Sensitive and Efficient PCR

Amplification of a plasmid dilution series demonstrates that the sensitivity and PCR efficiency (99.7%) of Fast SYBR Green Master Mix in fast mode is equivalent to Power SYBR Green PCR Master Mix in standard mode on the Applied Biosystems StepOnePlus™ Real-Time PCR System [1]. In addition, the Fast SYBR Green Master Mix outperformed other commercially available master mixes, giving higher PCR efficiency.

The presence of nonspecific PCR products and primer-dimers can reduce amplification efficiency, limit dynamic range, and ultimately affect the accuracy of the data. Fast SYBR Green Master Mix was formulated to minimize primer dimerization and nonspecific amplification, and to optimize the efficiency and accuracy of the reaction without compromising sensitivity (Figure 1).



RT-PCR products amplified with PGRMC1-specific primers and either the Fast SYBR Green Master Mix or one of two other commercial master mixes were compared. Only Fast SYBR Green Master Mix demonstrated target specific amplification using as little as 0.1 pg cDNA with no primer-dimer formation or nonspecific amplification (data not shown).

The high sensitivity provided by Fast SYBR Green Master Mix facilitates quantifying small differences in target amount between samples. An amplification plot of PCR targeting the RNase P gene in genomic DNA shows statistically significant discrimination down to a 1.33-fold difference between samples, with 99.7% confidence (Figure 1).

Applied Biosystems Fast SYBR Green Master Mix and Fast Real-Time PCR Systems enable researchers to increase sample throughput by decreasing thermal cycling times to as little as 35 min/run. The Fast SYBR Green Master Mix maintains the same high quality of real-time PCR analysis: sensitivity, specificity, and dynamic range of PCR amplification are comparable to reactions performed in standard thermal cycling mode.