A highly screening system at seedling-stage for genic male sterility in pepper hybrid breeding
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In pepper ( Capsicum annuum L.), utilizing heterosis is a highly effective strategy to boost yield. While GMS lines offer advantages for seed production by avoiding the limitations associated with cytoplasmic male sterile (CMS) lines, the use of GMS typically requires the labor-intensive removal of 50% of fertile plants during seed production. To address this challenge, we identified a anthocyanidin-absent (aa) mutant, with a green hypocotyl at seedling stage and yellow anthers.We further identified dihydroflavonol 4-reductase gene ( CaDFR) as a key gene regulating anthocyanin biosynthesis in pepper based on gene fine localization and VIGS experiments. Genetic segregation analysis revealed that CaDFR and CaDYT1 (a genic male sterile gene) had a closely linkage relationship. Building on this discovery, we developed an efficient hybrid seed production strategy using the green hypocotyl morphological marker from the aa mutant and the male sterile line gms1 ( CaDYT1 locus). In conclusion, we successfully cloned CaDFR , a key gene controlling hypocotyl and anther color in pepper. Additionally, we further proposed an efficient seed propagation strategy to accelerate hybrid seed production in the utilization of heterosis. This study not only deepens mechanistic insights into the genetic regulation of pepper pigmentation, but also establishes a practical framework for optimizing hybrid breeding protocols, thereby contributing to simplifying the pepper breeding process and sustainable cultivation.