** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** THE USE OF NITROGENOUS FERTILIZER HAS BEEN INCREASING IN AGRICULTURE PRODUCTION WORLDWIDE. IT IS ESTIMATED THAT UP TO 70% OF THE APPLIED NITROGEN IS LOST FROM THE PLANT-SOIL SYSTEM. NITROGEN NOT ABSORBED BY THE PLANTS WILL LIKELY CONTRIBUTE TO SOIL AND WATER POLLUTION THROUGH SURFACE RUN-OFF, DENITRIFICATION, AND VOLATILIZATION. REDUCING NITROGEN INPUT WHILE MAINTAINING CROP PRODUCTIVITY IS CRUCIAL FOR AGRICULTURE PRODUCTION AND ENVIRONMENTAL PROTECTION. PERENNIAL RYEGRASS IS A WIDELY USED TURF, FORAGE, AND PASTURE GRASS SPECIES THAT HAS TREMENDOUS ECONOMIC VALUE. HOWEVER, THIS GRASS SPECIES HAS SHALLOW ROOTS, WHICH CAN RESTRICT NITROGEN UPTAKE AND UTILIZATION, ESPECIALLY UNDER NITROGEN-LIMITING CONDITIONS. THE OVERARCHING RATIONALE OF THE PROPOSED RESEARCH IS TO FILL A CRITICAL RESEARCH GAP IN UNDERSTANDING THE MECHANISMS OF PERENNIAL RYEGRASS RESPONSE TO LOW NITROGEN INPUT. WE WILL INTEGRATE THE PHYSIOLOGICAL, MOLECULAR, AND GENOMIC APPROACHES TO IDENTIFY GENES FOR NITROGEN USE EFFICIENCY. THE SPECIFIC OBJECTIVES ARE TO: 1) CONDUCT A GENOME-WIDE ASSOCIATION STUDY OF LOW NITROGEN RESPONSE OF PERENNIAL RYEGRASS NATURAL POPULATION; 2) EXAMINE THE TRANSCRIPTOME PROFILE IN GENOTYPES WITH CONTRASTING LOW NITROGEN RESPONSE AND NITROGEN USE EFFICIENCY; 3) IDENTIFY KEY GENES FOR NITROGEN USE EFFICIENCY BY INTEGRATING A GENOME-WIDE ASSOCIATION STUDY AND TRANSCRIPTOMIC ANALYSIS, AND 4) VALIDATE THE BIOLOGICAL FUNCTIONS OF KEY GENES USING MUTANTS OF MODEL GRASS SPECIES AND PERENNIAL RYEGRASS. THE LONG-TERM BENEFITS OF THIS PROJECT WILL BE FACILITATING THE DEVELOPMENT OF LOW-NITROGEN TOLERANT PLANT GERMPLASM, PROMOTING SUSTAINABILITY OF PERENNIAL GRASS PRODUCTION, AND IMPROVING THE ENVIRONMENT.