Article ID Journal Published Year Pages File Type
2090881 Journal of Microbiological Methods 2008 12 Pages PDF
Abstract

Fungal communities are key components of soil, but the study of their ecological significance is limited by a lack of appropriated methods. For instance, the assessment of fungi occurrence and spatio-temporal variation in soil requires the analysis of a large number of samples. The molecular signature methods provide a useful tool to monitor these microbial communities and can be easily adapted to capillary electrophoresis (CE) allowing high-throughput studies. Here we assess the suitability of CE-FLA (Fragment Length Polymorphism, denaturing conditions) and CE-SSCP (Single-Stranded Conformation Polymorphism, native conditions) applied to environmental studies since they require a short molecular marker and no post-PCR treatments. We amplified the ITS1 region from 22 fungal strains isolated from an alpine ecosystem and from total genomic DNA of alpine and infiltration basin soils. The CE-FLA and CE-SSCP separated 17 and 15 peaks respectively from a mixture of 19 strains. For the alpine soil-metagenomic DNA, the FLA displayed more peaks than the SSCP and the converse result was found for infiltration basin sediments. We concluded that CE-FLA and CE-SSCP of ITS1 region provided complementary information. In order to improve CE-SSCP sensitivity, we tested its resolution according to migration temperature and found 32 °C to be optimal. Because of their simplicity, quickness and reproducibility, we found that these two methods were promising for high-throughput studies of soil fungal communities.

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