DNA was extracted from fecal samples or fecal donor suspensions as described previously [27 (link), 31 (link)]. In brief, DNA was quantified by using Quant-iT PicoGreen reagent (Invitrogen, UK) and adjusted to 1 ng per μL. Then, the total eubacterial loads, as well as the main bacterial groups abundant in the murine and human intestinal microbiota including enterobacteria, enterococci, lactobacilli, bifidobacteria, Bacteroides/Prevotella species, Clostridium coccoides group, Clostridium leptum group, and Mouse Intestnal Bacteroides, were assessed by quantitative real-time polymerase chain reaction (qRT-PCR) with species-, genus-, or group-specific 16S rRNA gene primers (Tib MolBiol, Germany) as described previously [10 (link), 30 (link), 32 (link)] and numbers of 16S rRNA gene copies per ng DNA of each sample determined.
Protocol detail
Quantification of Intestinal Microbiota
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16s rrna
Bacterial
Bacteroides
Bifidobacteria
Clostridium
Clostridium coccoides
Donor
Enterobacteria
Enterococci
Eubacterial
Fecal
Gene
Human
Intestinal microbiota
Lactobacilli
Mouse
Murine
Picogreen
Prevotella
Primers
Quantitative real time polymerase chain reaction
Rrna gene
Corresponding Organization : Charité - Universitätsmedizin Berlin
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Variable analysis
independent variables
- None explicitly mentioned
- Total eubacterial loads
- Abundance of bacterial groups (enterobacteria, enterococci, lactobacilli, bifidobacteria, Bacteroides/Prevotella species, Clostridium coccoides group, Clostridium leptum group, Mouse Intestinal Bacteroides)
- DNA concentration (adjusted to 1 ng per μL)
- Positive control: Not specified
- Negative control: Not specified
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