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Cyanobacteria are a key constituent of biocrusts, communities dominated by lichens, mosses and associated microorganisms, which are prevalent in drylands worldwide and that largely determine their functioning. Despite their importance, there are large gaps in our knowledge of the composition and diversity of cyanobacteria associated with biocrusts, particularly in areas such as the Mediterranean Basin. We studied the diversity of these cyanobacteria in a gypsiferous grassland from Central Spain using both morphological identification after cultivation and genetic analyses with the 16 S rRNA gene. Nine different morphotypes were observed, eight corresponding to filamentous, and one to unicellular cyanobacteria. We found cyanobacterial genera typical of biocrust communities, such as Microcoleus and Trichocoleus, and N-fixing cyanobacteria such as Scytonema and Nostoc. Genetic information allowed us to identify cultures belonging to recently described genera such as Roholtiella, Nodosilinea and Mojavia. We also describe two new phylotypes of Microcoleus and Scytonema, which are key genera contributing to ecosystem functioning in biocrust-dominated ecosystems worldwide.