ABSTRACT
The fatty acid profile of plant oils determines their quality and uses. Saturated fatty acids are often not desirable from the standpoints of nutrition and some industrial applications. Camelina sativa is a re-emerged oilseed crop, however its oil needs to be improved to meet different application requirements. In this study, saturated fatty acids were greatly reduced by down-regulating genes encoding the fatty acyl-ACP thioesterases (FATB). An artificial microRNA (amiFATB) was created by replacing a microRNA sequence in the camelina Csa-miR159a gene with a FATB gene specific sequence. Seed-specific expression of amiFATB caused a 45% reduction of palmitic acid (16:0) and a 38% reduction of stearic acid (18:0) compared to wildtype seeds. The total saturated fatty acid content was decreased by 35% from 14.6% to 9.4% of total fatty acids. When amiFATB was expressed in a high-oleic acid transgenic line, it caused further increased oleic acid content. This work demonstrates that the FATB genes in camelina can be effectively knocked down by an artificial microRNA targeting gene-specific sequences, thus provides an additional tool to improve seed oils for desired properties.
Subject(s)
Camellia/genetics , Fatty Acids/genetics , Gene Expression Regulation, Plant , Gene Knockdown Techniques , Plant Proteins/genetics , Seeds/genetics , Thiolester Hydrolases/genetics , Camellia/chemistry , Down-Regulation , Fatty Acids/analysis , Genes, Plant , MicroRNAs/genetics , Plants, Genetically Modified/chemistry , Plants, Genetically Modified/genetics , Seeds/chemistryABSTRACT
Camelina sativa is a re-emerging low-input oilseed crop that has great potentials. It is necessary to ameliorate camelina oils for optimized fatty acid composition that can meet different application requirements. Camelina seed contains significant amounts of C20-C24 very long-chain fatty acids (VLCFAs) that may not be desirable. We demonstrated that these VLCFAs can be effectively reduced by deactivating the Fatty Acid Elongase1 (FAE1) in camelina. The allohexaploid camelina contains three alleles of FAE1 genes. Ethyl methanesulfonate (EMS) induced mutation at the FAE1-B gene caused over 60% reduction of VLCFAs in seed. Homozygous knockout mutants were successfully created in a single generation by simultaneously targeting three FAE1 alleles using the CRISPR technology with an egg cell-specific Cas9 expression. VLCFAs were reduced to less than 2% of total fatty acids compared to over 22% in the wild type, and the C18 unsaturated fatty acids were concomitantly increased. The fae1 mutants were indistinguishable from wild type in seed physiology and plant growth. This study demonstrated that the CRISPR/Cas9 technology can be effectively applied to the polyploid crop camelina to rapidly obtain desired traits such as optimal fatty acid composition in its seed oil. Knocking out FAE1 also provides a means to increase the levels of oleic acid or α-linolenic acid in camelina oils that are desirable for industrial or food/feed uses.
