This paper aims, with a botanical transect, to contribute to define the boundary between the Mediterranean biogeographical area and the flora of the Alpine domain. The transect, realized in 2018, is located in the Maritimes Alps, in the Roya Valley at the northernmost limit of the Mediterranean in contact with a mountain ecoregion. Our objective was to get as close as possible to the integral flora (Tracheophytes), that counts a corpus of 529 species. A 40 km long transect divided in 150 sampling sites led to 7333 observations of presence, analyzed by Factorial Composant Analysis (FCA) and frequency analysis. Statistics revealed a significant distinction of two floras with a clear spatial boundary not crossed by 76% of the species. The Transect revealed a clear biogeographic limit of biogeographical area, which seems to converge well with the limit of the Mediterranean Hotspot as defined by Myers (2000).