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Real-Time qPCR & Reverse Transcription

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Fast Real-Time PCR


Fast Real-time PCR

About the Figure: Turning waste into H2 biofuel. Researchers assessed the microbiome of waste activated sludge generated at water treatment facilities (shown in color in the figure above) using pyrosequencing, then used Premix Ex Taq™ (Probe qPCR) on an ABI 7500 FAST system to identify methanogens by fast real-time PCR. Nakazawa, T., et al. (2011) Fungal Genetics Biol. 48:939.

Less waiting for results...without sacrificing data quality

Whether you require a high-throughput workflow, need results faster, or just want to get more experiments done in a day, Takara® master mixes can help you accomplish fast real-time PCR by reducing your run times. Takara premixes for SYBR® and TaqMan® detection:

• Are compatible with fast real-time protocols and instruments, including the Applied Biosystems 7500 Fast Real-Time PCR System
• Provide high assay sensitivity, specificity, and efficiency
• Provide consistent and reproducible results
 Try a sample of SapphireAmp Fast PCR Master Mix

Experimental Example: Using Premix Ex Taq (Probe qPCR) for Fast Real-Time PCR

PCR efficiency

Figure 1. PCR efficiency using Premix Ex Taq (Probe qPCR). A portion of the GAPDH gene was amplified by real-time PCR from cDNA synthesized using 1 pg to 100 ng of total RNA. The standard curves are plotted.
  Cut extension times from 30 seconds down to 20 seconds with no reduction in reaction efficiency when using Premix Ex Taq (Probe qPCR) for TaqMan® probe detection. For maximal time savings, the master mix can be used with fast mode thermal cycling conditions.
In Figure 1, reaction efficiency was assessed using various extension times. The same efficiency was obtained with extension times of 30, 25, and 20 seconds using the Applied Biosystems 7500 Fast Real-Time PCR System (top) and extension times of 30 and 20 seconds on the Roche LightCycler® 480 (bottom).


Experimental Example: Using SYBR® Premix Ex Taq (Tli RNase H Plus) for Fast Real-Time PCR

amplification curves

Figure 2. Fast real-time PCR measurement of OGT gene expression. OGT was measured from cDNA synthesized from cardiac progenitor cells by real-time PCR. Amplification products were measured using SYBR® Green detection with either SYBR® Premix Ex Taq (Tli RNase H Plus) (purple) or a fast SYBR® master mix from another manufacturer (blue) using a 20 second extension step.
  Takara SYBR® master mixes are also compatible with fast cycling protocols. In Figure 2, real-time PCR measurement of OGT gene expression was performed with a 7900HT Fast Real-Time PCR System (Applied Biosystems) using a 20
second extension step. Takara SYBR® Premix Ex Taq provided similar results to another manufacturer's fast SYBR® Green master mix. Moreover, SYBR® Premix Ex Taq provided consistent, reproducible results. For additional experimental details, download the entire application note.


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Real-time PCR overview

Application Note

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