Mutations in the FAD3 Genes of the Brassica C Genome: Can Linolenic Acid be Virtually Eliminated from B. napus Canola Seed Oil?.

Edible oil containing α-linolenic acid (ALA) offers health benefits; however, this fatty acid rapidly oxidizes resulting in oil which is unstable under frying conditions. Therefore, decreasing the content of ALA in Brassica napus (AC genome, n = 19) canola seed oil is desirable. ALA content in seed oil is regulated by the FATTY ACID DESATURASE 3 (FAD3) gene, which encodes an enzyme that catalyses the conversion of linoleic acid (LA) into ALA. The B. napus genome carries at least six FAD3 genes, all of which might be involved in the biosynthesis of ALA. Therefore, mutations in all of these genes could be required to reduce the content of ALA to near zero in B. napus. This would be an incredibly difficult task requiring an extremely large mutagenized population. Conversely, B. oleracea (C genome, n = 9) and B. rapa (A genome, n = 10), which are the progenitor species of B. napus, carry only half the number of FAD3 genes; therefore, it would be far easier to achieve this in these species. Once the desired mutant gene(s) is identified in the parental species, it could subsequently be utilized in B. napus breeding. With this view, B. oleracea seeds were treated with 5% and 0.5% ethyl-methane-sulphonate (EMS) and two mutagenized populations were generated. Selection for low ALA content was carried out in these populations up to the M₇ generation and mutant lines with <2.0% ALA were obtained. Molecular analysis of the lines generated from 5% EMS treatment revealed that the mutation was due to a single nucleotide substitution from G to A in exon 3 of the ‘class b’ FAD3 (BoFAD3-1) gene. In the case of the low ALA lines derived from the 0.5% EMS treatment, the phenotype was due to a nonsense mutation within the ‘class a’ FAD3 gene (BoFAD3-2). Expression of the coding regions of these FAD3 genes (BoFAD3-1 and BoFAD3-2) from the mutant B. oleracea lines in yeast (Saccharomyces cerevisiae) cultures resulted in significantly reduced conversion of LA to ALA. Thus, the mutant B. oleracea lines developed in this research could potentially be used in breeding for the development of B. napus cultivars with further reductions in ALA content, as well as in the breeding of low-ALA B. carinatacultivars.