Multi-Locus GWAS Mapping and Candidate Gene Analysis of Anticancer Peptide Lunasin in Soybean (Glycine max L. Merr.)

Soybean (<i>Glycine max</i>) peptide lunasin exhibits significant cancer-preventive, antioxidant, and hypocholesterolemic effects. This study aimed to identify quantitative trait nucleotides (QTNs) associated with lunasin content and to annotate the candidate genes in the soybean genome. The mapping panel of 144 accessions was gathered from the USDA Soybean Germplasm Collection, encompassing diverse geographical origins and genetic backgrounds, and was genotyped using SoySNP50K iSelect Beadchips. The lunasin content in soybean seeds was measured using the enzyme-linked immunosorbent assay (ELISA) method, with lipid-adjusted soybean flour prepared from seeds obtained from the Germplasm Resource Information Network (GRIN) of USDA-ARS in 2003 and from North Carolina in 2021, respectively. QTNs significantly related to lunasin content in soybean seeds were detected on 15 chromosomes, with LOD scores greater than 3.0, explaining various phenotypic variations identified using the R package mrMLM (v4.0). Significant QTNs on chromosomes 3, 13, 16, 18, and 20 were consistently identified across multiple models as being significantly associated with soybean lunasin content, based on assessment data from two years. Twenty-nine candidate genes were found, with 12 identified in seeds from 2003 and 17 from 2021. Our study is an important effort to understand the genetic basis and functional genes for lunasin production in soybean seeds.