Development of high oleic and low linoleic acid transgenics in a zero erucic acid Brassica juncea L. (Indian mustard) line by antisense suppression of the fad2 gene

A zero erucic acid (C22: 1) line of Brassica juncea (VH486), adapted to the agronomic conditions of Northern India, has been modified for its fatty acid composition in the seed oil with antisense constructs using the sequence of fad2 gene of B. rapa. The full-length B. rapa fad2 cDNA sequence was determined by 5' and 3' RACE of a partial sequence available in the EST database. Construct pASfad2.1 contained 315 to 1251 bp and construct pASfad2.2 contained 1 to 1251 bp fragment of the fad2 gene, both in antisense orientation, driven by a truncated napin promoter. Analysis of the levels of linoleic acid (C18: 2) in the BC1 seeds of single-copy transgenics showed that the construct pASfad2.2 gave better suppression of the fad2 gene as compared to the construct pASfad2.1. The BC1 transgenic seeds containing the pASfad2.2 construct segregated into two distinct classes of C18: 2 > 20% (putative null homozygotes) and C18: 2 < 20% (putative heterozygotes) in a 1: 1 ratio, while the T1 seeds segregated into three classes, C18: 2 > 20%, C18: 2 between 12% and 20%) and C18: 2 < 12% (putative homozygotes) in a 1: 2: 1 ratio. Putative homozygous T1 seeds (C18: 2 < 12% analyzed by the half-seed method) of four of the transgenic lines were grown to establish T2 homozygous lines. These had ca. 73% C18: 1 and 8 to 9% each of C18: 2 and C18: 3 (alpha-linolenic acid) fractions in comparison to ca. 53% C18: 1, 24% C18: 2 and 16% C18: 3 in the parental line VH486.