Detailed study of strata associated with the glauconite-rich Floyds Knob Bed in the western Appalachian and eastern Illinois basins have corroborated previous interpretations that the unit is a widespread, largely synchronous marker horizon. However, in some areas there are multiple glauconite beds; in others a distinct bed is lacking, but the glauconite is dispersed throughout many beds, forming an interval rather than a distinct bed. In Kentucky and adjacent states, the Floyds Knob interval, in upper parts of the Lower-Middle Mississippian Borden-Grainger delta sequence and in lower parts of the Fort Payne carbonate sequence, was deposited at the end of loading-type relaxation during a flexural cycle in the Neoacadian (final) tectophase of the Acadian Orogeny. Tectonic influence, combined with a major late Osagean sea-level lowstand, created conditions that generated sediment starvation and shallower seas across widespread parts of the western Appalachian and eastern Illinois basins. In the absence of major sediment influx, glauconite was deposited uniformly across many major depositional settings, ranging from delta-platform to basinal environments. Especially important, however, is the newly reported occurrence of the Floyds Knob interval in basinal Fort Payne environments from south-central Kentucky, where it is represented by a thick, pelletal, glauconite-rich horizon that separates clastics at the base of the Fort. Payne Formation from carbonates at top. The study also provides the first-ever radiometric dating of the Floyds Knob glauconites, which suggests a late Osagean origin. These results support the existing biostratigraphic studies that point to a late Osagean origin for the Floyds Knob interval.