Automated DNA Extraction for Real-Time PCR

We evaluated the MagNA Pure LC System (Roche Applied Science) for automated DNA extraction for use in mutation detection and allele quantification. The system uses magnetic silica beads that bind DNA to a silica surface and transfer DNA through various steps of the extraction process. This system can be linked to LightCycler assays for further automation. The LightCycler is an enclosed thermocycler with continuous monitoring for PCR quantification and allele discrimination (1)(2)(3). Combined MagNA Pure/LightCycler automation of sample preparation, amplification, and detection has the potential to reduce human error in sample tracking and to standardize DNA extraction and inoculation into the PCR mixture. The MagNA Pure/LightCycler system has been evaluated for infectious disease applications, detecting PCR product by crossing point (4)(5). In this study we compared the MagNA Pure LC DNA extraction with the Qiagen 9604 BioRobot (Qiagen) for real-time mutation detection and allele quantification. We evaluated 280 samples with the MagNA Pure System and Qiagen 9604 BioRobot under an Internal Review Board-approved protocol for instrument validation. Genomic DNA was extracted from 200 μL of EDTA-anticoagulated whole blood by use of the MagNA Pure LC DNA Isolation Kit I (Roche Diagnostics), with its high-performance DNA protocol, and by the QIAamp 9604 DNA Blood BioRobot Kit (Qiagen) according to manufacturer’s instructions. Both methods initially use red blood cell lysis and a proteinase K procedure, and the DNA binds to a silica-based support. The MagNA Pure separates contaminants from DNA by use of magnetic beads, whereas the 9604 BioRobot uses a vacuum manifold.