RNAi-mediated Down-regulation Of BnFAE1 Expression And Erucic Acid Accumulation In Brassica Napus L.

Erucic acid accumulated in rapeseeds is nutritionally unhealthy and undesirable for human consumption, resulting in development of new cultivars with low erucic acid (LEAC) as a source of oils.Erucic acid (C22:1), a monounsaturated very long chain fatty acid (VLCFA), is the major constituent of storage fatty acids in seeds of the members of Cruciferae. The biosynthesis of VLCFA such as erucic acid has been elucidated in B. rapa, B. napus, Crambe abyssinicum and Simmondsia chinensis. In vitro, chain length of C18 fatty acid as precursors are elongated by an enzyme complex known as elongase complex, leading to the synthesis of C22 fatty acid. The rate-limiting enzyme of the elongase complex,β-ketoacyl CoA synthase (KCS), was shown to be encoded by fatty acid elongase 1 (FAE1) gene and responsible for the biosynthesis of erucic acid.In the recent years, RNA interference (RNAi) has proven to be an effective strategy to knock out homologous genes in a wide range of plant species. Based on its principle, a new generation of vector, consisting of an inverted target sequence separated by an intron as a loop, is required to efficiently improve gene inactivation. FAE1 gene is a rate-limiting gene concerned in erucic acid biosynthesis in Brassica. To inhibit its biosynthesis in rapeseed, an RNAi vector pBERi containing an inverted target sequence of BnFAE1.1 gene, an intron of RBCS gene from pea between the inverted sequences was constructed, and transformed into rape mediated by Agrobacterium-infection. The desired sequence integrated into rape(Brassica napus L.) genome DNA evidenced by histo-chemic staining, PCR and Southern blotting analysis. FAE1 gene in T1 developing seeds from 19 transgenic lines was significantly weakened, which was proved by semi-quantitative RT-PCR. Furthermore, erucic acid content in transgenic seeds only accounted for 0.18-16.02% of total fatty acids, and that was greatly lower than that in wild type rapeseeds and pCAMBIA3301 transgenic rapeseeds (about 40%). In sum, expression of FAE1 gene, and biosynthesis of erucic acid in rapeseeds are efficiently down-regulated mediated by FAE1 RNAi. This study provides valuable data for developing new rape germplasm free of erucic acid via gene engineering.