Wild barley cytoplasms reduce grain weight plasticity, with environment-dependent cytonuclear epistasis at the

Although cytoplasmic genetic variation influences agronomic traits, its potential for breeding applications remains unexplored. We validated cytoplasmic effects on grain traits in barley and tested their portability across nuclear backgrounds and environments. The cytonuclear multi-parent population validation panel (CMPPV) and Magal reciprocal validation panel (MRVP) were evaluated in replicated field trials across two thermally contrasting Israeli environments. The wild B1K-50-04 cytoplasm consistently increased thousand grain weight (TGW) by 6-11% and grain width (GW) by 2-3% relative to the corresponding cultivated cytoplasm within each reciprocal comparison, explaining up to 9.5% of TGW, and 8% of GW variance. These effects persisted in segregating populations derived from crosses with the elite cultivar Magal, demonstrating portability across nuclear backgrounds. Three-way analysis of marker × cytoplasm × environment interactions across a targeted set of loci identified a chromosome 5 H cluster of significant marker effects, including genotype × cytoplasm × environment signals near the Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-026-01673-6.