Mycobiology.
2003 Dec;31(4):196-199.
A Reliable "Direct from Field" PCR Method for Identification of Mycorrhizal Fungi from Associated Roots
- Affiliations
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- 1Institute of Applied Botany/Plant Anatomy and Physiology, Center for Environmental Research and Technology (UFT), University of Bremen, Germany. carsten.harms@uni-bremen.de
- 2Graduate School of Biological Science and Education, Korea National University of Education, Chung-Buk 360-791, Korea.
- 3University of Bremen, Institute of Molecular Biology and Bioanalytics, Leobener Street, 28359 Bremen, Germany.
Abstract
- A very reliable and specific method for the identification of fungi in ectotrophic mycorrhizal symbiosis was developed using a specific PCR assay based on the amplification of the ITS1 region. To obtain specific data, an ITS-diagnostic assay was carried out that reveals genera and species specific sequences. Here, an application of one method is presented, which covers the identification of pure mycelia, basidiocarps as well as mixed samples such as ectomycorrhizal roots that were mingled with remains of the host plant. For this purpose a protocol was established that allowed the extraction of DNA from single mycorrhizal roots. In order to perform a specific ITS analysis we generated a new ITS-primer (ITS8) by a multiple alignment of five different genera and species of mycorrhizal fungi. The utilization of ITS1 and ITS8 resulted in specific PCR amplicons, which were characterized by sequencing without purification steps, even when the template DNA was associated with roots.
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