Figure 1.
Nucleotide variability in 18S rRNA genes.
Shannon entropy values of all eukaryotic alignment positions from the SILVA database along the 18S rRNA gene of Saccharomyces cerevisiae. Red dots mark consecutive nucleotide positions where at least 90% of ≥10 nt have entropy values lower than 0.2. The highly variable regions of 18S rRNA gene are denoted V1 to V9. In total, 100 primers targeting eukaryotes from the literature as well as those designed in this study are positioned along the reference sequence. The direction of the arrows indicates the orientation of the primers. The color denotes the melting temperature, and the thickness of the arrows represents the eukaryote universality of the primers.
Figure 2.
Variable regions of the 18S rRNA gene.
The amplicon from the optimal primers selected in this study was included. Expected coverage of biodiversity is shown as a function of cut-off values for sequence similarity. V2, V4 and V9 generated the best results at all sequence identity cut-off levels.
Table 1.
Primers generated following the in silico analysis.
Figure 3.
Empirical primer test on marine sediments.
Taxonomic coverage of reads from 454 pyrosequencing on pooled DNA extracts from three different marine sediments from the Norwegian continental shelf. The figure indicates lower rank levels of the taxonomic tree of the best hits after running BLAST on representative sequences of 6823 OTUs against the SILVA database.