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Summary: •Although the molecular phylogeny, evolution and biodiversity of arbuscular mycorrhizal fungi (AMF) are becoming clearer, phylotaxonomically reliable sequence data are still limited. To fill this gap, a data set allowing resolution and environmental tracing across all taxonomic levels is provided.•Two overlapping nuclear DNA regions, totalling c. 3 kb, were analysed: the small subunit (SSU) rRNA gene (up to 1800 bp) and a fragment spanning c. 250 bp of the SSU rDNA, the internal transcribed spacer (ITS) region (c. 475–520 bp) and c. 800 bp of the large subunit (LSU) rRNA gene. Both DNA regions together could be analysed for 35 described species, the SSU rDNA for c. 76 named and 18 as yet undefined species, and the ITS region or LSU rDNA, or a combination of both, for c. 91 named and 16 as yet undefined species.•Present phylogenetic analyses, based on the three rDNA markers, provide reliable and robust resolution from phylum to species level. Altogether, 109 named species and 27 cultures representing as yet undefined species were analysed.•This study provides a reference data set for molecular systematics and environmental community analyses of AMF, including analyses based on deep sequencing. See also the Commentary by Young [ABSTRACT FROM AUTHOR]