Isoflavone is a secondary metabolite of the soybean phenylpropyl
biosynthesis pathway with physiological activity and is beneficial
to human health. In this study, the isoflavone content of 205 soybean
germplasm resources from 3 locations in 2020 showed wide phenotypic
variation. A joint genome-wide association study (GWAS) and weighted
gene coexpression network analysis (WGCNA) identified 33 single-nucleotide
polymorphisms and 11 key genes associated with soybean isoflavone
content. Gene ontology enrichment analysis, gene coexpression, and
haplotype analysis revealed natural variations in the Glyma.12G109800 (GmOMT7) gene and promoter
region, with Hap1 being the elite haplotype. Transient overexpression
and knockout of GmOMT7 increased and decreased the
isoflavone content, respectively, in hairy roots. The combination
of GWAS and WGCNA effectively revealed the genetic basis of soybean
isoflavone and identified potential genes affecting isoflavone synthesis
and accumulation in soybean, providing a valuable basis for the functional
study of soybean isoflavone.