Rapid and reliable high-throughput methods of DNA extraction for use in barcoding and molecular systematics of mushrooms |
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Authors: | Dentinger Bryn T M Margaritescu Simona Moncalvo Jean-Marc |
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Affiliation: | Department of Ecology and Evolutionary Biology, 25 Willcocks St., University of Toronto, Toronto, ON, Canada M5S 3B2 Department of Natural History, 100 Queen's Park, Royal Ontario Museum, Toronto, ON, Canada M5S 2C6. |
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Abstract: | We present two methods for DNA extraction from fresh and dried mushrooms that are adaptable to high-throughput sequencing initiatives, such as DNA barcoding. Our results show that these protocols yield ∼85% sequencing success from recently collected materials. Tests with both recent (<2 year) and older (>100 years) specimens reveal that older collections have low success rates and may be an inefficient resource for populating a barcode database. However, our method of extracting DNA from herbarium samples using small amount of tissue is reliable and could be used for important historical specimens. The application of these protocols greatly reduces time, and therefore cost, of generating DNA sequences from mushrooms and other fungi vs. traditional extraction methods. The efficiency of these methods illustrates that standardization and streamlining of sample processing should be shifted from the laboratory to the field. |
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Keywords: | FTA fungi internal transcribed spacer internal transcribed spacer proteinase K rDNA ribosomal RNA Whatman |
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