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Background  Identifying species of organisms by short sequences of DNA has been in the center of ongoing discussions under the terms DNA barcoding or DNA taxonomy. A C-terminal fragment of the mitochondrial gene for  cytochrome oxidase subunit I  ( COI ) has been proposed as universal marker for this purpose among animals.    Results  Herein we present experimental evidence that the mitochondrial  16S rRNA  gene fulfills the requirements for a universal DNA barcoding marker in amphibians. In terms of universality of priming sites and identification of major vertebrate clades the studied  16S  fragment is superior to  COI . Amplification success was 100% for  16S  in a subset of fresh and well-preserved samples of Madagascan frogs, while various combination of  COI  primers had lower success rates. COI  priming sites showed high variability among amphibians both at the level of groups and closely related species, whereas  16S  priming sites were highly conserved among vertebrates. Interspecific pairwise  16S  divergences in a test group of Madagascan frogs were at a level suitable for assignment of larval stages to species (1–17%), with low degrees of pairwise haplotype divergence within populations (0–1%).    Conclusion  We strongly advocate the use of  16S rRNA  as standard DNA barcoding marker for vertebrates to complement  COI , especially if samples  a priori  could belong to various phylogenetically distant taxa and false negatives would constitute a major problem.

Comparative performance of the 16S rRNA gene in DNA barcoding of amphibians.