A common practice in real-time PCR is to add a known amount of control DNA after DNA extraction. This monitors PCR inhibition but has no value as an extraction control. The ideal situation is to have the test sample and internal control undergo the same processing prior to real-time PCR (Fig. 1). Bioline have specially developed a DNA Extraction Control (DEC), which more closely mimics the test sample, as compared to spike controls. The genetic material from the test sample and the DEC is simultaneously extracted by common extraction methods, with the extraction control being as sensitive to inhibition and extraction failure as the test sample. The DEC cells are of a known concentration, containing the Internal Control DNA sequence. This sequence contains no known homology to any organism and importantly, has minimal interference with detection of sample DNA. The DEC cells are spiked into lysis buffer with the target sample, prior to DNA extraction. Control Mix (primers and probe) is then added to the reaction mix before amplification. Signal derived from the Internal Control DNA confirms the success of the extraction step (Fig. 2). DEC also monitors co-purification of PCR inhibitors (Fig. 3) that may cause biased or false amplification patterns. DEC offers the quality control assurance required for the entire workflow from sample extraction to real-time amplification and analysis.
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