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Methods

Use of Stable Isotopes To Measure the Metabolic Activity of the Human Intestinal Microbiota

Nicole Reichardt, Andrew R. Barclay, Lawrence T. Weaver, Douglas J. Morrison
Nicole Reichardt
1Section of Child Health, Division of Developmental Medicine, University of Glasgow, Yorkhill Hospitals, Glasgow G3 8SJ
2Stable Isotope Biochemistry Laboratory, Scottish Universities Environmental Research Centre, East Kilbride, Glasgow G75 0QF, United Kingdom
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Andrew R. Barclay
1Section of Child Health, Division of Developmental Medicine, University of Glasgow, Yorkhill Hospitals, Glasgow G3 8SJ
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Lawrence T. Weaver
1Section of Child Health, Division of Developmental Medicine, University of Glasgow, Yorkhill Hospitals, Glasgow G3 8SJ
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Douglas J. Morrison
2Stable Isotope Biochemistry Laboratory, Scottish Universities Environmental Research Centre, East Kilbride, Glasgow G75 0QF, United Kingdom
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  • For correspondence: douglas.morrison@glasgow.ac.uk
DOI: 10.1128/AEM.05573-11
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    Fig. 1.

    Origin of C atoms in purine and pyrimidine de novo synthesis and 13C labeling by use of [13C]urea as the tracer substance.

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    Fig. 2.

    13C enrichment of the 16S rRNA of total bacteria (measured using probe EUB338) after 24 h of incubation of 1% human fecal sample with either [13C]glycine (1.5 mg/ml), NaH13CO3 (0.679 mg/ml), or [13C]urea (0.939 mg/ml) in mineral medium with 10 g/liter oligofructose. Values are given as means ± standard deviations; **, P < 0.01.

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    Fig. 3.

    13C enrichment of the 16S rRNAs of total bacteria (A), the Bacteroides-Prevotella group (B), bifidobacteria (C), the Eubacterium rectale-Clostridium coccoides group (D), and the Clostridium leptum subgroup (E) after 6 h of incubation of 1% human fecal sample with [13C]urea (0.939 mg/ml) in mineral medium without oligofructose (MM − OF) or with 10 g/liter oligofructose (MM + OF) and in Wilkins-Chalgren broth without oligofructose (WC − OF) or with 10 g/liter oligofructose (WC + OF). **, P < 0.01 versus the control (MM − OF); a, P < 0.05 versus MM + OF.

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  • Table 1.

    Biotinylated oligonucleotide probes targeting 16S rRNA and hybridization temperatures

    Probe nameSequence (5′–3′)Target organismsHybridization temp (°C)Reference
    EUB338Biotin-GCTGCCTCCCGTAGGAGTMost eubacteria461
    Bac303Biotin-CCAATGTGGGGGACCTTBacteroides-Prevotella group4617
    Bif164Biotin-CATCCGGCATTACCACCCBifidobacteria4513
    EREC482Biotin-GCTTCTTAGTCARGTACCGEubacterium rectale-Clostridium coccoides group508
    Clep866Biotin-GGTGGATWACTTATTGTGClostridium leptum subgroup4014
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Use of Stable Isotopes To Measure the Metabolic Activity of the Human Intestinal Microbiota
Nicole Reichardt, Andrew R. Barclay, Lawrence T. Weaver, Douglas J. Morrison
Applied and Environmental Microbiology Nov 2011, 77 (22) 8009-8014; DOI: 10.1128/AEM.05573-11

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Use of Stable Isotopes To Measure the Metabolic Activity of the Human Intestinal Microbiota
Nicole Reichardt, Andrew R. Barclay, Lawrence T. Weaver, Douglas J. Morrison
Applied and Environmental Microbiology Nov 2011, 77 (22) 8009-8014; DOI: 10.1128/AEM.05573-11
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