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Applied and Environmental Microbiology, October 2005, p. 6193-6198, Vol. 71, No. 10
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.10.6193-6198.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Phylum- and Class-Specific PCR Primers for General Microbial Community Analysis

Christopher B. Blackwood,* Adam Oaks, and Jeffrey S. Buyer

Sustainable Agricultural Systems Laboratory, USDA-ARS, Beltsville Agricultural Research Center, Bldg. 001 Rm. 140, Beltsville, Maryland 20705

Received 31 January 2005/ Accepted 6 June 2005

Amplification of a particular DNA fragment from a mixture of organisms by PCR is a common first step in methods of examining microbial community structure. The use of group-specific primers in community DNA profiling applications can provide enhanced sensitivity and phylogenetic detail compared to domain-specific primers. Other uses for group-specific primers include quantitative PCR and library screening. The purpose of the present study was to develop several primer sets targeting commonly occurring and important groups. Primers specific for the 16S ribosomal sequences of Alphaproteobacteria, Betaproteobacteria, Bacilli, Actinobacteria, and Planctomycetes and for parts of both the 18S ribosomal sequence and the internal transcribed spacer region of Basidiomycota were examined. Primers were tested by comparison to sequences in the ARB 2003 database, and chosen primers were further tested by cloning and sequencing from soil community DNA. Eighty-five to 100% of the sequences obtained from clone libraries were found to be placed with the groups intended as targets, demonstrating the specificity of the primers under field conditions. It will be important to reevaluate primers over time because of the continual growth of sequence databases and revision of microbial taxonomy.

* Corresponding author. Present address: School of Natural Resources and the Environment, University of Michigan, 440 Church St., Ann Arbor, MI 48109-1041. Phone: (734) 647-0481. Fax: (734) 936-2195. E-mail: cbwood{at}umich.edu.

Applied and Environmental Microbiology, October 2005, p. 6193-6198, Vol. 71, No. 10
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.10.6193-6198.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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