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Five simple guidelines for establishing basic authenticity and reliability of newly generated fungal ITS sequences

Journal article
Authors R. Henrik Nilsson
Leho Tedersoo
Kessy Abarenkov
Martin Ryberg
Erik Kristiansson
Martin Hartmann
Conrad L. Schoch
Johan A. A. Nylander
Johannes Bergsten
Teresita M. Porter
Ari Jumpponen
Parag Vaishampayan
Otso Ovaskainen
Nils Hallenberg
Johan Bengtsson-Palme
Karl Martin Eriksson
Karl-Henrik Larsson
Ellen Larsson
Urmas Kõljalg
Published in MycoKeys
Volume 4
Pages 37-63
ISSN 1314-4057
Publication year 2012
Published at Institute of Neuroscience and Physiology, Department of Physiology
Department of Mathematical Sciences, Mathematical Statistics
Department of Biological and Environmental Sciences
Pages 37-63
Language en
Keywords ITS, sequence reliability, sequence quality control, fungi, databases, barcoding
Subject categories Soil biology, Medical microbiology, Biological Systematics, Terrestrial ecology, Bioinformatics and Systems Biology, Botany, Microbiology


Molecular data form an important research tool in most branches of mycology. A non-trivial proportion of the public fungal DNA sequences are, however, compromised in terms of quality and reliability, contributing noise and bias to sequence-borne inferences such as phylogenetic analysis, diversity assessment, and barcoding. In this paper we discuss various aspects and pitfalls of sequence quality assessment. Based on our observations, we provide a set of guidelines to assist in manual quality management of newly generated, near-full-length (Sanger-derived) fungal ITS sequences and to some extent also sequences of shorter read lengths, other genes or markers, and groups of organisms. The guidelines are intentionally non-technical and do not require substantial bioinformatics skills or significant computational power. Despite their simple nature, we feel they would have caught the vast majority of the severely compromised ITS sequences in the public corpus. Our guidelines are nevertheless not infallible, and common sense and intuition remain important elements in the pursuit of compromised sequence data. The guidelines focus on basic sequence authenticity and reliability of the newly generated sequences, and the user may want to consider additional resources and steps to accomplish the best possible quality control. A discussion on the technical resources for further sequence quality management is therefore provided in the supplementary material.

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