAT2 than VfFAD2. The relationship between VfFAD2 expression and linoleic acid (C18:2) increases in R. glutinis (R (2) = 0.98) and A. thaliana (R (2) = 0.857) transformants was statistically linear. The VfDGAT2 expression level was statistically correlated with increased total fatty acid content in R. glutinis (R (2) = 0.962) and A. thaliana (R (2) = 0.8157) transformants. With a similar expression level between single- and two-gene transformants, VfFAD2-VfDGAT2 co-transformants showed a higher linolenic acid (C18:3) yield in R. glutinis (174.36 % increase) and A. thaliana (14.61 % increase), and eicosatrienoic acid (C20:3) was enriched (17.10 % increase) in A. thaliana. Our data suggest that VfFAD2-VfDGAT2 had a synergistic effect on UFA metabolism in R. glutinis, and to a lesser extent, A. thaliana. These results show promise for further genetic engineering of plant lipids to produce desirable UFAs.
通过基因工程生产含有不饱和脂肪酸（UFA）的有价值脂质，在食品和工业应用方面具有巨大潜力。然而，利用单一酶对植物进行基因改造以生产所需UFA的努力，取得的成功有限。关键酶脂肪酸去饱和酶（FAD）和二酰甘油酰基转移酶（DGAT）分别负责植物中UFA的生物合成（一个推动过程）以及将脂肪酸组装成脂质（一个拉动过程）。为了研究它们在UFA积累中的作用，从油桐种子中克隆的VfFAD2和VfDGAT2基因进行了融合，并通过根癌农杆菌转化到粘红酵母和拟南芥中。实时定量PCR显示转化体中基因表达水平存在差异，VfDGAT2的表达水平远高于VfFAD2。在粘红酵母（R² = 0.98）和拟南芥（R² = 0.857）转化体中，VfFAD2表达与亚油酸（C18:2）增加之间的关系具有统计学上的线性。在粘红酵母（R² = 0.962）和拟南芥（R² = 0.8157）转化体中，VfDGAT2表达水平与总脂肪酸含量增加具有统计学相关性。在单基因和双基因转化体表达水平相似的情况下，VfFAD2 - VfDGAT2共转化体在粘红酵母中显示出更高的亚麻酸（C18:3）产量（增加174.36%），在拟南芥中也有增加（增加14.61%），并且在拟南芥中二十碳三烯酸（C20:3）得到富集（增加17.10%）。我们的数据表明，VfFAD2 - VfDGAT2对粘红酵母中的UFA代谢具有协同作用，对拟南芥的协同作用较小。这些结果为进一步通过基因工程改造植物脂质以生产所需UFA展现出了前景。
