Publication Date:
2024-02-03
Description:
The aim of this study was to assess potential candidate gene regions and corresponding universal primer pairs as secondary DNA barcodes for the fungal kingdom, additional to ITS rDNA as primary barcode. Ampli\xef\xac\x81cation ef\xef\xac\x81ciencies of 14 (partially) universal primer pairs targeting eight genetic markers were tested across 〉\xe2\x80\xaf1\xe2\x80\xaf500 species (1\xe2\x80\xaf931 strains or specimens) and the outcomes of almost twenty thousand (19\xe2\x80\xaf577) polymerase chain reactions were evaluated. We tested several well-known primer pairs that amplify: i) sections of the nuclear ribosomal RNA gene large subunit (D1\xe2\x80\x93D2 domains of 26/28S); ii) the complete internal transcribed spacer region (ITS1/2); iii) partial \xce\xb2-tubulin II (TUB2); iv) \xce\xb3-actin (ACT); v) translation elongation factor 1-\xce\xb1 (TEF1\xce\xb1); and vi) the second largest subunit of RNA-polymerase II (partial RPB2, section 5\xe2\x80\x936). Their PCR ef\xef\xac\x81ciencies were compared with novel candidate primers corresponding to: i) the fungal-speci\xef\xac\x81c translation elongation factor 3 (TEF3); ii) a small ribosomal protein necessary for t-RNA docking; iii) the 60S L10 (L1) RP; iv) DNA topoisomerase I (TOPI); v) phosphoglycerate kinase (PGK); vi) hypothetical protein LNS2; and vii) alternative sections of TEF1\xce\xb1. Results showed that several gene sections are accessible to universal primers (or primers universal for phyla) yielding a single PCR-product. Barcode gap and multi-dimensional scaling analyses revealed that some of the tested candidate markers have universal properties providing adequate infra- and inter-speci\xef\xac\x81c variation that make them attractive barcodes for species identi\xef\xac\x81cation. Among these gene sections, a novel high \xef\xac\x81delity primer pair for TEF1\xce\xb1, already widely used as a phylogenetic marker in mycology, has potential as a supplementary DNA barcode with superior resolution to ITS. Both TOPI and PGK show promise for the Ascomycota, while TOPI and LNS2 are attractive for the Pucciniomycotina, for which universal primers for ribosomal subunits often fail.
Keywords:
DNA barcoding
;
ITS supplement
;
molecular taxonomy
;
phylogeny
;
species identi\xef\xac\x81cation
;
universal primers
Repository Name:
National Museum of Natural History, Netherlands
Type:
info:eu-repo/semantics/article
Format:
application/pdf
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