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| Detection of functional genes and their expression in soil ecosystems: nifH a marker for nitrogen-fixing microbial populations |
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Franco Widmer
Molecular Ecology, Swiss Federal Research Station for Agroecology and Agriculture (FAL-Reckenholz), Zurich, Switzerland
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Molecular tools allow to analyze microbial populations in soil without the limitations of cultivation dependent approaches. The molecular ecology approach is based on the extraction of nucleic acids directly from soil samples and on the detection of specific marker genes. Development of robust DNA extraction and PCR protocols and the establishment of phylogenetic markers based on the ribosomal RNA gene represent milestones in the development of this approach.
Analysis of phylogenetic markers poses limits to the detection of functional groups of microorganisms that perform specific functions in an ecosystem. Functional genes, therefore represent attractive marker genes for such purposes. In addition, if RNA is extracted from soil, the expression dynamics of these functional marker genes can be described.
Biological nitrogen fixation is a key ecological function exerted exclusively by some prokaryotes. Therefore this functional group represents an ideal model system for the development of the functional marker gene approach. The specific marker gene nifH, encoding the enzyme nitrogenase reductase, has been established for the specific detection of nitrogen-fixing microorganisms.
We developed specific nifH mRNA detection in soil as a means to measure nifH gene expression in soil. NifH mRNA expression was correlated to nitrogen-fixation activity in a soil microcosm system with Azotobacter vinelandii as diazotrophic model organism. In addition subgroup-specific detection protocols were developed for detection at a higher resolution than allowed by the commonly used universal nifH detection protocols. Our data demonstrate the power and feasibility of detecting functional groups of microorganisms in soil and of analyzing their marker gene expression at the mRNA level.
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References
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WIDMER F., SHAFFER B.T., PORTEOUS L.A., and SEIDLER R.J. (1999).
Analysis of nifH gene pool complexity in soil and litter at a Douglas fir forest site in the Oregon Cascade mountain range.
Appl. Environ. Microbiol. 65:373-380
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SHAFFER B.T., WIDMER F., PORTEOUS L.A., and SEIDLER R.J. (2000).
Temporal and spatial distribution of the nifH gene of N2 fixing bacteria in forests and clear-cuts in Western Oregon.
Microb. Ecol. 39:12-21
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