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qPCR 2x Blue Master Mix + ROX

Description

This product is similar to qPCR 2x Blue Master Mix, but it is supplemented with ROX (6-carboxy-X-rhodamine) reference dye. ROX serves as an internal reference for normalization of the fluorescent signal when using real-time PCR instruments, which can detect ROX. ROX allows for correction of the well-to-well variation due to pipetting inaccuracies and fluorescence fluctuations. The presence of ROX does not interfere with qPCR amplification (G. Wang, E. Becker, C. Mesa, Can. J. Microbiol. 53, 391-397, 2007). ROX has different emission spectrum (the excitation/emission maxima are at 586 nm/605 nm, respectively) when compared to SYBR Green I and other probes used for qPCR analyses.

Rapid preparation (2x concentrated)

• All components of the qPCR 2x Blue Master Mix + ROX are 2x concentrated (optimized reaction buffer, nucleotides, Taq DNA polymerase, anti-Taq monoclonal antibody, additives, and stabilizers). This facilitates rapid preparation of the PCRs. The samples are prepared simply by mixing an aliquot of the Mix with oligonucleotide primers, selected DNA probes, template DNA, and PCR H₂O (included).

Hot start

• The product contains monoclonal antibody anti-Taq, which binds to Taq DNA polymerase and thus inactivates its enzymatic activity. After the first denaturation cycle, the antibody is irreversibly inactivated and Taq DNA polymerase regains enzymatic activity. This decreases formation of nonspecific DNA amplicons.

ROX Reference Dye ROX in qPCR mixes is used in two concentrations, depending on instrument used.

qPCR 2x Blue Master Mix + ROX 500 - Final concentration 500 nM when using Applied Biosystems 7000, 7300, 7700, 7900HT, StepOne, StepOnePlus and othr PCR machines.

qPCR 2x Blue Master Mix + ROX 50 - Final concentration 50 nM when using Applied Biosystems 7500, 7500Fast, Stratagene cyclers and other machines..

Sensitive detection

• This product is optimized for sensitive detection of DNA fragments amplified during qPCR from genomic DNA or cDNA obtained by reverse transcription.

Technical data

Components and packaging

• 1 tube with 0.5 ml qPCR 2x Blue Master Mix + ROX 500 or ROX 50 (this amount is sufficient e.g. for 40 reactions, 25 µl each).
• 1 tube with 1.5 ml PCR H₂O.

Storage

• At temperature -20^oC ± 5^oC. Material can be repeatedly defrosted.

Composition

• The Mix is 2x concentrated: 150 mM Tris-HCl, pH 8. 8 (at 25oC), 40 mM (NH4)2SO4, 0.02% Tween 20, 5 mM MgCl2, 400 µM dATP, 400 µM dCTP, 400 µM dGTP, 400 µM dTTP, Taq DNA polymerase (50 U/ml), monoclonal antibody anti-Taq (38 nM), ROX (1 µM or 0.1 µM), stabilizers and additives.

Purity and quality control

• Each batch of qPCR 2x Blue Master Mix + ROX is tested for amplification of a single copy gene in genomic DNA.

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