Open AccessA novel metabarcoding primer pair for environmental DNA analysis of Cephalopoda (Mollusca) targeting the nuclear 18S rRNA region
Author(s) -
Daniëlle de Jonge,
Véronique Merten,
Till Bayer,
Oscar Puebla,
Thorsten B. H. Reusch,
Henk-Jan T. Hoving
Publication year - 2021
Publication title -
royal society open science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.84
H-Index - 51
ISSN - 2054-5703
DOI - 10.1098/rsos.201388
Subject(s) - cephalopod , biology , environmental dna , primer (cosmetics) , octopus (software) , evolutionary biology , 18s ribosomal rna , biodiversity , genbank , ribosomal rna , genetics , zoology , gene , ecology , chemistry , physics , organic chemistry , quantum mechanics
Cephalopods are pivotal components of marine food webs, but biodiversity studies are hampered by challenges to sample these agile marine molluscs. Metabarcoding of environmental DNA (eDNA) is a potentially powerful technique to study oceanic cephalopod biodiversity and distribution but has not been applied thus far. We present a novel universal primer pair for metabarcoding cephalopods from eDNA, Ceph18S (Forward: 5′-CGC GGC GCT ACA TAT TAG AC-3′, Reverse: 5′-GCA CTT AAC CGA CCG TCG AC-3′). The primer pair targets the hypervariable region V2 of the nuclear 18S rRNA gene and amplifies a relatively short target sequence of approximately 200 bp in order to allow the amplification of degraded DNA. In silico tests on a reference database and empirical tests on DNA extracts from cephalopod tissue estimate that 44–66% of cephalopod species, corresponding to about 310–460 species, can be amplified and identified with this primer pair. A multi-marker approach with the novel Ceph18S and two previously published cephalopod mitochondrial 16S rRNA primer sets targeting the same region (Jarman et al . 2006 Mol. Ecol. Notes. 6 , 268–271; Peters et al . 2015 Mar. Ecol. 36 , 1428–1439) is estimated to amplify and identify 89% of all cephalopod species, of which an estimated 19% can only be identified by Ceph18S . All sequences obtained with Ceph18S were submitted to GenBank, resulting in new 18S rRNA sequences for 13 cephalopod taxa.