Abstract:

Mycelia were cultivated from a fruiting body of puffballs which were collected from the Eucalyptus grandis forest, and the genomic DNA was extracted as template. Subsequently, the internal transcribed spacer (ITS) region in rDNA gene was amplified, cloned and sequenced. The phylogenetic tree of puffballs was first built based on different part of ITS region. The results showed that six species of puffballs were cultivated successfully using a PDA medium after comparing the sequences of puffballs fruiting body and mycelium ITS regions. The rDNA-ITS region of different puffballs was 607 bp to 766 bp in length. Analysis of phylogenesis revealed that the Scleroderma had close genetic relationships with Pisolithus, Calvatia, Lycoperdon and Vascellum. The three undetermined species, Scleroderma sp.11-1, Scleroderma sp.2-2 and Scleroderma sp.5-2, could probably be S. aurantium. The sequence of ITS1-5.8S rDNA-ITS2 can be used as an evolutionary marker between different genus, and ITS2 as an evolutionary marker in genus. Thus, this work laid a foundation for studying the ectotrophic mycorrhiza, the mechanisms of action, the classification of puffballs and its exploitation and utilization.

Key words: Eucalyptus grandis forest, puffballs, phylogenetic relationships, ITS region, Sichuan Province