QTL mapping for hundred-seed weight in soybean using high-density SLAF-seq technology

Hundred-seed weight is a critical yield component and quality trait in soybean that directly influences grain yield potential and market value. To dissect the genetic architecture underlying seed weight variation, we conducted comprehensive quantitative trait loci (QTL) mapping using a recombinant inbred line (RIL) population of 325 F₂:₅ lines derived from a cross between Qihuang 34 (high seed weight parent, mean 23.2 g) and Dongsheng 16 (low seed weight parent, mean 18.6 g). High-density genotyping was performed using specific-locus amplified fragment sequencing (SLAF-seq), generating 2,841 polymorphic markers distributed across 20 linkage groups spanning 2,387.6 cM with an average marker density of 0.84 cM. Multi-environment field trials were conducted in Harbin (2022-2023) and Sanya (2023/2024) to evaluate hundred-seed weight across contrasting environmental conditions. QTL analysis identified 11 significant QTLs distributed across 8 chromosomes (4, 6, 7, 13, 15, 18, 19, and 20), with individual QTLs explaining 2.47-8.59% of phenotypic variance, modest but typical effect sizes for polygenic seed weight traits that collectively account for 62% of total phenotypic variation. The major QTL qSW19-1 on chromosome 19 was detected in 2024 with the highest significance (LOD = 9.72, PVE = 8.59%), representing a novel locus in an infrequently reported chromosomal region for seed weight. Two QTLs (qSW6-1 and qSW7-1) demonstrated cross-year stability, indicating reliable targets for marker-assisted selection. Broad-sense heritability for seed weight was estimated at 0.68, indicating substantial genetic control with moderate genotype × environment interactions. Candidate gene analysis within major QTL regions identified Glyma.19G195400 (cell wall invertase) as the most promising candidate, showing 3.8-fold higher expression in the high seed weight parent during seed filling and strong correlation with phenotype (r = 0.68). Additional validated candidates include Glyma.19G194300 (PEBP family protein), Glyma.19G193400 (bZIP transcription factor), and Glyma.06G095100 (Myb DNA-binding protein), suggesting complex regulatory networks control seed weight. qRT-PCR validation identified four genes with differential expression between contrasting parents, including a cell wall invertase gene with a 3.2-fold difference in expression. These findings expand the genetic landscape of soybean seed weight control, provide molecular markers for breeding applications, and identify functionally validated candidate genes for future mechanistic studies.