Consider upgrading to BlazeTaq™ SYBR Green qPCR Mix 2.0 for higher amplification efficiency.
- Uniform reaction conditions reduce experimental design time enabling earlier journal submissions
- High amplification efficiency and sensitivity even for low-copy genes means reliable quantitation every time
- Absence of non-specific amplification* and no primer-dimers* ensures reproducible and ready-to-publish data
Figure 2. The amplification efficiency and detection sensitivity of the 2X All-in-One™ qPCR Mix are assessed by standard curves made by gradient dilution of plasmid DNA from 5×106 to 5 molecules. The peak values from amplification and melting curves show that very high sensitivity can be obtained using All-in-One™ qPCR Mix which can detect as low as 5 molecules. At the same time, high amplification efficiency has also been shown by a good linear relationship among each concentration.
When used in combination with the All-in-One SYBR® Green qPCR Mix, the All-in-One primers deliver reliable and reproducible high performance in quantitative PCR assays.
|Agarose Gel Electrophoresis Validation Result
Figure 3. Forty-five pairs of gene-specific All-in-One™ qPCR primers were experimentally validated to yield a single dissociation curve peak and to generate a single amplification of the correct size for the targeted genes. A cDNA pool, containing reverse transcribed products of total RNA from 10 different human tissue (lung, liver, testicle, ovary, spleen, brain, placenta, pancreas, heart and mammary), was used as the qPCR validation template. qPCR was performed using 0.2 µM primer with 2X All-in-One qPCR Mix. Reactions were incubated for 10 min. at 95°C, followed by 40 cycles of 95°C for 10 sec.; 60°C, 20 sec. and 72°C, 15 sec. using Bio-Rad iQ5 Instrument. At the end of the last cycle, the temperature was increased from 72 to 95°C to produce a melting curve. Amplification curves, melting curves and agarose gel electrophoresis (validation result for positive in odd-lane and no template control (NTC) in even lane) shown for the 10 samples.* Non-specific amplification and absence of primer-dimers are ensured when All-in-One validated PCR primers and PCR mix are used together. † Primers validated automatically for human, mouse and rat. Inquire for validation of other species.