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Isolation of small-subunit rRNA for stable isotopic characterization

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MacGregor,  B. J.
Department of Molecular Ecology, Max Planck Institute for Marine Microbiology, Max Planck Society;

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Brüchert,  V.
Department of Biogeochemistry, Max Planck Institute for Marine Microbiology, Max Planck Society;

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Amann,  R.
Department of Molecular Ecology, Max Planck Institute for Marine Microbiology, Max Planck Society;

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MacGregor, B. J., Brüchert, V., Fleischer, S., & Amann, R. (2002). Isolation of small-subunit rRNA for stable isotopic characterization. Environmental Microbiology, 4(8), 451-464.


Cite as: http://hdl.handle.net/21.11116/0000-0001-D2DD-9
Abstract
Small-subunit ribosomal RNA (SSU rRNA) has several characteristics making it a good candidate biomarker compound: it is found in bacteria, archaea and eukaryotes; it is quickly degraded extracellularly, hence SSU rRNA extracted from a sample probably derives from the currently active population; it includes both conserved and variable regions, allowing the design of capture probes at various levels of phylogenetic discrimination; and rRNA sequences from uncultured species can be classified by comparison with the large and growing public database. Here we present a method for isolation of specific classes of rRNAs from mixtures of total RNA, employing biotin- labelled oligonucleotide probes and streptavidin-coated paramagnetic beads. We also show that the stable carbon isotope composition of Escherichia coli total RNA and SSU rRNA reflects that of the growth substrate for cells grown on LB, M9 glucose and M9 acetate media. SSU rRNA is therefore a promising biomarker for following the flow of carbon, and potentially nitrogen, in natural microbial populations. Some possible applications are discussed.