PrimeTime One-Step 4X Broad-Range Master Mix is a premium one-step qPCR mastermix that offers advanced capabilities including the flexibility to amplify from crude or purified samples.
Features & advantages
- Flexibility to amplify from either purified or crude samples in viral transport media (VTM)
- Excellent Ct and end-point fluorescence performance compared to leading competitors
- Proven performance for singleplex and multiplex (up to 5-plex) probe assays
- Compatible with multiple market-leading qPCR instruments
- Highly inhibitor resistant
Order information
A one-step, 4X concentrated master mix that contains a hot-start reverse transcriptase, hot-start DNA polymerase, dNTPs, MgCl2, enhancers, and stabilizers. Individual tubes of direct amplification enhancer and reference dyes are included.
| Cat-No. | Item | Size | Price (CHF) |
|---|---|---|---|
| 10011744 | PrimeTime 1-Step 4X Broad-Range Master Mix, 1 mL | 1 ml | 450.00 |
| 10011745 | PrimeTime 1-Step 4X Broad-Range Master Mix, 5 mL | 5 ml | 2'405.00 |
| 10011746 | PrimeTime 1-Step 4X Broad-Range Master Mix, 10 mL | 10 ml | 4'575.00 |
| 10011747 | PrimeTime 1-Step 4X Broad-Range Master Mix, 25 mL | 25 ml | price on request |
Product details
The PrimeTime One-Step 4X Broad-Range Master Mix is a ready-to-use, 4X concentrated master mix that is designed for use in probe-based, real-time quantitative PCR (qPCR). PrimeTime One-Step 4X Broad-Range Master Mix contains a mutant hot-start reverse transcriptase, a mutant hot-start DNA polymerase, dNTPs, MgCl2, enhancers, and stabilizers. An enhancer solution is also provided as an optional additive to neutralize PCR inhibitors, including nucleases in crude samples. A reference dye is provided as a separate component, making this Master Mix compatible for use on both reference dye-dependent, and independent, instrument systems.
Product data
Amplify from purified or crude samples in VTM (NP and saliva)
Figure 1. PrimeTime™ One-Step 4X Broad-Range Master Mix demonstrates comparable amplification of viral analytes between extracted and non-extracted human saliva samples. RT-qPCR probe assays containing probes and primers specific for FluA, FluB, SARS-CoV-2, and RNase P were used to amplify 30 copies of AccuPlex™ SARS-CoV-2, Flu A/B and RSV Verification Panel (SARS-CoV-2 data shown) material that had been extracted (dark blue) or direct amplified (light blue). The AccuPlex™ analyte verification panel was first diluted into negative human saliva matrix prior to RNA extraction or direct specimen amplification, and RNA extractions were performed automated using the KingFisher™ Flex 96 and MagMAX™ Viral/Pathogen II (MVP II) Nucleic Acid Isolation Kit. Reactions were amplified and analyzed using the QuantStudio 7 Flex instrument according to the manufacturer's instructions. Experiments were performed with multiple replicates (n = 20) for each condition.
Figure 2. PrimeTime One-Step 4X Broad-Range Master Mix demonstrates comparable amplification of viral analytes between extracted and non-extracted human nasopharyngeal samples. RT-qPCR probe assays containing probes and primers specific for FluA, FluB, SARS-CoV-2, and RNase P were used to amplify 30 copies of AccuPlex™ SARS-CoV-2, Flu A/B and RSV Verification Panel (SARS-CoV-2 data shown) material that had been extracted (dark blue) or direct amplified (light blue). The AccuPlex™ analyte verification panel was first diluted into negative human nasopharyngeal (NP) matrix prior to RNA extraction or direct specimen amplification, and RNA extractions were performed automated using the KingFisher Flex 96 and MagMAX™ Viral/Pathogen II (MVP II) Nucleic Acid Isolation Kit. Reactions were amplified and identified using the QuantStudio 7 Flex instrument according to the manufacturer's instructions. Experiments were performed with multiple replicates (n = 20) for each condition.
