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Characterization of the fan1 locus in soybean line A5 and development of molecular assays for high-throughput genotyping of FAD3 genes

Kristin Bilyeu
Molecular breeding 2013 v.33 pp. 895-907
alleles, genetic markers, genotyping, heterozygosity, linolenic acid, loci, marker-assisted selection, missense mutation, oil crops, single nucleotide polymorphism, soybean oil, soybeans, stearoyl-CoA desaturase
Soybean is one of the most important oil crops in the world, and reduced linolenic acid content of soybean oil will provide increased stability of the oil to consumers and food manufacturers and limit the amount of trans-fat to be used in the processed foods. The linolenic content in soybean seeds is controlled by three fatty acid desaturase (FAD) 3 enzymes, including FAD3A, B and C, with the FAD3A enzyme having the major effect on the final linolenic acid content in soybean seeds. The widely used soybean lines for breeding in the U.S.with 1% linolenic acid content have mutations in each of the three FAD genes derived from A5 (deletion of FAD3A), A26 and A23 (missense mutations in FAD3B and C, respectively). Although soybean line A5 has been released for 30 years, the extent and definition of the deletion of the FAD3A gene has not been characterized which prevented us from designing robust molecular markers for effective marker-assisted selection. Using a PCR-based genomic strategy, we have identified a 6.5 kbp deletion of the FAD3A gene in A5 and developed a TaqMan detection assay by targeting the deletion junction in A5 which could be used to distinguish the homozygotes and heterozygotes of the gene. In addition, based on mutant SNPs in FAD3B and FAD3C identified in A26 and A23, respectively, we have also developed TaqMan assays for high throughput marker-assisted selection (MAS). The TaqMan assays have proven to be a very effective platform to detect the mutant FAD3 alleles and thus will greatly facilitate the high-throughput MAS for development of soybean lines with reduced linolenic acid content.