محصولات مرتبط
Introduction
The RealQ Plus 2x Master Mix Green with high ROX™ is a singletube 2x reagent including all components necessary to perform real-time DNA amplification for DNA-binding dye based PCR. Just add your primers and DNA. The ROX™ internal reference dye level is optimized for real-time instruments that require high ROX™ as internal reference dye. See instrument capability
Detection limit of RealQ Plus Green with high ROX™ is approximately 1 copy. Quantification limit is approximately 24 copies (0.08 ng of human gDNA, correlating to 12 diploid genomes, with 2 gene copy per diploid genome)
. Real-time PCR is an important tool for SNP and gene expression analysis
Composition of RealQ Plus 2x Master Mix Green, High ROX
TEMPase Hot Start DNA Polymerase
Optimized buffer system including dNTPs, fluorescent dye and ROX™ reference dye
Recommended Storage and stability
Long term storage at -20 °C. Product expiry at -20 °C is stated on the label
Option: Store at +4 °C for up to 3 months
Quality Control
TEMPase Hot Start DNA Polymerase is tested for contaminating activities, with no traces of endonuclease activity, nicking activity or exonuclease activity. The RealQ Plus 2x Master Mix Green with high ROX™ is functionally tested for efficiency and absence of contaminating human genomic DNA
Pre-protocol Considerations
ROX™ Reference Dye
ROX™ is used as passive reference dye to compensate for nonPCR related variations in the fluorescence. The ROX™ fluorescence does not change during the course of the PCR reaction nor does it influence the PCR reaction. It provides a stable baseline to which samples are normalized
PCR Primers
It is important - especially in fluorescent DNA dye based quantitative PCR applications - to minimize the formation of non-specific amplification products. Particularly at low target concentration it is important to use the lowest possible primer concentration without compromising the efficiency of the PCR. The optimal concentration of primer pairs is the lowest concentration that results in the lowest Cq and an adequate fluorescence for a given target concentration with minimal or no formation of primer-dimers. The optimal concentrations of upstream and downstream primers are not always of equal molarity. Optimal concentrations of primers are in the range of 100 nM to 800 nM
Preventing Template Cross-Contamination
Due to the high sensitivity of quantitative PCR there is a risk of contaminating the reactions with the products of previous runs. To minimize this risk, tubes or plates containing reaction products should not be opened or analysed by gel electrophoresis in the same laboratory area used to set up reactions
Instrument compatibility
Real-time instruments which require high ROX™ such as: Applied Biosystems® 5700, 7000, 7300, 7700, 7900, 7900 HT, StepOne™ and StepOnePlus
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