Biosynthesis of very long chain polyunsaturated fatty acids in the leafy vegetable chicory

Abstract

The synthesis of very long chain polyunsaturated fatty acids (VLCPUFAs) was investigated in five cultivars of chicory. Genes for enzymes of the ω3/6 Δ 8-desaturation biosynthetic pathways for the formation of C20 VLCPUFAs were inserted into chicory by Agrobacterium-mediated transformation of leaf explants. Plants were transformed by genes encoding Δ 9-specific elongating activity from Isochrysis galbana, Δ 8-desaturase from Euglena gracilis and ∆ 5-desaturase from Mortierella alpina, either separately or in combination; transgenic plants were selected on culture medium containing glufosinate ammonium for those transformed with the ∆ 9-elongase gene alone or in combination with the ∆ 8-desaturase gene, or kanamycin for plants transformed with the ∆ 5-desaturase gene. PCR showed the presence of the transgenes within the genome of selected plants, with RT-PCR confirming gene expression. Gas Chromatography of fatty acid methyl esters extracted from freeze-dried leaves of transgenic plants quantified the synthesis of omega-6 arachidonic acid and its precursors eicosadienoic and dihomo-γ-linolenic acids, and omega-3 eicosapentaenoic acid together with its precursors eicosatrienoic and eicosatetraenoic acids. This is the first report of the production of VLCPUFAs in a leafy vegetable. Since VLCPUFAs are the precursors of prostaglandins, the formation of prostaglandins was also investigated in chicory following a second transformation event using the PGHS-1 gene from Mus musculus.

Metabolomics driven analysis of Erythrina lysistemon cell suspension culture in response to methyl jasmonate elicitation

“…Seeds were surface sterilized in 20% sodium hypochlorite solution for 30 min, washed three times in sterile purified water and placed on agarized Murashige and Skoog (MS; Caisson, Smithfield, USA) germination medium (1962) and incubated under 12 h light period and a temperature of 23 °C ± 1 °C in a culture room [16] . Leaves were excised from 28 d-old seedlings.…”

“…Explants were cultured on 25 mL aliquots of MS supplemented with either 1 mg L −1 or 2 mg L −1 of each of kinetin (Kin; Acros, Geel, Belgium) and 2,4-dichlorophenoxyacetic acid (2,4D; Acros, Belgium) in addition to 30 g L −1 sucrose (El-Nasr, Alexandria, Egypt), and semi-solidified with 0.8% (w/v) agar (Roko, Llanera – Asturias, Spain), pH 5.6, in a 9 cm diameter Petri dishes. The explants were transferred onto fresh media, until callus was produced [16] .…”