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Applied and Environmental Microbiology, December 2008, p. 7174-7182, Vol. 74, No. 23
0099-2240/08/$08.00+0     doi:10.1128/AEM.01298-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Quantification of Diatom and Dinoflagellate Biomasses in Coastal Marine Seawater Samples by Real-Time PCR

Anna Godhe,^1* Maria E. Asplund,¹ Karolina Härnström,¹ V. Saravanan,² Anuj Tyagi,² and Indrani Karunasagar²

Department of Marine Ecology, University of Gothenburg, Box 461, SE 405 30 Göteborg, Sweden,¹ Department of Fishery Microbiology, College of Fisheries, Karnataka Veterinary Animal and Fisheries Sciences University, P.B. No. 527, Mangalore 575 002, India²

Received 11 June 2008/ Accepted 2 October 2008

Two real-time PCR assays targeting the small-subunit (SSU) ribosomal DNA (rDNA) were designed to assess the proportional biomass of diatoms and dinoflagellates in marine coastal water. The reverse primer for the diatom assay was designed to be class specific, and the dinoflagellate-specific reverse primer was obtained from the literature. For both targets, we used universal eukaryotic SSU rDNA forward primers. Specificity was confirmed by using a BLAST search and by amplification of cultures of various phytoplankton taxa. Reaction conditions were optimized for each primer set with linearized plasmids from cloned SSU rDNA fragments. The number of SSU rDNA copies per cell was estimated for six species of diatoms and nine species of dinoflagellates; these were significantly correlated to the biovolumes of the cells. Nineteen field samples were collected along the Swedish west coast and subjected to the two real-time PCR assays. The linear regression of the proportion of SSU rDNA copies of dinoflagellate and diatom origin versus the proportion of dinoflagellate and diatom biovolumes or biomass per liter was significant. For diatoms, linear regression of the number of SSU rDNA copies versus biovolume or biomass per liter was significant, but no such significant correlation was detected in the field samples for dinoflagellates. The method described will be useful for estimating the proportion of dinoflagellate versus diatom biovolume or biomass and the absolute diatom biovolume or biomass in various aquatic disciplines.

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* Corresponding author. Mailing address: Department of Marine Ecology, University of Gothenburg, Box 461, SE 405 30 Göteborg, Sweden. Phone: 46 31 7862709. Fax: 46 31 786 2727. E-mail: anna.godhe{at}marecol.gu.se

Published ahead of print on 10 October 2008.

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Applied and Environmental Microbiology, December 2008, p. 7174-7182, Vol. 74, No. 23
0099-2240/08/$08.00+0     doi:10.1128/AEM.01298-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

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