Evaluation of hypotheses that relate environmental to evolutionary change across the Ediacaran-Cambrian transition has been hampered by a dearth of sections that preserve both the last appearance of Ediacaran body fossils and the first appearance of Treptichnus pedum within carbonate-rich strata suitable for chemostratigraphic studies. Here, we report two new exceptionally preserved latest Ediacaran fossil assemblages from the Deep Spring Formation at Mount Dunfee, Nevada (USA). Further, we report these occurrences in a high-resolution carbon isotope chemostratigraphic framework, permitting correlation on a regional and global scale. The lower of the two horizons, at the base of the Deep Spring Formation, hosts a body fossil assemblage that includes Gaojiashania, other vermiform body fossils, and possible Wutubus annularis interbedded with Cloudina shell beds. The upper of the two fossil horizons, in the Esmeralda Member of the Deep Spring Formation, contains Conotubus and occurs within the basal Cambrian negative carbon isotope excursion, establishing it as the youngest Ediacaran fossil assemblage discovered to date. This is the first report of Gaojiashania, Conotubus, and Wutubus in Laurentia, extending the known stratigraphic ranges and biogeographic distributions of these taxa to a global scale. These data refine the relative ages of defining characteristics of the Ediacaran-Cambrian boundary and confirm that a large perturbation to the carbon cycle and surface ocean conditions coincided with the extinction of Ediacaran organisms.