A Restricted Two-Stage Multi-Locus Multi-Allele Genome-Wide Association Study Reveals Genomic Loci and Candidate Genes Controlling Plant-Height-Related Traits in Soybean Under Normal and Shade Conditions

Soybean is an important global crop used for oil, food, and feed production. To increase yield and land-use efficiency, growers often plant soybean at a high density or use intercropping systems. Under these systems, soybeans frequently experience shade stress, which directly affects agronomic traits such as plant height. Although researchers have well documented the genetic basis of plant height under normal conditions, the loci responsible for height variation under shade stress remain largely unexplored. Therefore, we performed a restricted two-stage multi-locus multi-allele genome-wide association study (RTM-GWAS) using SNP linkage disequilibrium block (SNPLDB) markers to identify QTLs associated with soybean plant height under shade stress. We evaluated a natural population of 181 soybean accessions for plant height traits under both normal and shaded conditions across four environments for three years. Using the Soybean40K chip, we derived 11,463 SNPLDB markers and identified 42, 33, and 28 significant SNPLDBs associated with plant height, average internode length, and number of main-stem nodes, respectively. For each SNPLDB, we estimated haplotype (allele) effects and assembled QTL–allele matrices to summarize the population’s genetic composition. Four SNPLDB loci proved stable across multiple environments, exhibiting high −lg(p) values and explaining substantial phenotypic variation. Finally, we projected that 80 candidate genes resided within 180 kb of these stable loci, and we identified four strong candidate genes linked to plant height traits based on combined positional and functional evidence. These results clarify genetic factors that influence soybean height under shading and could aid development of high-yielding soybean varieties.