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Improved PCR-DGGE for high resolution diversity screening of complex sulfate-reducing prokaryotic communities in soils and sediments

Miletto, M. and Bodelier, P.L.E. and Laanbroek, H.J. (2007) Improved PCR-DGGE for high resolution diversity screening of complex sulfate-reducing prokaryotic communities in soils and sediments. Journal of Microbiological Methods, 70, 103-111. ISSN 0167-7012.

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Official URL: http://dx.doi.org/10.1016/j.mimet.2007.03.015

Abstract

In this study we evaluated a high resolution PCR-DGGE strategy for the characterization of complex sulfate-reducing microbial communities inhabiting natural environments. dsrB fragments were amplified with a two-step nested PCR protocol using combinations of primers targeting the dissimilatory (bi)sulfite reductase genes. The PCR-DGGE conditions were initially optimized using a dsrAB clone library obtained from a vegetated intertidal riparian soil along the river Rhine (Rozenburg, the Netherlands). Partial dsrB were successfully amplified from the same environmental DNA extracts used to construct the library, DGGE-separated and directly sequenced. The two approaches were in good agreement: the phylogenetic distribution of clones and DGGE-separated dsrB was comparable, suggesting the presence of sulfate-reducing prokaryotes (SRP) belonging to the families ‘Desulfobacteraceae,' ‘Desulfobulbaceae’ and ‘Syntrophobacteraceae,' and to the Desulfomonile tiedjei- and Desulfobacterium anilini-groups. The nested PCR-DGGE was also used to analyze sediment samples (Appels, Belgium) from a series of microcosms subjected to a tidal flooding regime with water of different salinity, and proved to be a valid tool also to monitor the SRP community variation over time and space as a consequence of environmental changes.

Item Type:Article
Institutes:Nederlands Instituut voor Ecologie (NIOO)
ID Code:4560
Deposited On:16 Sep 2009 02:00
Last Modified:24 Apr 2012 16:48

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