PCR ENZYMES - Real-time qPCR

RealQ Plus 2x Master Mix for Probe

RealQ Plus 2x Master Mix for Probe enables real-time-based DNA amplification with high specificity and efficiency.

Features

Promotes probe-based real-time PCR based detection
Enables multiplexing
Very high specificity
High stability and efficiency
Reliable quantification
Reaction setup at room temperature
Premixed all-in-one 2x solution

DESCRIPTION

RealQ Plus 2x Master Mix for Probe is composed of TEMPase Hot Start DNA Polymerase, dNTPs, dye and an optimized buffer system. Furthermore, this master mix is available with high, low or without ROXensuring optimal performance on most of the commonly used real-time PCR instruments. Just add DNA template, primers and probe.

The RealQ Plus 2x Master Mix for probe is optimized to TaqMan probes, but is also suitable with other probe chemistries such as Molecular Beacon and Scorpion.

Most probe-based detection methods take advantage of fluorescent resonance energy transfer (FRET) by quenching the signal of a fluorescent reporter in the absence of the desired target. During elongation, the quenching molecule is separated from the fluorescent reporter and a signal is emitted and monitored. Probe-based detection is significantly more specific since signal is only detected when the correct target is amplified.

HIGH EFFICIENCY AND PRECISION

Amplification plot of a 4-fold dilution series for a Pthr target (75 bp) amplified from human gDNA, starting with 80 ng down to 80 pg of gDNA, using RealQ Plus 2x Master Mix for Probe. Samples were made in triplicates. The results depicted here, show high precision and efficiencies close to 100 %.

MULTIPLEXING REAL-TIME PCR USING FOUR DNA TARGETS

To examine the multiplexing capacity of the RealQ Plus 2x Master Mix for Probe, a serial dilution of four different human gDNA targets were evaluated.The Pthr1 (FAM), DMD4 (HEX), DMD6 (Texas red) and DMD47 (Cy5) assays are shown in purple, green, pink and orange respectively. Data showing linear amplification plots of a set of four 5-fold serial dilutions of human gDNA ranging from 40 – 0.32 ng per PCR reaction. Data represents triplicates for each DNA dilution. Cycling was performed on a LightCycler® 96 SW 1.1 with the following protocol: 95°C, 15 min; followed by 50 cycles of 95°C, 15 s; 60°C, 60 s

Singleplex Vs Qudroplexvers 1 17122021 Jpg

High efficiency, high sensitivity multiplex real-time PCR results with RealQ Flex for Probe. Serial dilution of four different human gDNA targets were amplified either a single-plex real-time PCR, or a 4-target multiplexed real-time PCR. There is only a minor shift in Cq values for singleplexing vs multingplexing. Cycling was performed on a LightCycler® 96 SW 1.1 with the following protocol: 95°C, 15 min; followed by 50 cycles of 95°C, 15 s; 60°C, 60 s

STABILITY AT ROOM TEMPERATURE

Two plates were pre-assembled for qPCR reaction and incubated in darkness at room temperature for 48 and 72 hours. The results show high stability and complete inactivation of the TEMPase before hot start. The High Stability of the TEMPase Hot Start DNA Polymerase in RealQ master mixes allows the scientist to set up the real-time PCR reaction at room temperature and run the plate several hours later, when convenient.