Abstract
BackgroundDNA extraction is an important factor influencing the microbiome profile in fecal samples. Considering that the QIAamp DNA Stool Mini Kit, one of the most commonly used DNA extraction kits, is no longer manufactured, this study aimed to investigate whether a new commercially available kit, the QIAamp PowerFecal Pro DNA Kit, yields comparable microbiome profiles with those previously obtained using the QIAamp DNA Stool Mini Kit.ResultsWe extracted DNA from fecal samples of 10 individuals using three protocols (protocol P of the QIAamp PowerFecal Pro DNA Kit, and protocols SB and S of the QIAamp DNA Stool Mini Kit with and without an additional bead-beating step, respectively) in triplicate. Ninety extracted DNA samples were subjected to 16S rRNA gene sequencing. DNA quality measured by 260/280 absorbance ratios was found to be optimal in protocol P. Additionally, the DNA quantity and microbiome diversity obtained using protocol P were significantly higher than those of protocol S, however, did not differ significantly from those of protocol SB. Based on the overall microbiome profiles, variations between protocol P and protocol SB or S were significantly less than between-individual variations. Furthermore, most genera were not differentially abundant in protocol P compared to the other protocols, and the number of differentially abundant genera, as well as the degree of fold-changes were smaller between protocols P and SB than between protocols P and S.ConclusionsThe QIAamp PowerFecal Pro DNA Kit exhibited microbiome analysis results that were comparable with those of the QIAamp DNA Stool Mini Kit with a bead-beating step. These results will prove useful for researchers investigating the gut microbiome in selecting an alternative protocol to the widely used but discontinued kit.
Highlights
DNA extraction is an important factor influencing the microbiome profile in fecal samples
This study investigated whether the QIAamp PowerFecal Pro DNA Kit performed to the QIAamp DNA Stool Mini Kit with or without an additional bead-beating step in gut microbiome analysis
This study shows that the degree of microbiome variations caused by differences in the tested DNA extraction protocols was significantly less than that caused by inter-individual differences
Summary
DNA extraction is an important factor influencing the microbiome profile in fecal samples. The QIAamp PowerFecal Pro DNA Kit, recently manufactured by Qiagen, may prove useful owing to its improved technology for the efficient elimination of inhibitors in fecal samples, as well as the application of a novel bead tube to facilitate more efficient bacterial lysis [13]. This kit is compatible with an existing automated sample preparation instrument, QIAcube, and the downstream processes following bead-beating can be automated using the QIAcube. Before introducing a new protocol for the primary experiments, it is important, to determine whether the new protocol would be an appropriate replacement for gut microbiome studies in comparison with the established kits and protocols
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