Genetic transformation of the oilseed crop camelina using immature zygotic embryos

Camelina is an oilseed crop with particularly useful fatty acid and amino acid profiles of its seeds, high resilience to abiotic and biotic stresses, and a short life cycle. Previous genetic engineering approaches in camelina have largely relied on the floral dip method which is, however, associated with genotype-dependent efficiency and incompatibility with methods of direct biomolecule delivery.

Here, we established a novel method of transgenesis for camelina, taking advantage of the high regenerative capacity of immature embryos. Various culture conditions and treatments were experimentally validated, which included the duration of explant pre-cultivation, wounding of explants and its time of application, Agrobacterium strain and density of inoculum, acetosyringone concentration, duration of explant-Agrobacterium co-cultivation, as well as application time and concentration of the selective agent hygromycin. We provide convergent evidence of stable transgenicity and transgene inheritance by (1) selection for resistance to hygromycin, (2) PCR, (3) detection of the transgene product GFP, and (4) DNA gel blot analysis involving primary transgenic plants and segregating progeny. Primary transgenics examined in detail featured one to three T-DNA integration loci, with one to seven T-DNA copies being integrated in total per plant. The established method proved efficient across all three tested accessions including two current cultivars, whereby transformation efficiencies, determined as PCR-positive primary transgenic plants related to agro-inoculated explants, of between 13 and 17% were obtained.

With this method, we provide a viable platform for the functional validation of genes-of-interest and for biotechnological improvements of plant performance and quality features in camelina.