Diversity Measures in Environmental Sequences Are Highly Dependent on Alignment Quality—Data from ITS and New LSU Primers Targeting Basidiomycetes
| Autor: | Danuta Kapturska, Tesfaye Wubet, Dirk Krüger, Rolf Daniel, Christiane Fischer |
|---|---|
| Přispěvatelé: | Stajich, Jason E. |
| Jazyk: | angličtina |
| Rok vydání: | 2012 |
| Předmět: |
0106 biological sciences
Operational taxonomic unit Ecological Metrics Sequence analysis Science Entropy Molecular Sequence Data Sequence alignment Mycology Plant Science 010603 evolutionary biology 01 natural sciences Microbiology DNA Ribosomal 03 medical and health sciences Phylogenetics Agaricomycotina DNA Ribosomal Spacer Databases Genetic Environmental Microbiology Evolutionary Systematics Clade DNA Fungal Ribosomal DNA Biology Soil Microbiology 030304 developmental biology DNA Primers Genetics 0303 health sciences Evolutionary Biology molecular ecology phylogenetic analyses Multidisciplinary biology Phylogenetic tree Ecology Base Sequence Basidiomycota Botany Genetic Variation 15. Life on land Ribosome Subunits Large Eukaryotic biology.organism_classification Evolutionary biology Medicine Sequence Alignment Research Article |
| Zdroj: | PLoS ONE PLoS ONE, Vol 7, Iss 2, p e32139 (2012) |
| ISSN: | 1932-6203 |
| Popis: | 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. peerReviewed |
| Databáze: | OpenAIRE |
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