TY - JOUR
T1 - Microbial diversity in fecal samples depends on DNA extraction method
T2 - EasyMag DNA extraction compared to QIAamp DNA stool mini kit extraction
AU - Mirsepasi, Hengameh
AU - Persson, Søren
AU - Struve, Carsten
AU - Andersen, Lee O B
AU - Petersen, Andreas M.
AU - Krogfelt, Karen A.
PY - 2014
Y1 - 2014
N2 - Background: There are challenges, when extracting bacterial DNA from specimens for molecular diagnostics, since fecal samples also contain DNA from human cells and many different substances derived from food, cell residues and medication that can inhibit downstream PCR. The purpose of the study was to evaluate two different DNA extraction methods in order to choose the most efficient method for studying intestinal bacterial diversity using Denaturing Gradient Gel Electrophoresis (DGGE). Findings. In this study, a semi-automatic DNA extraction system (easyMag®, BioMérieux, Marcy I'Etoile, France) and a manual one (QIAamp DNA Stool Mini Kit, Qiagen, Hilden, Germany) were tested on stool samples collected from 3 patients with Inflammatory Bowel disease (IBD) and 5 healthy individuals.DNA extracts obtained by the QIAamp DNA Stool Mini Kit yield a higher amount of DNA compared to DNA extracts obtained by easyMag® from the same fecal samples. Furthermore, DNA extracts obtained using easyMag® seemed to contain inhibitory compounds, since in order to perform a successful PCR-analysis, the sample should be diluted at least 10 times. DGGE performed on PCR from DNA extracted by QIAamp DNA Stool Mini Kit DNA was very successful. Conclusion: QIAamp DNA Stool Mini Kit DNA extracts are optimal for DGGE runs and this extraction method yields a higher amount of DNA compared to easyMag®.
AB - Background: There are challenges, when extracting bacterial DNA from specimens for molecular diagnostics, since fecal samples also contain DNA from human cells and many different substances derived from food, cell residues and medication that can inhibit downstream PCR. The purpose of the study was to evaluate two different DNA extraction methods in order to choose the most efficient method for studying intestinal bacterial diversity using Denaturing Gradient Gel Electrophoresis (DGGE). Findings. In this study, a semi-automatic DNA extraction system (easyMag®, BioMérieux, Marcy I'Etoile, France) and a manual one (QIAamp DNA Stool Mini Kit, Qiagen, Hilden, Germany) were tested on stool samples collected from 3 patients with Inflammatory Bowel disease (IBD) and 5 healthy individuals.DNA extracts obtained by the QIAamp DNA Stool Mini Kit yield a higher amount of DNA compared to DNA extracts obtained by easyMag® from the same fecal samples. Furthermore, DNA extracts obtained using easyMag® seemed to contain inhibitory compounds, since in order to perform a successful PCR-analysis, the sample should be diluted at least 10 times. DGGE performed on PCR from DNA extracted by QIAamp DNA Stool Mini Kit DNA was very successful. Conclusion: QIAamp DNA Stool Mini Kit DNA extracts are optimal for DGGE runs and this extraction method yields a higher amount of DNA compared to easyMag®.
KW - Denaturing gradient gel electrophoresis
KW - DNA measurement
KW - easyMag®
KW - Fecal DNA extraction
KW - Inflammatory bowel disease
KW - NanoDrop®
KW - QIAamp DNA stool mini kit
KW - Qubit® system
U2 - 10.1186/1756-0500-7-50
DO - 10.1186/1756-0500-7-50
M3 - Journal article
C2 - 24447346
AN - SCOPUS:84892586947
SN - 1756-0500
VL - 7
JO - BMC Research Notes
JF - BMC Research Notes
IS - 1
M1 - 50
ER -
