Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.

The ribosomal DNA comprised of the ITS1-5.8S-ITS2 regions is widely used as a fungal marker in molecular ecology and systematics but cannot be aligned with confidence across genetically distant taxa. In order to study the diversity of Agaricomycotina in forest soils, we designed primers targeting th...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Dirk Krüger, Danuta Kapturska, Christiane Fischer, Rolf Daniel, Tesfaye Wubet
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/a7bc57283a5d488a80fae6b6756610ed
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:a7bc57283a5d488a80fae6b6756610ed
record_format	dspace
spelling	oai:doaj.org-article:a7bc57283a5d488a80fae6b6756610ed2021-11-18T07:27:11ZDiversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.1932-620310.1371/journal.pone.0032139https://doaj.org/article/a7bc57283a5d488a80fae6b6756610ed2012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22363808/?tool=EBIhttps://doaj.org/toc/1932-6203The ribosomal DNA comprised of the ITS1-5.8S-ITS2 regions is widely used as a fungal marker in molecular ecology and systematics but cannot be aligned with confidence across genetically distant taxa. In order to study the diversity of Agaricomycotina in forest soils, we designed primers targeting the more alignable 28S (LSU) gene, which should be more useful for phylogenetic analyses of the detected taxa. This paper compares the performance of the established ITS1F/4B primer pair, which targets basidiomycetes, to that of two new pairs. Key factors in the comparison were the diversity covered, off-target amplification, rarefaction at different Operational Taxonomic Unit (OTU) cutoff levels, sensitivity of the method used to process the alignment to missing data and insecure positional homology, and the congruence of monophyletic clades with OTU assignments and BLAST-derived OTU names. The ITS primer pair yielded no off-target amplification but also exhibited the least fidelity to the expected phylogenetic groups. The LSU primers give complementary pictures of diversity, but were more sensitive to modifications of the alignment such as the removal of difficult-to align stretches. The LSU primers also yielded greater numbers of singletons but also had a greater tendency to produce OTUs containing sequences from a wider variety of species as judged by BLAST similarity. We introduced some new parameters to describe alignment heterogeneity based on Shannon entropy and the extent and contents of the OTUs in a phylogenetic tree space. Our results suggest that ITS should not be used when calculating phylogenetic trees from genetically distant sequences obtained from environmental DNA extractions and that it is inadvisable to define OTUs on the basis of very heterogeneous alignments.Dirk KrügerDanuta KapturskaChristiane FischerRolf DanielTesfaye WubetPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 2, p e32139 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Dirk Krüger Danuta Kapturska Christiane Fischer Rolf Daniel Tesfaye Wubet Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.
description	The ribosomal DNA comprised of the ITS1-5.8S-ITS2 regions is widely used as a fungal marker in molecular ecology and systematics but cannot be aligned with confidence across genetically distant taxa. In order to study the diversity of Agaricomycotina in forest soils, we designed primers targeting the more alignable 28S (LSU) gene, which should be more useful for phylogenetic analyses of the detected taxa. This paper compares the performance of the established ITS1F/4B primer pair, which targets basidiomycetes, to that of two new pairs. Key factors in the comparison were the diversity covered, off-target amplification, rarefaction at different Operational Taxonomic Unit (OTU) cutoff levels, sensitivity of the method used to process the alignment to missing data and insecure positional homology, and the congruence of monophyletic clades with OTU assignments and BLAST-derived OTU names. The ITS primer pair yielded no off-target amplification but also exhibited the least fidelity to the expected phylogenetic groups. The LSU primers give complementary pictures of diversity, but were more sensitive to modifications of the alignment such as the removal of difficult-to align stretches. The LSU primers also yielded greater numbers of singletons but also had a greater tendency to produce OTUs containing sequences from a wider variety of species as judged by BLAST similarity. We introduced some new parameters to describe alignment heterogeneity based on Shannon entropy and the extent and contents of the OTUs in a phylogenetic tree space. Our results suggest that ITS should not be used when calculating phylogenetic trees from genetically distant sequences obtained from environmental DNA extractions and that it is inadvisable to define OTUs on the basis of very heterogeneous alignments.
format	article
author	Dirk Krüger Danuta Kapturska Christiane Fischer Rolf Daniel Tesfaye Wubet
author_facet	Dirk Krüger Danuta Kapturska Christiane Fischer Rolf Daniel Tesfaye Wubet
author_sort	Dirk Krüger
title	Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.
title_short	Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.
title_full	Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.
title_fullStr	Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.
title_full_unstemmed	Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.
title_sort	diversity measures in environmental sequences are highly dependent on alignment quality--data from its and new lsu primers targeting basidiomycetes.
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/a7bc57283a5d488a80fae6b6756610ed
work_keys_str_mv	AT dirkkruger diversitymeasuresinenvironmentalsequencesarehighlydependentonalignmentqualitydatafromitsandnewlsuprimerstargetingbasidiomycetes AT danutakapturska diversitymeasuresinenvironmentalsequencesarehighlydependentonalignmentqualitydatafromitsandnewlsuprimerstargetingbasidiomycetes AT christianefischer diversitymeasuresinenvironmentalsequencesarehighlydependentonalignmentqualitydatafromitsandnewlsuprimerstargetingbasidiomycetes AT rolfdaniel diversitymeasuresinenvironmentalsequencesarehighlydependentonalignmentqualitydatafromitsandnewlsuprimerstargetingbasidiomycetes AT tesfayewubet diversitymeasuresinenvironmentalsequencesarehighlydependentonalignmentqualitydatafromitsandnewlsuprimerstargetingbasidiomycetes
_version_	1718423428814340096

Diversity measures in environmental sequences are highly dependent on alignment quality--data from ITS and new LSU primers targeting basidiomycetes.

Ejemplares similares