Lower Ct and higher end-point fluorescence as compared to other commercially available mixes
Figure 3. PrimeTime One-Step 4X Broad-Range Master Mix provides more complete analyte identification from human saliva and nasopharyngeal matrix as compared to competitors. RT-qPCR probe assays containing probes and primers specific for Influenza A (shown), Influenza B, SARS-CoV-2, and RNaseP genes were used to amplify 30 copies of pooled viral analytes spiked into negative human nasopharyngeal (left) or saliva (right) matrix stored in viral transport medium. RT-qPCR reactions were run on the QuantStudio 7 Flex (n = 6).
5-plex amplification
The PrimeTime One-Step 4X Broad-Range Master Mix is suitable for multiplex qPCR. The example below shows amplification of a 5-plex reaction that demonstrates the ability of the master mix to handle these types of challenging experiments. Here we amplified 5 targets: CHEK1 (ROX), CSK (Tamara), HPRT (FAM), RNase P (Cy5) and SFRS9 (SUN). As seen, the master mix can amplify all targets over 8 logs or RNA template.
Figure 4. PrimeTime One-Step 4X Broad-Range Master Mix provides exceptional multiplexed (5-plex) RNA identification over 8 logs of RNA template. RT-qPCR probe assays containing probes and primers specific for CHEK1, CSK, HPRT, RNase P, and SFRS9 genes were used to amplify a 10-fold dilution series of pooled RNA Ultramer templates ranging from 108 copies, down to 10 copies. RT-qPCR reactions were run on the QuantStudio 7 Flex.
Inhibitor resistant
Samples that contain PCR inhibitors can cause issues with amplification and data analysis. The PrimeTime One-Step 4X Broad-Range Master Mix is designed to handle these types of samples and give consistent amplification results. The data below shows amplification of samples with common PCR inhibitors spiked-in to reactions.
Figure 5. PrimeTime One-Step 4X Broad-Range Master Mix has a higher tolerance to heparin inhibition than three other commercially available master mixes. RT-qPCR probe assays containing probes and primers specific for Flu A (shown), Flu B, SARS-CoV-2, and RNase P genes were used to amplify RNA template in the presence of heparin concentrations ranging from 0.78−25 U/mL. Each master mix was used according to the manufacturer’s instructions. RT-qPCR reactions were run on the QuantStudio 7 Flex (n = 6, per inhibitor concentration per master mix). Error bars represent standard deviation. *Indicates reactions where no amplification occurred in the presence of heparin.
Figure 6. PrimeTime One-Step 4X Broad-Range Master Mix has a higher tolerance to hematin inhibition than three other commercially available master mixes. RT-qPCR probe assays containing probes and primers specific for Flu A (shown), Flu B, SARS-CoV-2, and RNase P genes were used to amplify RNA template in the presence of hematin concentrations ranging from 1.88−60 µM. Each master mix was used according to the manufacturer’s instructions. RT-qPCR reactions were run on the QuantStudio 7 Flex (n = 6, per inhibitor concentration per master mix). Error bars represent standard deviation.
Figure 7. PrimeTime One-Step 4X Broad-Range Master Mix has a higher tolerance to humic acid inhibition than two out of three other commercially available master mixes. RT-qPCR probe assays containing probes and primers specific for Flu A (shown), Flu B, SARS-CoV-2, and RNase P genes were used to amplify RNA template in the presence of humic acid concentrations ranging from 1.88−60 ng/µL. Each master mix was used according to the manufacturer’s instructions. RT-qPCR reactions were run on the QuantStudio 7 Flex (n = 6, per inhibitor concentration per master mix). Error bars represent standard deviation.
Resources
| Filename | Download |
|---|---|
| IDT-flyer-primetime-one-step-4x-broad-range-master-mix-Lubio.pdf |  |
| IDT-protocol-primetime-one-step-4x-broad-range-master-mix-Lubio.pdf | |
| IDT-technical_note-Direct_qPCR_amplification_from_human_urine_samples_technical_note-Lubio.pdf | |
| 23_QP_Technical_Note_Direct_amplification_of_cell_culture_samples-F.pdf.pdf | |
For research use only. Not for use in diagnostic procedures. Unless otherwise agreed to in writing, IDT does not intend for these products to be used in clinical applications and does not warrant their fitness or suitability for any clinical diagnostic use. Purchaser is solely responsible for all decisions regarding the use of these products and any associated regulatory or legal obligations.