PacBio sequencing of Glomeromycota rDNA: a novel amplicon covering all widely used ribosomal barcoding regions and its applicability in taxonomy and ecology of arbuscular mycorrhizal fungi
Language English Country Great Britain, England Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
PubMed
33780549
DOI
10.1111/nph.17372
Knihovny.cz E-resources
- Keywords
- Archaeosporales, Glomeromycota, PacBio, long-read metabarcoding, mycorrhizal fungi distribution, third-generation sequencing,
- MeSH
- DNA, Fungal genetics MeSH
- Phylogeny MeSH
- Glomeromycota * genetics MeSH
- Fungi genetics MeSH
- Mycorrhizae * genetics MeSH
- DNA, Ribosomal genetics MeSH
- Sequence Analysis, DNA MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- DNA, Fungal MeSH
- DNA, Ribosomal MeSH
There is no consensus barcoding region for determination of arbuscular mycorrhizal fungal (AMF) taxa. To overcome this obstacle, we have developed an approach to sequence an AMF marker within the ribosome-encoding operon (rDNA) that covers all three widely applied variable molecular markers. Using a nested PCR approach specific to AMF, we amplified a part (c. 2.5 kb) of the rDNA spanning the majority of the small subunit rRNA (SSU) gene, the complete internal transcribed spacer (ITS) region and a part of the large subunit (LSU) rRNA gene. The PCR products were sequenced on the PacBio platform utilizing Single Molecule Real Time (SMRT) sequencing. Employing this method for selected environmental DNA samples, we were able to describe complex AMF communities consisting of various glomeromycotan lineages. We demonstrate the applicability of this new 2.5 kb approach to provide robust phylogenetic assignment of AMF lineages without known sequences from pure cultures and to consolidate information about AMF taxon distributions coming from three widely used barcoding regions into one integrative dataset.
Faculty of Science Charles University Prague Prague CZ 128 44 Czech Republic
Institute of Botany of the Czech Academy of Sciences Průhonice CZ 252 43 Czech Republic
Institute of Microbiology of the Czech Academy of Sciences Prague CZ 142 20 Czech Republic
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