Increasing erucic acid content in the seed oil of rapeseed (Brassica napus L.) by combining selection for natural variation and transgenic approaches

Abstract

Erucic acid (22:1) obtained from the seed oil of high erucic acid rapeseed (HEAR) is of interest for the oleochemical industry. It is used in the plastic film, nylon, lubricant and emollient industries. Currently available conventional HEAR cultivars contain about 50% 22:1 in the seed oil. A substantial increase of the 22:1 content of the rapeseed oil would significantly reduce processing costs and could increase market prospects. Crossing of conventional HEAR to rapeseed with reduced contents of linoleic acid (18:2) and linolenic acid (18:3) did result in recombinant high erucic low polyunsaturated fatty acid (HELP) F3-plants which, however, did not show an increased 22:1 content compared to the parental HEAR genotype. This indicated that the â-ketoacyl-CoA synthase (KCS; fae1 gene) activity, the enzyme responsible for the fatty acid elongation from oleic acid (18:1) to eicosenoic acid (20:1) and to erucic acid, may be limiting. Furthermore, the rapeseed native lysophosphatidic acid acyltransferase (Bn-LPAAT) does not accept erucic acid as a substrate for insertion into the central sn-2 position of the triacylglycerol backbone. Here, the LPAAT enzyme from Limnanthes douglasii (Ld-LPAAT) has been found to preferentially insert erucic acid into the sn-2 position. The main objective of the present study was to increase 22:1 content in the seed oil of rapeseed through the genetic combination of alleles from a HELP rapeseed type with transgenic rapeseed over expressing the Bn-fae1 and expressing the Ld-LPAAT gene and to study trait inheritance in segregating populations. To this end, two separate experiments were conducted using HELP line and two different sources of transgenic rapeseed carrying one respectively two transgene copies of Ld-LPAAT-Bn-fae1. In both cases the Ld LPAAT and the Bn-fae1 gene were under control of the seed specific napin promoter. In the second experiment F1-plants derived from the cross between transgenic 361.2B and 6575 1 HELP were used to produce F2-seeds. 220 randomly chosen F2-seed s were sown in the green house and F3-seeds were harvested from the selfed F2-plants. F3-seeds from 41 F2-plants with the highest erucic acid content in the seed oil along with the parental lines were tested in a greenhouse experiment in a randomized complete block design with five replicates. F4-seeds obtained after selfing were analyzed for seed quality traits as described above. The results from the fatty acid analysis of the F2-plants (F3-seeds) showed a large variation in erucic content ranging from 44 to 72%. F3-lines (F4-seeds) were identified which contained up to 72% erucic acid (mean) in the seed oil. This compares favourably with the 63.2% of the transgenic parent 361.2B and 49.6% of the non-transgenic 6575-1 HELP parents. The 72% erucic acid content achieved in the present study mark a major breakthrough in breeding high erucic acid rapeseed.