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Combining nested PCR and restriction digest of the internal transcribed spacer region to characterize arbuscular mycorrhizal fungi on roots from the field

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Abstract

Identification of arbuscular mycorrhizal fungi (AMF) on roots is almost impossible with morphological methods and, due to the presence of contaminating fungi, it is also difficult with molecular biological techniques. To allow broad investigation of the population structure of AMF in the field, we have established a new method to selectively amplify the internal transcribed spacer (ITS) region of most AMF with a unique primer set. Based on available sequences of the rDNA, one primer pair specific for AMF and a few other fungal groups was designed and combined in a nested PCR with the already established primer pair ITS5/ITS4. Amplification from contaminating organisms was reduced by an AluI restriction after the first reaction of the nested PCR. The method was assessed at five different field sites representing different types of habitats. Members of all major groups within the Glomeromycota (except Archaeosporaceae) were detected at the different sites. Gigasporaceae also proved detectable with the method based on cultivated strains.

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Acknowledgements

This work was kindly supported by a grant from the Deutsche Forschungsgemeinschaft (GRK 266/1–96). We wish to thank the group of Dr. M. Vosátka (Průhonice), especially M. Janoušková and Z. Sýkorová for supplying culture material of Glomus intraradices and Gigaspora rosea. Many thanks to Boris Börstler and Cristina Popescu for their assistance in the field and the laboratory and also many thanks to Prof. K. Turnau, Dr. V. Temperton, H. Renker and the anonymous reviewers for critically reading and improving the manuscript.

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Authors and Affiliations

  1. Department of Environmental Sciences, Institute of Ecology, University of Jena, Dornburger Str. 159, 07743 , Jena, Germany

    Carsten Renker, Michael Kaldorf & François Buscot

  2. Department of Systematic Botany, Albrecht von Haller Institute of Plant Sciences, University of Göttingen, Untere Karspüle 2, 37073 , Göttingen, Germany

    Jochen Heinrichs

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  1. Carsten Renker
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  2. Jochen Heinrichs
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  3. Michael Kaldorf
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  4. François Buscot
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Correspondence to François Buscot.

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Renker, C., Heinrichs, J., Kaldorf, M. et al. Combining nested PCR and restriction digest of the internal transcribed spacer region to characterize arbuscular mycorrhizal fungi on roots from the field. Mycorrhiza 13, 191–198 (2003). https://doi.org/10.1007/s00572-002-0214-5

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  • DOI: https://doi.org/10.1007/s00572-002-0214-5

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