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Abstract：
Triacylglycerols (TAGs) are the storage oils of plant seeds, and these lipids provide energy for seed germination and valuable oils for human consumption. Three diacylglycerol acyltransferases (DGAT1, DGAT2, and DGAT3) and phospholipid:diacylglycerol acyltransferases participate in the biosynthesis of TAGs. DGAT1 and DGAT2 participate in the biosynthesis of TAGs through the endoplasmic reticulum (ER) pathway. In this study, we functionally characterized CsDGAT1 and CsDGAT2 from camelina (Camelina sativa). Green fluorescent protein-fused CsDGAT1 and CsDGAT2 localized to the ER when transiently expressed in Nicotiana benthamiana leaves. To generate Csdgat1 and Csdgat2 mutants using the CRISPR/Cas9 system, camelina was transformed with a binary vector carrying Cas9 and the respective guide RNAs targeting CsDGAT1s and CsDGAT2s via the Agrobacterium-mediated floral dip method. The EDD1 lines had missense and nonsense mutations in the CsDGAT1 homoeologs, suggesting that they retained some CsDGAT1 function, and their seeds showed decreased eicosaenoic acid (C20:1) contents and increased C18:3 contents compared to the wild type (WT). The EDD2 lines had a complete knockout of all CsDGAT2 homoeologs and a slightly decreased C18:3 content compared to the WT. In conclusion, CsDGAT1 and CsDGAT2 have a small influence on the seed oil content and have an acyl preference for C20:1 and C18:3, respectively. This finding can be applied to develop oilseed plants containing high omega-3 fatty acids or high oleic acid.
摘要：
三酰基甘油（TAG）是植物种子的储存油，这些脂质为种子发芽提供能量，并为人类消费提供宝贵的油脂。三种二酰基甘油酰基转移酶（DGAT1，DGAT2和DGAT3）和磷脂：二酰基甘油酰基转移酶参与TAG的生物合成。DGAT1和DGAT2通过内质网（ER）途径参与TAG的生物合成。在这项研究中,我们在功能上表征了csDGAT1和csDGAT2来自camelina（camelinasativa）。绿色荧光蛋白融合的CsDGAT1和CsDGAT2在烟草叶片中瞬时表达时位于ER。要使用CRISPR/Cas9系统生成Csdgat1和Csdgat2突变体，通过农杆菌介导的花浸方法，用携带Cas9和靶向CsDGAT1s和CsDGAT2s的相应指导RNA的二元载体转化山茶花。EDD1系在CsDGAT1同系物中具有错义和无义突变，表明他们保留了一些CsDGAT1功能，与野生型（WT）相比，它们的种子显示出二十碳烯酸（C20：1）含量降低，C18：3含量增加。与WT相比,EDD2品系具有所有CsDGAT2同系物的完全敲除和稍微降低的C18:3含量。总之,CsDGAT1和CsDGAT2对种子油含量的影响很小，并且分别对C20：1和C18：3具有酰基偏好。这一发现可应用于开发含有高ω-3脂肪酸或高油酸的油料种子植物。
