Using terminal restriction fragment length polymorphism (T-RFLP) to identify mycorrhizal fungi: a methods review
- 1.7k Downloads
Terminal restriction fragment length polymorphism (T-RFLP) is an increasingly widely used technique in mycorrhizal ecology. In this paper, we review the technique as it is used to identify species of mycorrhizal fungi and distinguish two different versions of the technique: peak-profile T-RFLP (the original version) and database T-RFLP. We define database T-RFLP as the use of T-RFLP to identify individual species within samples by comparison of unknown data with a database of known T-RFLP patterns. This application of T-RFLP avoids some of the pitfalls of peak-profile T-RFLP and allows T-RFLP to be applied to polyphyletic functional groups such as ectomycorrhizal fungi. The identification of species using database T-RFLP is subject to several sources of potential error, including (1) random erroneous matches of peaks to species, (2) shared T-RFLP profiles across species, and (3) multiple T-RFLP profiles within a species. A mathematical approximation of the risk of the first type of error as a function of experimental parameters is discussed. Although potentially less accurate than some other methods such as clone libraries, the high throughput of database T-RFLP permits much greater replication and may, therefore, be preferable for many ecological questions, particularly when combined with other techniques such as cloning.
KeywordsArbuscular mycorrhiza Ectomycorrhiza Molecular identification Terminal restriction fragment length polymorphism (T-RFLP)
Valuable discussions and sharing of data and concepts with P. Avis, S. Branco, I. Edwards, G. Forrester, D. Park, R. Smissen, and many others have contributed to this review. R. Molina and two reviewers provided helpful comments. The authors were supported by funds from the Foundation for Research, Science and Technology (FRST, NSOF) of New Zealand.
- Artursson V, Finlay RD, Jansson JK (2005) Combined bromodeoxyuridine immunocapture and terminal-restriction fragment length polymorphism analysis highlights differences in the active soil bacterial metagenome due to Glomus mosseae inoculation or plant species. Environ Microbiol 7:1952–1966CrossRefPubMedGoogle Scholar
- FitzJohn RG, Dickie IA (2007) TRAMPR: an R package for analysis and matching of terminal-restriction fragment length polymorphism (TRFLP) profiles. Mol Ecol Notes (in press). DOI 10.1111/j.1471-8286.2007.01744.x
- Klamer M, Roberts MS, Levine LH, Drake BG, Garland JL (2002) Influence of elevated CO2 on the fungal community in a coastal scrub oak forest soil investigated with terminal-restriction fragment length polymorphism analysis. Appl Environ Microbiol 68:4370–4376CrossRefPubMedPubMedCentralGoogle Scholar
- R Development Core Team (2006) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. www.R-project.orgGoogle Scholar