AEM Accepts, published online ahead of print on 10 November 2006

This Article 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 HighWire Citing Articles via Google Scholar Google Scholar Articles by Gao, H. Articles by Zhou, J. Search for Related Content PubMed PubMed Citation Articles by Gao, H. Articles by Zhou, J. Agricola Articles by Gao, H. Articles by Zhou, J.

 Previous Article  |  Next Article 

Appl. Environ. Microbiol. doi:10.1128/AEM.01771-06
Copyright (c) 2006, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Microarray-based Analysis of Microbial Community RNAs by Whole Community RNA Amplification (WCRA)

Haichun Gao, Zamin K. Yang, Terry J. Gentry, Liyou Wu, Christopher W. Schadt, and Jizhong Zhou^*

Environmental Sciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee, Center for Microbial Ecology, Michigan State University, East Lansing, Michigan, Institute for Environmental Genomics and Department of Botany and Microbiology, University of Oklahoma, Norman, Oklahoma, 73019; Department of Soil and Crop Sciences, Texas A&M University, College Station, Texas

^* To whom correspondence should be addressed. Email: jzhou{at}ou.edu.

A new approach, termed whole community RNA amplification (WCRA), was developed to provide sufficient amounts of mRNAs from environmental samples for microarray analysis. The method employs fusion primers (6-9 random nucleotides with an attached T7 promoter) for the first strand synthesis. The shortest primers (T7N6S) gave the best results in terms of yields and representativeness of amplification. About 1200-1800 fold amplifications were obtained with the RNA templates ranging from 10 to 100 ng and very representative detections were obtained with the total RNAs of 50-100 ng. Evaluation with Shewanella oneidensis fur strain revealed that the developed amplification method was able to preserve the original abundance relationships of mRNAs. In addition, to understand whether representative detection of RNAs can be achieved with mixed community samples, amplification biases were evaluated with a mixture of equal quantities of RNAs (100 ng each) from four bacterial species, and representative amplifications were also obtained. Finally, the developed method was applied to the active microbial populations in a denitrifying fluidized bed reactor (FBR) used for denitrification of contaminated groundwaters and ethanol stimulated groundwater samples for uranium reduction. The expressed genes are consistent with the expected functions of the bioreactor and groundwater system, suggesting that this approach is useful in analyzing the functional activities of microbial communities. This is one of the first demonstrations that microarray-based technology can be used to successfully detect the activities of microbial communities from real environmental samples in a high-throughput fashion.

This article has been cited by other articles:

• Liu, Y., Gilchrist, A., Zhang, J., Li, X.-F. (2008). Detection of Viable but Nonculturable Escherichia coli O157:H7 Bacteria in Drinking Water and River Water. Appl. Environ. Microbiol. 74: 1502-1507 [Abstract] [Full Text]