ISSN:
1550-7408
Source:
Blackwell Publishing Journal Backfiles 1879-2005
Topics:
Biology
Notes:
Recent culture-independent studies based on small-subunit ribosomal RNA (SSU rRNA) gene analysis revealed the existence of completely new clades of protists. The main problems with this approach are to correlate sequences from environmental rRNA genes with the organisms they belong to and then to detect the ecological role of these organisms in the environment. In order to overcome such problems we chose an alternative approach allowing us both a molecular characterization of uncultivable organisms with a low relative abundance in environmental samples, and a morphological analysis, even if restricted. The experimental protocol consists of two steps: an initial observation and photo-taking of the single cell under the DIC (Differential Interferential Contrast) microscope and then PCR amplification and direct sequencing of the 18S rRNA gene of the same cell. The advantages of this method are the possibility to: (1) establish a precise link between morphology and gene sequence; (2) detect the possible occurrence of highly similar species within the studied population; (3) avoid the insertion of Taq-polymerase errors in the gene sequence; and (4) detect possible polymorphisms in the gene under examination. Such an approach was used to sequence the 18S rRNA gene of organisms belonging to the class Karyorelictea that comprises several uncultivable ciliates with limited distinctive features. Gene sequences analysis revealed an unexpected genetic variability in trachelocercids and, in particular, the existence of polymorphisms in the SSU rRNA gene of a group of them. Such specimens show a similar morphology and, as a result of phylogenetic analysis, they form a constant clade.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1111/j.1550-7408.2005.05202003_6_2.x
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