Journal
JOURNAL OF FUNGI
Volume 2, Issue 2, Pages -Publisher
MDPI
DOI: 10.3390/jof2020012
Keywords
soil fungi; direct PCR; barcoding; fungal isolates; yeasts; reference library construction
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Funding
- Austrian Science Fund (FWF) project Growth response of Pinus cembra to experimentally modified soil temperatures at the treeline [P22836-B16]
- Tiroler Wissenschaftsfond project How to open a Treasure Box: Barcoding of the Mycological Collection IB [TWF P718017]
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Fungal pure cultures identified with both classical morphological methods and through barcoding sequences are a basic requirement for reliable reference sequences in public databases. Improved techniques for an accelerated DNA barcode reference library construction will result in considerably improved sequence databases covering a wider taxonomic range. Fast, cheap, and reliable methods for obtaining DNA sequences from fungal isolates are, therefore, a valuable tool for the scientific community. Direct colony PCR was already successfully established for yeasts, but has not been evaluated for a wide range of anamorphic soil fungi up to now, and a direct amplification protocol for hyphomycetes without tissue pre-treatment has not been published so far. Here, we present a colony PCR technique directly from fungal hyphae without previous DNA extraction or other prior manipulation. Seven hundred eighty-eight fungal strains from 48 genera were tested with a success rate of 86%. PCR success varied considerably: DNA of fungi belonging to the genera Cladosporium, Geomyces, Fusarium, and Mortierella could be amplified with high success. DNA of soil-borne yeasts was always successfully amplified. Absidia, Mucor, Trichoderma, and Penicillium isolates had noticeably lower PCR success.
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