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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
Author(s) -
Kolaříková Zuzana,
Slavíková Renata,
Krüger Claudia,
Krüger Manuela,
Kohout Petr
Publication year - 2021
Publication title -
new phytologist
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.742
H-Index - 244
eISSN - 1469-8137
pISSN - 0028-646X
DOI - 10.1111/nph.17372
Subject(s) - biology , internal transcribed spacer , glomeromycota , ribosomal rna , dna barcoding , phylogenetic tree , amplicon , ribosomal dna , thaumarchaeota , evolutionary biology , gene , computational biology , dna sequencing , genetics , botany , 16s ribosomal rna , polymerase chain reaction , arbuscular mycorrhizal , bacteria , symbiosis
Summary 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.