Comparison of five assays for DNA extraction from bacterial cells in human faecal samples
Tessa Gryp, Griet Glorieux, Marie Joossens, Mario Vaneechoutte
Abstract
AIM: To determine the most effective DNA extraction method for bacteria in faecal samples. MATERIALS AND RESULTS: This study assessed five commercial methods, that is, NucliSens easyMag, QIAamp DNA Stool Mini kit, PureLink Microbiome DNA purification kit, QIAamp PowerFecal DNA kit and RNeasy PowerMicrobiome kit, of which the latter has been optimized for DNA extraction. The DNA quantity and quality were determined using Nanodrop, Qubit and qPCR. The PowerMicrobiome kit recovered the highest DNA concentration, whereby this kit also recovered the highest gene copy number of Gram positives, Gram negatives and total bacteria. Furthermore, the PowerMicrobiome kit in combination with mechanical pre-treatment (bead beating) and with combined enzymatic and mechanical pre-treatment (proteinase K+mutanolysin+bead beating) was more effective than without pre-treatment. CONCLUSION: From the five DNA extraction methods that were compared, the PowerMicrobiome kit, preceded by bead beating, which is standard included, was found to be the most effective DNA extraction method for bacteria in faecal samples. SIGNIFICANCE AND IMPACT OF THE STUDY: The quantity and quality of DNA extracted from human faecal samples is a first important step to optimize molecular methods. Here we have shown that the PowerMicrobiome kit is an effective DNA extraction method for bacterial cells in faecal samples for downstream qPCR purpose.