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PMID: 14602639 Published · ppublish English Comparative Study Evaluation Study Journal Article Research Support, U.S. Gov't, Non-P.H.S.

Web-based phylogenetic assignment tool for analysis of terminal restriction fragment length polymorphism profiles of microbial communities.

Applied and environmental microbiology ·Vol. 69 ·No. 11 ·2003-11-00 ·Pages 6768-76

Kent AD, Smith DJ, Benson BJ, Triplett EW

Abstract

Culture-independent DNA fingerprints are commonly used to assess the diversity of a microbial community. However, relating species composition to community profiles produced by community fingerprint methods is not straightforward. Terminal restriction fragment length polymorphism (T-RFLP) is a community fingerprint method in which phylogenetic assignments may be inferred from the terminal restriction fragment (T-RF) sizes through the use of web-based resources that predict T-RF sizes for known bacteria. The process quickly becomes computationally intensive due to the need to analyze profiles produced by multiple restriction digests and the complexity of profiles generated by natural microbial communities. A web-based tool is described here that rapidly generates phylogenetic assignments from submitted community T-RFLP profiles based on a database of fragments produced by known 16S rRNA gene sequences. Users have the option of submitting a customized database generated from unpublished sequences or from a gene other than the 16S rRNA gene. This phylogenetic assignment tool allows users to employ T-RFLP to simultaneously analyze microbial community diversity and species composition. An analysis of the variability of bacterial species composition throughout the water column in a humic lake was carried out to demonstrate the functionality of the phylogenetic assignment tool. This method was validated by comparing the results generated by this program with results from a 16S rRNA gene clone library.

MeSH Terms
Bacteria/classification,genetics Computational Biology/methods DNA, Bacterial/analysis,genetics,isolation & purification Ecosystem Fresh Water/microbiology Gene Library Internet Phylogeny Polymorphism, Restriction Fragment Length RNA, Ribosomal, 16S/genetics
Chemicals
DNA, Bacterial RNA, Ribosomal, 16S
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Kent Angela D
Microbial Observatory of the North Temperate Lakes Long-Term Ecological Research Site (NTL-LTER), Center for Limnology, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
Smith Dan J
Benson Barbara J
Triplett Eric W
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Article Info
Journal
Applied and environmental microbiology
Abbr.
Appl Environ Microbiol
ISSN
0099-2240
DOI
10.1128/AEM.69.11.6768-6776.2003
Published
2003-11-00
Pages
6768-76
Language
English
Region
United States
NLM ID
7605801
PMCID
PMC262325
Subset
IM
External Links
PubMed source DOI: 10.1128/AEM.69.11.6768-6776.2003 PMC Full Text View journal details
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