Phenotypic information for eight important traits described by Ghamkhar et al. (2011) (flowering time, hardseededness, leaf marks, calyx tube pigmentation, stem hairiness and the isoflavonoids, formononetin, genistein, and biochanin A) was associated with specific regions in the revised high‐density SNP linkage map (Tables S18 and S19) previously described by Hirakawa et al., 2016;. Significant associations were then mapped onto the advanced assembly (Tsub_Refv2.0) to identify possible candidate genes (Figure 6; Table 2; Table 3). The effective anchoring of sequence scaffolds onto the high‐density SNP linkage map in a high‐quality chromosome‐level genome assembly was a major factor in the identification of loci governing key traits. For example, QTLs for leaf marks (LM), calyx pigmentation (CP) and the isoflavonoids [formononetin (FO), genistein (GT) and biochanin A (BCA)] were found to map adjacently to a region on Chr 5. The linkage between these traits was demonstrated by Francis and Millington (1965), who used mutagenesis on cv. Geraldton, which has a high formononetin content, a prominent leaf mark and anthocyanin pigmentation of leaves, calyx tubes, stipules and stems to produce the low formononetin cv. Uniwager, which concomitantly lost its leaf mark and anthocyanin pigmentation. Co‐localization of the QTLs identified for the traits in the present study explains the observation of breeders that isoflavone content is linked to pigmentation traits (leaf and calyx) (Figure 6; Table 3) (Francis and Millington, 1965). This information illustrates links between the new assembly, the high‐density linkage map and key quantitative traits that can assist future marker‐assisted selection and comparative mapping with other species to improve forage legumes and increase livestock productivity.