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Revealing genetic diversity of eukaryotic microorganisms in aquatic environments by denaturing gradient gel electrophoresis

Van Hannen, E.J. and Van Agterveld, M.P. and Gons, H.J. and Laanbroek, H.J. (1998) Revealing genetic diversity of eukaryotic microorganisms in aquatic environments by denaturing gradient gel electrophoresis. Journal of Phycology, 34, 206-213. ISSN 0022-3646.

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A new Eucarya-specific 18S rDNA primer set was constructed and tested using denaturing gradient gel electrophoresis to analyze the genetic diversity of eukaryotic microorganisms in aquatic environments. All eukaryal lines of descent exhibited four or fewer nucleotide mismatches in the forward primer sequence, except for the microspora line of descent. The reverse primer annealed to a more conserved region with fewer than two nucleotide mismatches. Genomic DNA from test organisms with different numbers of nucleotide mismatches were amplified to test primer specificity. Relatively low annealing temperatures allowed the amplification of sequences with up to four nucleotide mismatches while still maintaining specificity for the eukaryal domain. Denaturing gradient gel electrophoresis was used to separate similarly sized PCR products of environmental samples, and the obtained banding patterns were converted to a binary format for statistical comparisons. Cluster analysis of these patterns showed similar results to a cluster analysis based on environmental variables. This approach provides an analytical tool to study the population structure and molecular ecology of eukaryotic microbial communities inhabiting aquatic environments. [KEYWORDS: 18S rRNA; aquatic environments; community structure; DGGE; Eucarya; genetic diversity; nucleotide mismatches; Southern blot hybridization 16s ribosomal-rna; polymerase chain-reaction; marine picoplankton; flow-cytometry; identification; fragments; bacteria; probes; phytoplankton; populations]

Item Type:Article
Institutes:Nederlands Instituut voor Ecologie (NIOO)
ID Code:10690
Deposited On:25 Nov 2011 01:00
Last Modified:31 Mar 2014 10:59

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