Figure 1.
Entropy plot of 42,109 soil derived 16S rRNA gene sequence alignment.
Hypervariable regions indicated as designated by Baker et al. [24] E. coli nucleotide numbering. Sequence area presented excludes poorly supported areas from the beginning and end of the sequences (due to nearly full sequences) and thus excludes the V9 region.
Figure 2.
16S rRNA gene sequence conservation of soil derived sequences.
A) Nucleic acid base composition of the 16S rRNA gene consensus sequence of the 41,109 RDP database soil derived sequences for 90% conservation cutoff value. Red background positions include hypervariable stretches as reported in reference [24] and expanded in the current study, while green background positions are proposed primer designing sites in reference [11]. The IUPAC system was used for denoting per base variability (degeneracies) and lower-case letters are used for nucleotide positions where gaps participated by more than 10% in the position throughout the sequence alignment. B) Comparison of present study results for 95% sequence conservation with the ones provided in reference [11] for 90% sequence conservation. Letter color coding referring to differences found on sequences of this study compared to that of reference [11]: red) increased variability; blue) altered degeneracy without variability increase; green) reduced variability; grey) although presence of two nucleotides in that position is implied in reference [11], these are missing in the published table.
Figure 3.
Distribution of commonly screened V region fragment lengths.
Fragment lengths including the examined hypervariable regions for all screened (41,109) sequences. Sequence fragments were plotted according to length ascending order.
Figure 4.
Pearson correlation tests between corresponding sequence distances of examined V regions and FL variants.
All tests were significant (P<001). Test correlation index (r) values and linear models (presented with solid lines) used to describe overall trends are provided above and below each plot. Local relationships between corresponding sequence distances of the FL and other datasets are expressed with the non-parametric LOWESS (locally weighted regression and smoothing scatterplots) regression analysis plotting (dot-dashed lines), while the ideal y = x correlation is also plotted (dashed lines).
Figure 5.
Taxonomy classification depth comparisons among V region datasets and the FL variants.
Figure 6.
Over or under representation of phyla in the examined datasets.
Values are expressed as percentage of the taxonomical annotations obtained for the FL sequence variants. Taxa were characterized into groups according to the existing sequence numbers as indicated in Table 1.
Table 1.
Classification of the full-length sequences and their trimmed to the examined V region variants.
Figure 7.
Taxonomy, OTU (3% sequence distance) analysis and Unifrac results of the performed simulation.
A) PCA results of matrix generated by sample distances based on classified sequence relative abundance (left) and presence absence (right) for the V regions and FL datasets. B) Similarly to A for OTU relative abundance (left) and presence absence (right). C) PCA results for matrices generated using the weighted (left - phylotype relative abundance based) and unweighted (right - phylotype occurrence based) Unifrac analysis result distances between samples for the V regions and FL datasets.
Figure 8.
Study methods overview.