ROX Reference Dye

ROX Reference Dye provides users of the ABI PRISM® 7700 an effective alternative for normalization of fluorescent reporter signal in real-time quantitative PCR or RT-PCR without modifying the instrument’s default analysis parameters. ROX Reference Dye is supplied at 50X concentration. It is composed of a glycine conjugate of 5-carboxy-X-rhodamine, succinimidyl ester (25 μM) in 20 mM Tris- HCl (pH 8.4), 0.1 mM EDTA, 0.01% Tween® 20. It may be used with Platinum® Taq DNA polymerase, Platinum® Quantitative PCR SuperMix-UDG, Platinum® Quantitative RT-PCR ThermoScript™ One-Step System, and SuperScript® One-Step RT-PCR with Platinum® Taq.

ROX Reference Dye should be stored in the dark at either −30°C to −10°C or 2°C to 8°C. Storage at 2°C to 8°C avoids the necessity of thawing the solution before assembling the PCR.

ROX Reference Dye is stable at room temperature for several weeks.

Protocol

Set up quantitative PCR / RT-PCR reactions according to the protocol provided with each PCR reagent system. Multiple reactions MUST be assembled as a master mixture.

Include 1 μL of ROX Reference Dye for each 50-μL PCR. Adjust volumes as appropriate to account for addition of the reference dye. Mix thoroughly prior to addition to the master mixture.