Comparison of individual libraries The Shared OTUs and Similarity (SONS) program [24] was used to compare the unfractioned sample with each of the %G+C fractions and with the combined sequence data from the fractions (Table 3). Using a 98% similarity criterion for the phylotypes, at least 80% of sequences from %G+C fractions GSK126 cell line 30–35 and 35–40 were shared with the unfractioned sample (Vobs values). However, for two of the high %G+C Seliciclib order content fractions with %G+C content from 55 to 65, the Vobs values were considerably lower (32–33%). When comparing the combined sequence data from the fractioned sample with the unfractioned sample, a higher percentage of sequences

and OTUs in the unfractioned were shared. Table 3 Results from library comparisons with SONS [24]. Library A Unfractioned Uobs a Vobs b Aotu_shared c Botu_shared d Library B Fr G+C 25–30% 0.41 0.40 0.22 0.34 Library B Fr G+C 30–35% 0.59 0.83 0.40 0.56 Library B Fr G+C 35–40% 0.67 0.82 0.44 0.64

Library B Fr G+C 40–45% 0.72 0.75 0.45 0.51 Library B Fr G+C 45–50% 0.62 0.63 0.33 0.40 Library B Fr G+C 50–55% 0.34 0.64 0.20 0.40 Library B Fr G+C 55–60% 0.18 0.33 0.13 0.34 Library B Fr G+C 60–65% 0.44 0.32 0.17 0.36 Library B Fr G+C 65–70% 0.68 0.53 0.39 0.39 Library B Fr G+C 70–75% 0.69 0.67 0.42 Vadimezan 0.47 Library B Fr G+C 25–75%e 0.92 0.60 0.81 0.26 a. Fraction of sequences observed in shared OTUs in library A b. Fraction of sequences observed in shared OTUs in library B c. Fraction of shared OTUs in library A d. Fraction of shared OTUs in library B e. The combined

G+C fractions Shannon entropies of clone libraries of the %G+C profiled sample The %G+C fractions 50–55 and 55–60 had comparatively low Shannon entropies (Additional file 2), indicating lower diversity, and were abundant with bifidobacteria (Figure 2, Additional file 1). The peripheral %G+C fractions and the %G+C fraction 45–50 with sequences affiliating mainly with Clostridium clusters IV and XIV had comparatively higher diversity according to Shannon entropies. The peripheral fraction from the low %G+C end (25–30% G+C content) contained a substantial proportion of Firmicutes that do not belong to the Clostridum clusters IV and XIV. It had the highest Shannon entropy (Additional file 2), indicating rich diversity, and did Niclosamide not reach a plateau in the rarefaction curves (data not shown), which means that more OTUs would have been likely to appear after further sequencing. Discussion For a comprehensive evaluation of the human intestinal microbiota, 16S rRNA gene clone libraries were constructed from a %G+C fractioned pooled faecal DNA sample of 23 healthy subjects followed by a sequence analysis of 3199 clones. Previously, only selected fractions of such profiles have been sequenced and analysed.