Database: The C. gracea KCS sequence information has been submitted to the EMBL ⁄ GenBank under accession number: bankit1110928.Keywords: 3-keto-acyl-CoA synthase, Cardamine graeca, Nervonic acid, Brassica carinata, Brassica napus, Arabidopsis thaliana, Health and Industry. SummaryNervonic acid 24:1 D15 (cis-tetracos-15-enoic acid) is a very long-chain monounsaturated fatty acid and exists in nature as an elongation product of oleic acid.There is an increasing interest in production of high nervonic acid oils for pharmaceutical, nutraceutical and industrial applications. Using a polymerase chain reaction approach, we have isolated a gene from Cardamine graeca L., which encodes a 3-ketoacyl-CoA synthase (KCS), the first component of the elongation complex involved in synthesis of nervonic acid. Expression of the Cardamine KCS in yeast resulted in biosynthesis of nervonic acid, which is not normally present in yeast cells.We transformed Arabidopsis and Brassica carinata with the Cardamine KCS under the control of the seed-specific promoter, napin. The T 3 generations of transgenic Arabidopsis and B. carinata plants expressing the Cardamine KCS showed that seed-specific expression resulted in relatively large comparative increases in nervonic acid proportions in Arabidopsis seed oil, and 15-fold increase in nervonic acid proportions in B. carinata seed oil. The highest nervonic acid level in transgenic B. carinata lines reached 44%, with only 6% of residual erucic acid. In contrast, similar transgenic expression of the Cardamine KCS in high erucic B. napus resulted in 30% nervonic acid but with 20% residual erucic acid. Experiments using the Lunaria KCS gene gave results similar to the latter. In both cases, the erucic acid content is too high for human or animal consumption. Thus, the Cardamine KCS: B. carinata high nervonic ⁄ highly reduced erucic transgenic seed oils will be the most suitable for testing in pharmaceutical ⁄ nutraceutical applications to improve human and animal health.

A cDNA encoding the Arabidopsis extraplastidic linoleate desaturase (FAD3) was overexpressed in the seeds of wild‐type Arabidopsis and in a mutant line that accumulates high levels of oleic acid. In the transformed wild‐type plants, linolenic acid (18:3Δ9,12,15) increased from 19% to nearly 40% of total seed fatty acids, with a corresponding decrease in linoleate content (18:2Δ9,12). In the high oleate mutant, a large increase in the level of a fatty acid identified by gas‐chromatography/mass‐spectrometry as mangiferic acid (18:2Δ9,15) was observed. The results demonstrate that the polymethylene‐interrupted dienoic fatty acid, mangiferic acid, can be produced in seed oil through the overexpression of a fatty acid n‐3 desaturase.

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The cloning and characterization of a lyso-phosphatidic acid acyltransferase (LPAT2; EC 2.3.1.51) from Tropaeolum majus from a 20,000 EST collection is described. The 1358 bp TmLPAT2 gene encodes a 42.6 kD polypeptide; the primary sequence has a membrane bound O-acyltransferase, MBOAT, Box I motif, and Boxes II, III and IV typical of LPATs from various species. Unlike many LPAT2s, the gene was constitutively expressed in all tissues. The TmLPAT2 functionality was confirmed by expression in a yeast LPAT deletion (SLC1-) mutant. The TmLPAT2 could use a range of acyl-CoAs as acyl donor, including 22:1-CoA and 20:1-CoA and either 18:1-LPA or 22:1-LPA as acyl acceptor. This new LPAT2 could enable the production of Brassica seed oils with enhanced levels of very long-chain fatty acids.