This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Weinbauer, M. G. Articles by Höfle, M. G. Search for Related Content PubMed PubMed Citation Articles by Weinbauer, M. G. Articles by Höfle, M. G. Agricola Articles by Weinbauer, M. G. Articles by Höfle, M. G.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, March 2002, p. 1082-1087, Vol. 68, No. 3
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.3.1082-1087.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Simultaneous Extraction from Bacterioplankton of Total RNA and DNA Suitable for Quantitative Structure and Function Analyses

Department of Environmental Microbiology, AG Microbial Ecology, GBF-German Research Center for Biotechnology, D-38100 Braunschweig, Germany

Received 22 May 2001/ Accepted 10 October 2001

The aim of this study was to develop a protocol for the simultaneous extraction from bacterioplankton of RNA and DNA suitable for quantitative molecular analysis. By using a combined mechanical and chemical extraction method, the highest RNA and DNA yield was obtained with sodium lauryl sarcosinate-phenol or DivoLab-phenol as the extraction mix. The efficiency of extraction of nucleic acids was comparatively high and varied only moderately in gram-negative bacterial isolates and bacterioplankton (RNA, 52 to 66%; DNA, 43 to 61%); significant amounts of nucleic acids were also obtained for a gram-positive bacterial isolate (RNA, 20 to 30%; DNA, 20 to 25%). Reverse transcription-PCR and PCR amplification products of fragments of 16S rRNA and its genes were obtained from all isolates and communities, indicating that the extracted nucleic acids were intact and pure enough for community structure analyses. By using single-strand conformation polymorphism of fragments of 16S rRNA and its gene, community fingerprints were obtained from pond bacterioplankton. mRNA transcripts encoding fragments of the enzyme nitrite reductase gene (nir gene) could be detected in a pond water sample, indicating that the extraction method is also suitable for studying gene expression. The extraction method presented yields nucleic acids that can be used to perform structural and functional studies of bacterioplankton communities from a single sample.