Present address: CSIRO Plant Industry, GPO Box 1600, Canberra, ACT 2601, Australia.
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Directed terminal restriction analysis tool (DRAT): an aid to enzyme selection for directed terminal-restriction fragment length polymorphisms
Article first published online: 29 JUL 2011
DOI: 10.1111/j.2041-210X.2011.00139.x
© 2011 The Authors. Methods in Ecology and Evolution © 2011 British Ecological Society
Additional Information
How to Cite
Roberts, D. M., Schofield, P. G., Donn, S. and Daniell, T. J. (2012), Directed terminal restriction analysis tool (DRAT): an aid to enzyme selection for directed terminal-restriction fragment length polymorphisms. Methods in Ecology and Evolution, 3: 24–28. doi: 10.1111/j.2041-210X.2011.00139.x
Correspondence site: http://www.respond2articles.com/MEE/
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Present address: CSIRO Plant Industry, GPO Box 1600, Canberra, ACT 2601, Australia.
Publication History
- Issue published online: 1 FEB 2012
- Article first published online: 29 JUL 2011
- Received 1 February 2011; accepted 16 June 2011Handling Editor: Robert Freckleton
Keywords:
- directed T-RFLP;
- design software;
- diagnostic tool;
- environmental monitoring
Summary
1. T-RFLP is an established tool for high-throughput studies of microbial communities, which can, with care and practical validation, be enhanced to aid identification of specific organisms in a community by associating T-RFs from experimental runs with predicted T-RFs from a set of existing sequences. A barrier to this approach is the laborious process of selecting diagnostic restriction enzyme(s) for further validation.
2. Here, we describe directed terminal restriction analysis tool (DRAT), a software tool that aids the design of directed terminal-restriction fragment length polymorphism (DT-RFLP) strategies, to separate DNA targets based on restriction enzyme polymorphisms. The software assesses multiple user-supplied DNA sequences, ranks optimal restriction endonucleases for separating targets and provides summary information including the length of diagnostic terminal restriction fragments. A worked example suggesting enzymes uniquely separating selected arbuscular mycorrhizal fungal groups is presented.
3. This tool greatly facilitates identification of diagnostic restriction enzymes for user-designated groups within complex populations and provides expected product sizes for all designated groups.