DNA Extraction Systems Compared for Stool Samples

Five different DNA extractions systems have been tested and compared for fecal sample collections.

The purified DNA was analyzed by polymerase chain reaction (PCR) directed toward the relevant bacterial organisms present in the stool samples.

In a study carried out at the Statens Serum Institute, (Copenhagen, Denmark), five commercial DNA extraction systems were tested and compared on 81 clinical stool samples that were culture positive. The DNA was analyzed by PCR directed at the following organisms: diarrheagenic Escherichia coli, Campylobacter jejuni, Salmonella enterica, or Clostridium difficile.

The following five commercial DNA extraction systems were included in the study: EasyMag and BioRobot EZ1 are semiautomatic systems based on bacterial lysis followed by DNA extraction by magnetic beads with bound silica particles; Bugs'n Beads is a manual procedure that first isolates whole bacteria, by specific affinity of coated magnetic beads, followed by lysis and DNA extraction on the same beads; QIAamp DNA Stool Kit is a manual procedure that extracts DNA from lysed bacteria on spin columns with bound silica; and ChargeSwitch is a manual procedure that extracts DNA from lysed bacteria by magnetic particles coated with the ChargeSwitch matrix.

The results showed that conventional multiplex PCR combined with the extraction systems EasyMag Specific A, (bioMérieux; Marcy I'Etoile, France), BioRobot EZ1and QIAamp (Qiagen; Hilden, Germany), ChargeSwitch (Invitrogen; Carlsbad, CA, USA), EasyMag generic (bioMérieux), and Bugs'n Beads, (Genpoint; Oslo, Norway), were able to identify, 91%, 89%, 88%, 85%, 85% and 62% respectively, of the pathogens originally identified by conventional culture-based methods. When TaqMan mPCR was combined with the EasyMag Specific A protocol, 99% of the samples were correctly identified.

The results demonstrate that the extraction efficiencies can vary significantly among different extraction systems, careful optimization may have a significant positive effect, and the use of sensitive and specific detection methods like TaqMan PCR is an ideal choice for this type of analysis. The study was published in March 2011, in Diagnostic Microbiology and Infectious Disease.

Related Links:
Statens Serum Institute
bioMérieux
Qiagen
Invitrogen
Genpoint