Evaluating the potential of 18S rDNA clone libraries to complement pyrosequencing data of marine protists with near full-length sequence information

Abstract

Sequencing of 18S rDNA clone libraries and 454-pyrosequencing are valuable methods used to describe microbial diversity. The massively parallel 454-pyrosequencing generates vast amounts of ribosomal sequence data and has the potential to uncover more organisms, even rare species. However, the relatively short sequence lengths of ∼500 bp are suboptimal for taxonomic annotation and phylogenetic analyses. In this study, we assessed the potential of 18S ribosomal clone libraries to complement corresponding 454-pyrosequencing data with near full-length sequence information. This involved a comparison of protist community compositions in five polar samples suggested by 18S rDNA clone libraries, with the corresponding community compositions suggested by 454-pyrosequencing. The study was conducted with four Arctic water samples, focusing on the eukaryotic picoplankton (0.4–3 µm), and with one sample collected in the Southern Ocean, examining the entire size spectrum (> 0.4 µm). For all individual samples, the protist community compositions suggested by the two different approaches showed significant similarities. Around 70% of the sequences detected by sequencing of clone libraries were also present in the 454-pyrosequencing data set. However, the clone library sequences reflected only ∼20% of the abundant biosphere identified by 454-pyrosequencing and identified ribosomal sequences that were not detected in the 454-pyrosequencing data sets.

Key words:

• 18S rDNA near full-length clones
• genetic diversity
• polar regions

Acknowledgements

We thank the captain and crew of the RV Polarstern for their support during the cruises. We are very grateful to Stephan Frickenhaus, Fabian Kilpert and Bank Beszteri for their bioinformatical support. We also want to thank Annika Schroer, Anja Nicolaus and Kerstin Oetjen for excellent technical support in the laboratory. We are grateful to Steven Holland for providing access to the program Analytic Rarefaction 1.3.

Funding

This study was accomplished within the Young Investigator Group PLANKTOSENS (VH-NG-500), funded by the Initiative and Networking Fund of the Helmholtz Association.

Supplementary material

Supplementary material for this article is available via the Supplemental tab of the article's online page at http://dx.doi.org/10.1080/17451000.2013.852685.

Editorial responsibility: Hongyue Dang

Additional information

Funding

Funding: This study was accomplished within the Young Investigator Group PLANKTOSENS (VH-NG-500), funded by the Initiative and Networking Fund of the Helmholtz Association.