Isolation and characterization of the Delta(12)-fatty acid desaturase in peanut (Arachis hypogaea L.) and search for polymorphisms for the high oleate trait in Spanish market-type lines

An understanding of the molecular mechanisms that are responsible for incre ased oleic acid accumulation would open avenues to alter peanut fatty acid composition and allow detection of polymorphic regions which can be used fo r marker assisted selection (MAS). Delta (12)-Fatty acid desaturase (FAD) w as isolated and characterized from genotypes having a low or high oleic to linoleic acid O/L ratio - genotypes, Tamspan 90 (T-90) and F435-2-2 (F435), respectively. Southern blots showed three to four copies per haploid genom e, and no major differences in organization between the two parental lines. Approximately 3525 bp was isolated from both genotypes, including a genomi c walk toward the promoter region. The Delta (12)-Fad contains a putative i ntron, the coding region at the 3' end, and an open reading frame (ORF) of 1140 bp encoding 379 amino acids. Comparisons of the coding sequences from the high and low oleic acid genotypes revealed several single nucleotide po lymorphisms (SNPs). Two polymorphisms appear to be associated with the high O/L trait. The first is an "A" insertion 442 bp after the start codon. The "A" insertion shifts the amino acid reading frame, probably resulting in a truncated, inactive protein and the loss of one of three histidine boxes b elieved to be involved in metal ion complexation required for the reduction of oxygen. Another polymorphism at 448 bp from the start codon results in an amino acid change. The region containing the polymorphisms was amplified from leaf tissue of several independently derived backcross lines (IDBLs). Most high O/L lines had either the "A" insertion or the amino acid substit ution.