Fig. 1. Integrated data workflow of Mutagenesis Directed Breeding Platform (MDBP) toolkit. A‒D, Interfaces of four core modules: Pool Construction generates a standard pooling plan based on field layout (A); Mutation Screening imports the pooling plan to map variants to field plots (B); KASP Primer Design uses mutation results to generate primer design and plate map files (C); and Individual Plant Location uses the design file as a linkage key to trace KASP results back to specific field plants (D). E, Schematic overview of MDBP for rapid screening of target gene mutations in the heavy-ion-mutagenized hybrid rice restorer line R8022. Pooling was performed by combining four experimental plots, each consisting of 96 rice plants (arrayed in 12 rows × 8 plants). DNA was extracted from the mixed pool and subjected to targeted gene sequencing, followed by mutation screening using the Mutation Screening module. For pools where mutations were detected, leaf samples from the corresponding plots were mapped into 96-well plates according to their original field arrangement. Each plate was then genotyped using automatically generated KASP markers, and the results were accurately mapped back to the original field plots and row-plant positions, thereby precisely identifying the individual plants carrying the target gene mutations. KASP, Kompetitive allele-specific PCR; VCF, Variant call format; Mut, Mutant; FAM, 6-Carboxyfluorescein; HEX, Hexachlorofluorescein.