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Sequence variation in nuclear ribosomal small subunit, internal transcribed spacer and large subunit regions of Rhizophagus irregularis and Gigaspora margarita is high and isolate‐dependent
Author(s) -
Thiéry Odile,
Vasar Martti,
Jairus Teele,
Davison John,
Roux Christophe,
Kivistik PaulaAnn,
Metspalu Andres,
Milani Lili,
Saks Ülle,
Moora Mari,
Zobel Martin,
Öpik Maarja
Publication year - 2016
Publication title -
molecular ecology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.619
H-Index - 225
eISSN - 1365-294X
pISSN - 0962-1083
DOI - 10.1111/mec.13655
Subject(s) - biology , internal transcribed spacer , ribosomal rna , rhizophagus irregularis , genetics , ribosomal dna , amplicon , genetic variation , gene , phylogenetics , polymerase chain reaction , symbiosis , arbuscular mycorrhizal , bacteria
Arbuscular mycorrhizal ( AM ) fungi are known to exhibit high intra‐organism genetic variation. However, information about intra‐ vs. interspecific variation among the genes commonly used in diversity surveys is limited. Here, the nuclear small subunit ( SSU ) rRNA gene, internal transcribed spacer ( ITS ) region and large subunit ( LSU ) rRNA gene portions were sequenced from 3 to 5 individual spores from each of two isolates of Rhizophagus irregularis and Gigaspora margarita . A total of 1482 Sanger sequences (0.5 Mb) from 239 clones were obtained, spanning ~4370 bp of the ribosomal operon when concatenated. Intrasporal and intra‐isolate sequence variation was high for all three regions even though variant numbers were not exhausted by sequencing 12–40 clones per isolate. Intra‐isolate nucleotide variation levels followed the expected order of ITS > LSU > SSU , but the values were strongly dependent on isolate identity. Single nucleotide polymorphism ( SNP ) densities over 4 SNP /kb in the ribosomal operon were detected in all four isolates. Automated operational taxonomic unit picking within the sequence set of known identity overestimated species richness with almost all cut‐off levels, markers and isolates. Average intraspecific sequence similarity values were 99%, 96% and 94% for amplicons in SSU , LSU and ITS , respectively. The suitability of the central part of the SSU as a marker for AM fungal community surveys was further supported by its level of nucleotide variation, which is similar to that of the ITS region; its alignability across the entire phylum; its appropriate length for next‐generation sequencing; and its ease of amplification in single‐step PCR.