Neufeld, Josh D., Vohra, Jyotsna, Dumont, Marc G., Lueders, Tillmann, Manefield, Mike, Friedrich, Michael W. and Murrell, J. C. (J. Colin). (2007) DNA stable-isotope probing. Nature, 2 (4). pp. 860-866. ISSN 1754-2189

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Abstract

Stable-isotope probing is a method used in microbial ecology that provides a means by which specific functional groups of organisms that incorporate particular substrates are identified without the prerequisite of cultivation. Stable-isotope-labeled carbon (C-13) or nitrogen (N-15) sources are assimilated into microbial biomass of environmental samples. Separation and molecular analysis of labeled nucleic acids ( DNA or RNA) reveals phylogenetic and functional information about the microorganisms responsible for the metabolism of a particular substrate. Here, we highlight general guidelines for incubating environmental samples with labeled substrate and provide a detailed protocol for separating labeled DNA from unlabeled community DNA. The protocol includes a modification of existing published methods, which maximizes the recovery of labeled DNA from CsCl gradients. The separation of DNA and retrieval of unlabeled and labeled fractions can be performed in 4-5 days, with much of the time being committed to the ultracentrifugation step.

Item Type:	Journal Article
Subjects:	Q Science > QD Chemistry
Journal or Publication Title:	Nature
Publisher:	NATURE PUBLISHING GROUP
ISSN:	1754-2189
Date:	2007
Volume:	2
Number:	4
Number of Pages:	7
Page Range:	pp. 860-866
Identification Number:	10.1038/nprot.2007.109
Publication Status:	Published
URI:	http://wrap.warwick.ac.uk/id/eprint/30610

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