Abstract
Polyporus umbellatus is one of the most widely used and precious medicinal fungi and the underground sclerotia are known to be with great medicinal value. However, the molecular mechanisms involved in sclerotial development are poorly understood. In the present study, we constructed a forward suppression subtractive hybridization (SSH) cDNA library of Polyporus umbellatus to identify genes expressing differently between mycelium and sclerotia. In this library, a total of 1202 clones were sequenced, assembled into 222 contigs and 524 singletons which were further searched against the NCBI nonredundant (NR) protein database (E-value cutoff, 10−5). Based on sequence similarity with known proteins, 378 sequences between mycelium and sclerotial were identified and classified into different functional categories through Gene Ontology (GO), Clusters of orthologous Groups of proteins (COGs). We have finally identified a majority of differentially expressed genes (constituting 5.6% of the present library) between the two different periods. An expression level of 32 selected expressed sequence tags (ESTs) generated from the above SSH cDNA library was studied through RT-PCR. This study provides the first global overview of genes putatively involved in Polyporus umbellatus sclerotial development and provides a preliminary basis for further functional research in terms of regulated gene expression in sclerotial production.
Highlights
Polyporus umbellatus, known as Grifola umbellata, belongs to Polyporaceae in the class of Basidiomycetes and as a species of wood-rotting fungi [1]
Snowwhite interwoven hyphae with abundant aerial mycelia appeared at day 60 of cultivation (Figure 1b) which indicated that the P. umbellatus sclerotial were successful induced in the artificial conditions
We reported a combination of suppression subtractive hybridization (SSH) library and real-time qPCR techniques to determine gene expression patterns during sclerotium production
Summary
Known as Grifola umbellata, belongs to Polyporaceae in the class of Basidiomycetes and as a species of wood-rotting fungi [1]. We have demonstrated that the split-plate culture method was an effective way to induce P. umbellatus sclerotial production [18] and Ca2+ signal transduction was found to play an important role in P. umbellatus sclerotial formation [19] These attempts have opened up the possibility to understand the explicit factors affecting P. umbellatus sclerotial development. Identifying the molecular factors affecting sclerotial development and determining their roles will be beneficial for protecting the wild P. umbellatus sclerotial resources. Expressed genes were examined by sequencing, analyzing expressed sequence tags (ESTs) present in the library and quantitative real-time polymerase chain reaction (qRT-PCR) analysis These new EST resources are an important addition to publically available resource especially in relation to the study of sclerotial development
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