Lynch, M. and R.G. Thorn
Presented at the All Scientist Meeting (2003-09-12 )
A molecular approach was used to analyze the composition of communities of soil basidiomycetes in Kellogg Biological Station LTER agroecosystems that differ in tillage history. This approach combined soil DNA extraction using a FastPrep method modified to increase recovery of fungal DNA, PCR amplification using basidiomycete-specific primers, cloning and RFLP screening of mixed PCR products, and sequencing of unique PCR products. These fungi, primarily responsible for lignocellulose degradation in plant litter, are not easily cultured from soil and as such have been greatly under-represented in surveys of soil fungi. 10 g subsamples from soil cores were washed through sieves of 250 and 53 μm mesh to remove bacteria and most fungal spores. DNA was extracted from washed organics retained on the 53 μm mesh sieve and basidiomycete-specific PCR primers yielding 2.4 kb amplicons that span the nuclear ribosomal ITS region and approximately 1000 bp into both the 18S and 25S genes were used to obtain templates for cloning. 12 randomly selected transformed colonies were subjected to RFLP screening and unique transformants then sequenced and subjected to phylogenetic analysis. Preliminary results show higher rates of amplification from historically tilled (old field succession; 68%) and never tilled sites (83%) compared to conventional tillage (38%). Similarly, the number of species per sample was greater in no till and never tilled or historically tilled regimes than in conventional tillage. Phylogenetic analysis was performed to determine community composition and to evaluate the diversity of species within and between treatments. This molecular approach should facilitate investigations in soil fungal diversity by avoiding issues surrounding traditional culture methods and provide insight to the community structure of soil basidiomycetes, and their sensitivity to disturbance by tillage.
Back to meeting | Show |