ROX Reference Dye is used to normalize real-time RT-PCR and PCR assays. It provides an internal reference to which the reporter dye signal can be normalized. This signal normalization is necessary to correct for well-to-well signal fluctuations, often due to the design of the instrument.
ROX Reference Dye is particularly recommended for use in combination with FastStart TaqMan Probe Master and FastStart SYBR Green Master on real-time PCR instruments from Applied Biosystems (ABI), in order to readily use their default parameter analysis. These instruments are available equipped with either a Halogen lamp-based light source (ABI PRISM 7000 Sequence Detection System, ABI 7300/7500 Real-Time PCR Systems) or a Laser-based light source (ABI PRISM 7700 Sequence Detection System, ABI 7900HT Real-Time PCR System). While both FastStart TaqMan Probe Master (Rox) and FastStart SYBR Green Master (Rox) can be readily used for the ABI 7500 System, it is necessary to increase the ROX concentration to 300 - 600 nM for the ABI 7000 System, the ABI 7300 System, the ABI 7700 System, and the ABI 7900HT System.
ROX Reference Dye from Roche Applied Science is supplied in a “non-reactive” form, i.e., it is not able to react with e.g., amino groups of proteins like the ROX formulations supplied by other providers. In addition, it is proven not to inhibit PCR at a final concentration of up to 1000 nM. ROX Reference Dye is not required when using the LightCycler Instruments.
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