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ABSTRACT DNA metabarcoding of zooplankton biodiversity is used increasingly for monitoring global ocean ecosystems, requiring comparable data from different research laboratories and ocean regions. The MetaZooGene Intercalibration Experiment (MZG‐ICE) was designed to examine1 and analyse patterns of variation of DNA sequence data resulting from multi‐gene metabarcoding of 10 zooplankton samples carried out by 10 research groups affiliated with the Scientific Committee for Ocean Research (SCOR). Aliquots of DNA extracted from the 10 zooplankton samples were distributed to MZG‐ICE groups for metabarcoding of four gene regions: V1‐V2, V4 and V9 of nuclear 18S rRNA and mitochondrial COI. Molecular protocols and procedures were recommended; substitutions were allowed as necessary. Resulting data were uploaded to a common repository for centralised statistics and bioinformatics. Based on proportional sequence numbers for abundant phyla, overall patterns of variation were consistent across many—but not all—MZG‐ICE groups. V9 showed highest similarity, followed (in order) by V4, V1‐V2, and COI. Outlier data were hypothesised to result from the use of different PCR protocols and sequencing platforms, and possible contamination. MZG‐ICE results indicated that DNA metabarcoding data from different laboratories and research groups can provide reliable, accurate and valid descriptions of biodiversity of zooplankton throughout the ocean. Recommendations included: pre‐screening QA/QC of raw data, detailed records for laboratory protocols, reagents, and instrumentation, and centralised bioinformatics and multivariate statistics. In the absence of universal agreement on standardised protocols or best practices, intercalibration is the best way forward toward validation of DNA metabarcoding of zooplankton diversity for global ocean monitoring. 

Abstract Foraminifera are a species-rich phylum of rhizarian protists that are highly abundant in many marine environments and play a major role in global carbon cycling. Species recognition in Foraminifera is mainly based on morphological characters and nuclear 18S ribosomal RNA barcoding. The 18S rRNA contains variable sequence regions that allow for the identification of most foraminiferal species. Still, some species show limited variability, while others contain high levels of intragenomic polymorphisms, thereby complicating species identification. The use of additional, easily obtainable molecular markers other than 18S rRNA will enable more detailed investigation of evolutionary history, population genetics and speciation in Foraminifera. Here we present the first mitochondrial cytochrome c oxidase subunit 1 (COI) gene sequences (“barcodes”) of Foraminifera. We applied shotgun sequencing to single foraminiferal specimens, assembled COI, and developed primers that allow amplification of COI in a wide range of foraminiferal species. We obtained COI sequences of 49 specimens from 17 species from the orders Rotaliida and Miliolida. Phylogenetic analysis showed that the COI tree is largely congruent with previously published 18S rRNA phylogenies. Furthermore, species delimitation with ASAP and ABGD algorithms showed that foraminiferal species can be identified based on COI barcodes.