** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** THIS PROPOSAL IS GOING TO ADDRESS AREA 1: MANAGEMENT PRACTICES TO MINIMIZE ENVIRONMENTAL RISK OF GE ORGANISMS OF THE USDA BRAG STANDARD RESEARCH PROGRAM. A PROBLEM IN TOMATO IS THE SUBSTANTIAL POLLEN-MEDIATED GENE FLOW (PMGF) FROM GENETICALLY ENGINEERED (GE) TOMATO TO NON-GE TOMATO. USING ARABIDOPSIS AS A PROOF-OF-CONCEPT RESEARCH, WE SEEK TO GENETICALLY ENGINEER TOMATO PRODUCING CRISPR POLLEN THAT CAN DESTROYSEEDDEVELOPMENT IN THE NON-GE PLANTS AFTER DOUBLE FERTILIZATION. THE GE LINES WILL BE USED FOR SELFING OR RECIPROCAL CROSSING WITH THE NON-GE ARABIDOPSIS OR TOMATO PLANTS TO TEST THE EFFECT ONSEED ABORTION. THE BEST LINE IN TOMATO WILL BE USED FOR FIELD TRIALS TO TEST PHENOTYPIC STABILITY AND INTERFERENCE WITH SEED PRODUCTION, AND MEASURE THE EXTENT OF PMGF FROM THE GE TO NON-GE PLANTS IN A REAL WORLD SETTING. THE EXPECTED OUTCOME OF THIS PROJECT IS THE ENGINEERED CRISPR POLLEN IN EACH SPECIES, WHICH IN TURN WILL HIGHLY LIMIT PMGF FROM GE PLANTS WITHOUT INTERFERENCE WITH SEED PRODUCTION. IT COULD BE USED AS THE BACKGROUND SOURCE FOR GENETIC ENGINEERING OF ANY TRAITS. THEREFORE, THE RESEARCH DIRECTLY ADDRESSES KEY BARRIERS TO THE DEVELOPMENT OF REGULATORY AND BEST MANAGEMENT PLANS AND SUSTAINABLE PRODUCTION IN TOMATO, WHICH COULD BE APPLICABLE TO OTHER GE CROPS.