Widespread use of legume green manures is limited by a lack of reliable varieties that meet the needs of farmers. Improving symbiotic N fixation (SNF) in open‐pollinated legume species poses a challenge because normal methods used for assessing SNF conflict with breeding practices such as seed production and removing low‐quality sires. We investigated methods for measuring symbiotic N fixation that are compatible with breeding practice for two major legume cover crops: hairy vetch (Vicia villosa Roth) and crimson clover (Trifolium incarnatum L.). We collected two nondestructive samples at the early‐flowering stage (Meristem and Wedge) and a sample at seed harvest (Seed). Our objective was to determine which combination of measurements best predicted symbiotic N fixation (% N derived from atmosphere [Ndfa]) and total N content at 50% flowering. Our statistical approach accommodated the problem that target traits and seed measurements cannot be measured in the same individuals. Overall, the Wedge sample provided the best prediction for Ndfa at 50% flowering. The best predictions for total N plant–1 came from combining vigor ratings with aboveground biomass at seed harvest. In addition to specific recommendations for legume cover crop breeding programs, we provide a flexible statistical method for studying relationships between traits that cannot be measured jointly.

Grafting is a widely used agricultural technique that involves the physical joining of separate plant parts so they form a unified vascular system, enabling beneficial traits from independent genotypes to be captured in a single plant. This simple, yet powerful tool has been used for thousands of years to improve abiotic and biotic stress tolerance, enhance yield, and alter plant architecture in diverse crop systems. Despite the global importance and ancient history of grafting, our understanding of the fundamental biological processes that make this technique successful remains limited, making it difficult to efficiently expand on new genotypic graft combinations. One of the key determinants of successful grafting is the formation of the graft junction, an anatomically unique region where xylem and phloem strands connect between newly joined plant parts to form a unified vascular system. Here, we use an integrated imaging approach to establish a spatiotemporal framework for graft junction formation in the model crop Solanum lycopersicum (tomato), a plant that is commonly grafted worldwide to boost yield and improve abiotic and biotic stress resistance. By combining Positron Emission Tomography (PET), a technique that enables the spatio-temporal tracking of radiolabeled molecules, with high-resolution laser scanning confocal microscopy (LSCM), we are able to merge detailed, anatomical differentiation of the graft junction with a quantitative timeline for when xylem and phloem connections are functionally re-established. In this timeline, we identify a 72-hour window when anatomically connected xylem and phloem strands regain functional capacity, with phloem restoration typically preceding xylem restoration by about 24-hours. Furthermore, we identify heterogeneity in this developmental and physiological timeline that corresponds with microvariability in the physical contact between newly joined rootstock-scion tissues. Our integration of PET and confocal imaging technologies provides a spatio-temporal timeline that will enable future investigations into cellular and tissue patterning events that underlie successful versus failed vascular restoration across the graft junction.