Will I see a difference in my qPCR data
Normalization of Real-Time PCR Fluorescence Data with ROX Passive Reference Dye
Will I see a difference in my qPCR data if I change from an older real-time PCR reagent that contained a high or low ROX concentration to a Bio-Rad supermix containing a universal reference dye?
Users of Bio-Rad real-time PCR detection systems will not see any significant differences in their qPCR data because the new supermix formulations simply incorporate a universal passive reference dye, which is ignored by our optical detection systems. Users of ROX-dependent real-time PCR systems (for example, Life Technologies' systems) will now be able to use any of our universal real-time PCR reagents without having to determine which product to use.
If you were using an older product with ROX, you may see a slight shift in Cq values of less than ±0.5. However, since all the Cq values are shifted by the same amount, the relative quantitation data remain the same, with no impact on the final results.
See Normalization of Real-Time PCR Fluorescence Data with ROX Passive Reference Dye in our Applications and Technologies pages to learn why passive reference dyes are required for data normalization in some real-time PCR systems and not others. Although Bio-Rad real-time PCR detection systems do not require ROX for internal normalization, Bio-Rad’s universal real-time PCR supermixes are designed for high performance on all real-time PCR detection systems, both systems requiring ROX normalization and ROX-independent systems.
View our Understanding Real-Time Supermixes tutorial to learn more about qPCR data analysis and how to compare supermix performance.