Friday, July 9, 2010

When is “direct PCR” not direct?

We’ve seen a few companies promoting “direct PCR” products, where a sample can be added directly to a PCR mixture without first extracting DNA. This seems like a good idea in theory, but the reality is not as simple--especially for samples that contain PCR inhibitors.

Therefore, companies that offer kits for direct PCR have had to lengthen these procedures to contend with inhibitors. For example, one kit requires NaOH treatment for 10 minutes, followed by neutralization with Tris-HCl. A large PCR volume is needed and an additive must be included in the gel loading buffer prior to electrophoresis. A second direct PCR kit also requires a large PCR volume and an additive in the loading buffer. In addition, since the polymerase is not Taq-based, annealing temperature must be carefully optimized. For this kit, alternate protocols are offered for challenging templates or multiplex PCR consisting of adding Dilution Buffer and Additive, followed by vortexing, centrifugation, two incubations, and a second centrifugation.

Both kit protocols described above are based on processing animal tissue samples. For plant tissue, even more steps are needed to remove inhibitors.

In contrast, Epicentre’s QuickExtract™ products offer a rapid, efficient method to extract nucleic acids from virtually any sample for PCR-based assays, with most samples being processed in only 8 minutes. Our guaranteed FailSafe™ PCR System ensures reliable PCR results, the first time and every time. Together, these products provide a simpler and more effective method than direct PCR.

Special offer: Order the FailSafe PCR PreMix Selection Kit and get 5 ml of any QuickExtract product free.

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