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A Bioinformatics Pipeline for Sequence-Based Analyses of Fungal Biodiversity

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Fungal Genomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 722))

Abstract

The internal transcribed spacer (ITS) is the locus of choice with which to characterize fungal diversity in environmental samples. However, methods to analyze ITS datasets have lagged behind the capacity to generate large amounts of sequence information. Here, we describe our bioinformatics pipeline to process large fungal ITS sequence datasets, from raw chromatograms to a spreadsheet of operational taxonomic unit (OTU) abundances across samples. Steps include assembling of reads originating from one clone, identifying primer “barcodes” or “tags,” trimming vectors and primers, marking low-quality base calls and removing low-quality sequences, orienting sequences, extracting the ITS region from longer amplicons, and grouping sequences into OTUs. We expect that the principles and tools presented here are relevant to datasets arising from ever-evolving new technologies.

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Acknowledgments

We thank James Long for writing several of the original pipeline scripts and Dan Cardin for writing the tag-finder script. Niall Lennon and Chad Nusbaum of the Broad Institute, MA, spearheaded high-throughput Sanger sequencing of our fungal clone libraries. Lab members Michael Booth, Robert Burgess, Ian Herriott, Jack McFarland, and Ina Timling have assisted with testing and improving our pipeline and also provided valuable comments on earlier drafts of the chapter. This work was supported in part by the National Science Foundation under grant numbers EF-0333308 and ARC-0632332. Any opinions, findings and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect those of the National Science Foundation. This publication was also made possible by grant number 2P20RR016466 from the National Center for Research Resources (NCRR), a component of the National Institutes of Health (NIH).

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Authors and Affiliations

  1. Institute of Arctic Biology, University of Alaska, Fairbanks, AK, USA

    D. Lee Taylor

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  1. D. Lee Taylor
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  2. Shawn Houston
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Correspondence to D. Lee Taylor .

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Editors and Affiliations

  1. , Dept. of Botany and Plant Pathology, Purdue University, West State Street 915, West Lafayette, 47907, Indiana, USA

    Jin-Rong Xu

  2. Division of Agriculture, Department of Plant Pathology, University of Arkansas, North Campus Drive 495, Fayetteville, 72701, Arkansas, USA

    Burton H. Bluhm

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Taylor, D.L., Houston, S. (2011). A Bioinformatics Pipeline for Sequence-Based Analyses of Fungal Biodiversity. In: Xu, JR., Bluhm, B. (eds) Fungal Genomics. Methods in Molecular Biology, vol 722. Humana Press. https://doi.org/10.1007/978-1-61779-040-9_10

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  • DOI: https://doi.org/10.1007/978-1-61779-040-9_10

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  • Print ISBN: 978-1-61779-039-3

  • Online ISBN: 978-1-61779-040-9

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