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Use of group-specific PCR primers for identification of chrysophytes by denaturing gradient gel electrophoresis
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och geovetenskap. Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Umeå marina forskningscentrum (UMF). (UMFpub)
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och geovetenskap.
Vise andre og tillknytning
2005 (engelsk)Inngår i: Aquatic Microbial Ecology, ISSN 0948-3055, E-ISSN 1616-1564, ISSN 0948-3055, Vol. 39, nr 2, s. 171-182Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

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.

sted, utgiver, år, opplag, sider
Olendorf/Luhe: Inter-Research , 2005. Vol. 39, nr 2, s. 171-182
Emneord [en]
Chrysophytes, chrysophyceae, synurophyceae, heterotrophic nanoflagellates, denaturing gradient gel electrophoresis, dgge, pcr-primers, 18s ribosomal dna
Identifikatorer
URN: urn:nbn:se:umu:diva-4773DOI: 10.3354/ame039171OAI: oai:DiVA.org:umu-4773DiVA, id: diva2:144006
Tilgjengelig fra: 2005-10-26 Laget: 2005-10-26 Sist oppdatert: 2018-06-09bibliografisk kontrollert
Inngår i avhandling
1. Pelagic microorganisms in the northern Baltic Sea: Ecology, diversity and food web dynamics
Åpne denne publikasjonen i ny fane eller vindu >>Pelagic microorganisms in the northern Baltic Sea: Ecology, diversity and food web dynamics
2005 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

Heterotrophic microorganisms are important for the flow of carbon and nutrients in the sea. Bacteria, nanoflagellates and ciliates are relevant components of the pelagic food web. In order to be able to predict the outcome of e.g. eutrophication or climate change we need to know how the different components of the pelagic food web are regulated. With the focus on the northern Baltic Sea food web, this thesis deals with limitation and control of heterotrophic protists, the effect of resource heterogeneity on food web efficiency and diversity of nanoflagellates.

In-situ microcosm experiments showed that the net growth of heterotrophic flagellates were resource limited throughout the year. Field data confirmed that the abundance of flagellates was bottom-up controlled. Furthermore, field data also showed that the annual average biomass of protists, flagellates and ciliates increased with primary productivity. On a smaller seasonal scale temperature and bacterial biomass were able to explain most of the variation in flagellate biovolume. The temporal variation in ciliate biovolume could not be explained by any bottom-up factors like bacterial biomass, flagellate biomass or chlorophyll a. This and an in-situ microcosm experiment implied that the seasonal dynamics of ciliates were more regulated by predators like mesozooplankton.

The food web efficiency i.e. how much of production at the resource level is converted to production at the top trophic level, may be affected by specific size or type of resource. Indoor mesocosms revealed that the food web efficiency was 11 times lower when heterotrophic bacteria dominated basal production instead of nano- and micro-sized phytoplankton. This was due to a lengthening of the food web when pico-sized bacteria constituted the main resource.

The PCR-DGGE molecular biological method was used to study the diversity of heterotrophic or mixotrophic chrysomonads. The focus was set on chrysomonads due to their relatively large contribution to the nanoflagellate community. Group-specific PCR primers were optimized for the target group. A field survey in the northern Baltic Sea showed that a handful of chrysomonad sequences were present throughout the year. Significantly more chrysomonads were recorded in the basin with higher primary productive and salinity. In total 15-16 different chrysomonad sequences were recorded. Most of them matched uncultured chrysomonad clones.

sted, utgiver, år, opplag, sider
Umeå: Ekologi, miljö och geovetenskap, 2005. s. 36
Emneord
resource limitation, predation limitation, microbial food web, heterotrophic microorganisms, nanoflagellates, ciliates, protists, resource heterogeneity, chrysomonads, chrysophytes, group-specific PCR-DGGE primers, Baltic Sea
HSV kategori
Identifikatorer
urn:nbn:se:umu:diva-618 (URN)91-7305-957-9 (ISBN)
Disputas
2005-11-26, Stora hörsalen, KB3B1, KBC, 10:00
Opponent
Veileder
Tilgjengelig fra: 2005-10-26 Laget: 2005-10-26 Sist oppdatert: 2011-03-29bibliografisk kontrollert

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