Plant variety protection (PVP), or plant breeders’ rights, provides intellectual property protection (IPP) for cultivars. Technical requirements are distinctness, uniformity, and stable (DUS) reproduction. However, field trials are increasingly resource demanding and potentially inconclusive for soybean (Glycine max [L.] Merr.). Our objective was to establish methodologies using molecular markers to facilitate DUS testing while maintaining current IPP levels. We determined that DNA from 10–15 bulked plants represented cultivar genotype. Single nucleotide polymorphism (SNP) data were highly robust in the face of missing and mistyped data; concordances among five laboratories were >.9888. We used SNP, morphological, physiological, and pedigree information to examine 322 publicly available cultivars including 187 with PVPs. Associations among cultivars following multivariate analyses of genetic distances from SNP data and from pedigree kinship data were very similar. A SNP similarity of 98.6% was the maximum at which cultivars also differed for morphological characteristics. Many (38%) cultivar pairs with members >90% SNP similarity expressed different morphologies with SNP similarities ranging 96–98.6%. Of cultivars <96% SNP similar, only a single pair differed by a single morphological difference; all others differed by more than two morphological characteristics. A SNP similarity of 96% between soybean cultivars represents an initial and conservative point of demarcation between cultivars that have morphological differences and those that do not. Chronological monitoring of pedigree–kinship and SNP similarities showed little evidence that a lack of genetic diversity in F2 breeding populations contributed to challenges in DUS among U.S. soybean cultivars.

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