Single step molecular characterization of morphologically similar black truffle species

Species-specific internal ITS primers that amplify polymerase chain reaction (PCR) products of different lengths were selected to distinguish the morphologically similar ectomycorrhizal fungi T. melanosporum, T. brumale and T. indicum by aligning their internal transcribed spacer sequences and taking into account any incidence of intraspecific variability. In multiplex PCR experiments, the species-specific primers yielded the expected amplicons on template DNA isolated from the above mentioned species: while there was no amplification in PCR reactions carried our on fungal DNA from competing truffle species and host plants.