An all-in-one visual selection system for male-sterile line production in maize and rice

Heterosis has significantly improved crop yields, yet hybrid seed production remains hindered by labor-intensive manual emasculation. Although current male-sterility systems, such as cytoplasmic male sterility and environment-sensitive genic male sterility, have improved the efficiency of hybrid seed production, their limited genetic adaptability and high environmental dependence remain major challenges. Here, we report an all-in-one seed production technology (ASPT) that integrates CRISPR–Cas9, RUBY, and key seed production technology (SPT) components into a single vector, enabling efficient generation and propagation of male-sterile lines in both maize and rice. The engineered RUBY marker enables visual identification of male-sterile and maintainer lines, with an accuracy of 99.81% in automated seed sorting and 100% in secondary field screening. Notably, ASPT was successfully introduced into 21 genetically diverse elite maize inbred lines, demonstrating broad compatibility. ASPT enables scalable and precise propagation of male-sterile lines in both maize and rice, providing a broadly applicable strategy to advance hybrid seed production in crops.