High-throughput and functional SNP detection assays for oleic and linolenic acids in soybean

Soybean is a primary source of vegetable oil, accounting for 53 % of the total vegetable oil consumption in the USA in 2013. Soybean oil with high oleic acid and low linolenic acid content is desired, because it not only improves the oxidative stability of the oil, but also reduces the amount of undesirable trans fat production by the hydrogenation processing. The mutant FAD2-1A and FAD2-1B alleles contribute to an elevated oleic acid level, while mutations in FAD3A and FAD3C lead to a reduction in linolenic acid levels of soybean seed oil. Although single nucleotide polymorphism (SNP) SimpleProbe assays have been developed for these genes, breeder-friendly and high-throughput marker systems are needed for large-scale selections in soybean breeding programs. TaqMan or Kompetitive Allele Specific PCR assays were successfully developed to detect and discriminate mutant alleles of FAD2-1A (17D and PI 603452 sources) and FAD2-1B (PI 283327 source) for the high oleic acid trait, as well as those of FAD3A (CX1512-44 and C1640 sources) and FAD3C (CX1512-44 source) for low linolenic acid content soybean. The accuracy of the assays was validated using multiple populations as well as a panel of diverse soybean lines, and a perfect association was observed between the SNP alleles and their corresponding fatty acid phenotypes. These marker assays have been proven to be robust and reliable for detecting mutant and wild-type alleles and thus will assist forward and backcrossing breeding of soybean lines with desired oil profiles in an accurate and efficient manner.