A useful PCR primer set for the ectomycorrhizal fungus Tricholoma matsutake in wild pine rhizosphere based on the nuclear ribosomal DNA IGS2 sequence

Tricholoma matsutake is an edible ectomycorrhizal mushroom that forms a symbiotic association with Pinaceae trees by constructing a large extraradical mycelial area (called a shiro) in the soil. The detection of this fungal mycelium in the soil is crucial for estimating the success of outplanted mycorrhizal seedlings inoculated with T. matsutake under experimental conditions. Although several T. matsutake-specific DNA markers have been reported for efficient detection in the field, no comparative study has been conducted to assess their effectiveness. In the present study, we targeted the nuclear ribosomal DNA intergenic spacer 2 (IGS2) region for the detection of T. matsutake. The newly designed TmSP-I-2F/TmSP-I-2R primer pair, which targets a partial IGS2 sequence (543 bp), effectively detected T. matsutake from pine root and soil samples via PCR assay, outperforming other T. matsutake-specific primers. In combination with a PCR system targeting LTR DNA markers that were previously developed, a PCR system with the TmSP-I-2F/TmSP-I-2R primer pair set can expedite investigations of the dynamics of T. matsutake genets in mycorrhizas and shiro.