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ID: 241306.0, MPI für Evolutionsbiologie / Ecophysiology
Use of group-specific PCR primers for identification of chrysophytes by denaturing gradient gel electrophoresis
Authors:Berglund, Johnny; Jürgens, Klaus; Bruchmüller, Iris; Wedin, Mats; Andersson, Agneta
Language:English
Date of Publication (YYYY-MM-DD):2005-05-30
Title of Journal:Aquatic Microbial Ecology
Journal Abbrev.:Aquat Microb Ecol
Volume:39
Issue / Number:2
Start Page:171
End Page:182
Review Status:not specified
Audience:Not Specified
Abstract / Description:The chrysophytes Chrysophyceae and Synurophyceae are ecologically important groups of autotrophic, mixotrophic and heterotrophic flagellates. The smallest forms are difficult to identify by classical microscopy but have important functions both as primary producers and as consumers of bacteria in the aquatic food chain. Group-specific primers for amplification of the 18S small sub-unit rRNA gene were developed for analysis of chrysophyte diversity by denaturing gradient gel electrophoresis (DGGE). Two different primer pairs were tested. The first primer pair (EukC1-F-Chryso-R) primarily targeted Paraphysomonadaceae and Ochromonadales, which generally have heterotrophic or mixotrophic nutrition. The second primer pair (EukC2-F-Chryso-R) targeted both Chrysophyceae and Synurophyceae. The primer pairs were tested for PCR amplification of the 18S rRNA gene of 25 cultured chrysophyte species and 6 other closely related nanoplanktonic species. Both primer pairs performed well, since PCR products were obtained for the corresponding chrysophyte cultures. None of the non-chrysophyte species were amplified with these primers. PCR products of chrysophyte cultures could be separated by DGGE in a denaturing gradient from 40 to 60%. In order to test this PCR-DGGE system for natural planktonic systems, we used field samples from a brackish water area (Baltic Sea) and a freshwater lake. The most intense DGGE bands were excised, sequenced and compared to sequences in GenBank, All obtained sequences grouped within the chrysophytes. Thus, the method seems to be promising for examining chrysophyte diversity in planktonic systems.
Free Keywords:chrysophytes; Chrysophyceae; Synurophyceae; heterotrophic nanoflagellates; denaturing gradient gel electrophoresis; DGGE; PCR-primers; 18S ribosomal DNA
External Publication Status:published
Document Type:Article
Affiliations:MPI für Limnologie/Abt. Ökophysiologie
External Affiliations:Umea Univ, Dept Ecol & Environm Sci, Umea, S-90187 Sweden;
Umea Marine Sci Ctr, Hornefors, S-91020 Sweden;
Baltic Sea Res Inst Warnemunde, Rostock, D-18119 Germany
Identifiers:ISSN:0948-3055 [ID-No. 1]
LOCALID:2397/S 38402 [Listen-Nummer/S-Nummer]
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