This sequence dataset (FASTQ format) was obtained by metabarcoding an artificial (mock) fungal community. The ribosomal internal transcribed spacer region 1 (ITS1) was amplified with the ITS1F / ITS2 primer pair and sequenced on an Illumina MiSeq platform (v3 chemistry, 2x250 bp). The mock community was composed of an equimolar mixture of the DNA of 189 pure fungal strains isolated from soils, sporocarps or plant tissues. All the strains belonged to the subkingdom Dikarya: 87 Ascomycota strains, 99 Basidiomycota strains and 3 Mucoromycota strains. The mock community was replicated three times (replicates DikA, DikB, DikC) and the three replicates were sequenced independently. The six files correspond to the forward (R1) and reverse (R2) sequences of each replicate. The sequencing was funded by the INRA MEM metaprogramme (Meta-Omics of Microbial Ecosystems, MetaBAR project). This sequence dataset was used to compare several bioinformatic approaches (Pauvert et al. Bioinformatics matters: the accuracy of plant and soil community data is highly dependent on the metabarcoding pipeline. Submitted).