RNA to Real-Time PCR Results

Whether you use TaqMan® or SYBR® Green for real-time PCR, Applied Biosystems newest suite of offerings minimizes hands-on time and reduces error with one-step, single-tube protocols for reverse transcription and PCR.

NEW TaqMan® RNA-to-CT™ 1-Step Kit – From Good to Great!

The new TaqMan® RNA-to-CT 1-Step Kit shows improved sensitivity, efficiency, and dynamic range in real-time RT-PCR relative to TaqMan One-Step RT-PCR Master Mix Reagents. The dramatically improved performance of this new formulation provides results that are comparable to those from the TaqMan RNA-to-CT 2-Step Kit. These improvements hold true for a wide array of TaqMan Assays, including assays for challenging targets such as AT-rich, GC-rich, and long amplicons.

Consistent RNA Target Quantification

  • Accurate across a wide dynamic range
  • Sensitive low copy number detection and precise quantitation
  • Consistent results even with AT-rich, GC-rich and long amplicons
  • Fewer pipetting steps results in fewer errors
  • Validated with TaqMan Gene Expression Assays

Consistent Performance Improvement Over TaqMan One-Step RT-PCR Master Mix

Figure 1 illustrates the consistent performance of the new TaqMan RNA-to-CT 1-Step Kit with input RNA amounts covering a 6-log range compared to only a 4-log dynamic range with the TaqMan One-Step RT-PCR Master Mix Reagents. These data also show that PCR efficiency is not compromised compared to reactions run using the more traditional two-step RT-PCR strategy. Figure 2 shows the performance of the TaqMan RNA-to-CT 1-Step Kit in challenging reactions with long amplicons (Figure 2A), AT-rich targets (Figure 2B), and GC-rich targets (Figure 2C).



Figure 1. Dynamic Range and Efficiency of the TaqMan® RNA-to-CT™ 1-Step and 2-Step Kits vs. TaqMan One-Step RT-PCR Master Mix Reagents. The TaqMan® RNA-to-CT 1-Step Kit (blue) shows a wider dynamic range and improved sensitivity relative to the TaqMan One-Step RT-PCR Master Mix Reagents (orange) and comparable performance to the TaqMan RNA-to-CT 2-Step Kit (red). Amplifications are from a serial dilution of PPIA target using 100 ng to 0.1 pg of RNA input on the Applied Biosystems 7900HT Fast Real-Time PCR System.



Figure 2. Robust Amplification of Challenging Targets with the TaqMan® RNA-to-CT™1-Step Kit. Amplification plots are from serial dilutions using the TaqMan® RNA-to-CT™ 1-Step Kit on the Applied Biosystems 7900HT Fast Real-Time PCR System.


Power SYBR® Green RNA-to-CT 1-Step Kit— Industry-Leading Sensitivity and Specificity

For SYBR Green-based real-time RT-PCR, the Power SYBR Green RNA-to-CT 1-Step Kit is an easy-to-use, single-tube solution for reliable, high-quality real-time PCR data. A novel formulation helps prevent spurious results caused by primer dimers and enables reliable detection of targets at lower concentrations than other one-step kits using SYBR green-based detection chemistry.

  • Accurate across a wide dynamic range
  • Single tube reduces error and contamination caused by multiple pipetting steps
  • Highly specific and consistent results, even at low input amounts of target RNA



Figure 3. Power SYBR® Green RNA-to-CT™ 1-Step Kit Shows Better Efficiency, Precision, and Dynamic Range than Similar SYBR Green Products from Other Suppliers. RNA was isolated from BOSC23 cultured cells and amplified in three replicates on the Applied Biosystems StepOnePlus™ Real-Time PCR System using either Applied Biosystems Power SYBR Green RNA-to-CT 1-Step Kit and a one-step qRT-PCR kit from another supplier. The Power SYBR Green RNA-to-CT 1-Step Kit provides excellent PCR efficiency (>99%) and precise quantification over a 6-log range of input RNA quantities (100 fg to 100 ng). In comparison, the competitor kit shows inhibition of PCR at high quantities of target and nonspecific amplification at low quantities of target.


Scientific Contributors:

 Jennifer Berkman, Nathalie Koch,Yong Lee • Applied Biosystems Inc., Foster City, CA
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