Dissimilarity of Species and Forms of Planktonic Neocalanus Copepods Using Mitochondrial COI, 12S, Nuclear ITS, and 28S Gene Sequences

Background: A total of six Neocalanus species inhabit the oceans of the world. Of these, three species plus form variants (N. cristatus, N. plumchrus, N. flemingeri large form, and N. flemingeri small form), which constitute a monophyletic group among Neocalanus copepods, occur in the Northwestern Pacific off Japan. In the present study, we have tried to discriminate the three species plus form variants of Neocalanus copepods based on sequences of four DNA marker regions. Methodology/Principal Findings: Discrimination was performed based on the DNA sequence information from four genetic markers, including the mitochondrial COI, 12S, nuclear ITS, and 28S gene regions. Sequence dissimilarity was compared using both distance- and character-based approaches. As a result, all three species were confirmed to be distinct based on the four genetic marker regions. On the contrary, distinction of the form variants was only confirmed based on DNA sequence of the mitochondrial COI gene region. Conclusions/Significance: Although discrimination was not successful for the form variants based on the mitochondrial 12S, nuclear ITS, and 28S genes, diagnostic nucleotide sequence characters were observed in their mitochondrial COI gene sequences. Therefore, these form variants are considered to be an important unit of evolution below the species level, an

Zooplankton diversity analysis through single-gene sequencing of a community sample

<p>Abstract</p> <p>Background</p> <p>Oceans cover more than 70% of the earth's surface and are critical for the homeostasis of the environment. Among the components of the ocean ecosystem, zooplankton play vital roles in energy and matter transfer through the system. Despite their importance, understanding of zooplankton biodiversity is limited because of their fragile nature, small body size, and the large number of species from various taxonomic phyla. Here we present the results of single-gene zooplankton community analysis using a method that determines a large number of mitochondrial <it>COI </it>gene sequences from a bulk zooplankton sample. This approach will enable us to estimate the species richness of almost the entire zooplankton community.</p> <p>Results</p> <p>A sample was collected from a depth of 721 m to the surface in the western equatorial Pacific off Pohnpei Island, Micronesia, with a plankton net equipped with a 2-m²mouth opening. A total of 1,336 mitochondrial <it>COI </it>gene sequences were determined from the cDNA library made from the sample. From the determined sequences, the occurrence of 189 species of zooplankton was estimated. BLASTN search results showed high degrees of similarity (>98%) between the query and database for 10 species, including holozooplankton and merozooplankton.</p> <p>Conclusion</p> <p>In conjunction with the Census of Marine Zooplankton and Barcode of Life projects, single-gene zooplankton community analysis will be a powerful tool for estimating the species richness of zooplankton communities.</p

PCR Primers for Metazoan Mitochondrial 12S Ribosomal DNA Sequences

Background: Assessment of the biodiversity of communities of small organisms is most readily done using PCR-based analysis of environmental samples consisting of mixtures of individuals. Known as metagenetics, this approach has transformed understanding of microbial communities and is beginning to be applied to metazoans as well. Unlike microbial studies, where analysis of the 16S ribosomal DNA sequence is standard, the best gene for metazoan metagenetics is less clear. In this study we designed a set of PCR primers for the mitochondrial 12S ribosomal DNA sequence based on 64 complete mitochondrial genomes and then tested their efficacy. Methodology/Principal Findings: A total of the 64 complete mitochondrial genome sequences representing all metazoan classes available in GenBank were downloaded using the NCBI Taxonomy Browser. Alignment of sequences was performed for the excised mitochondrial 12S ribosomal DNA sequences, and conserved regions were identified for all 64 mitochondrial genomes. These regions were used to design a primer pair that flanks a more variable region in the gene. Then all of the complete metazoan mitochondrial genomes available in NCBI’s Organelle Genome Resources database were used to determine the percentage of taxa that would likely be amplified using these primers. Results suggest that these primers will amplify target sequences for many metazoans. Conclusions/Significance: Newly designed 12S ribosomal DNA primers have considerable potential for metazoa