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TAQuest™ FAST qPCR Master Mix for TaqMan Probes *High ROX*
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Application notes
TAQuest™ FAST qPCR Master Mix for TaqMan Probes is a ready-to-use 2X solution optimized for qPCR and 2-step RT-qPCR well suited to use for TaqMan gene expression assays. The master mix is compatible with FAST conditions, thus delivers results within 50 minutes for 40 cycles of PCR in a 20 uL reaction volume. The master mix provides all of the essential components including our proprietary TAQuest™ FAST Hot Start Taq DNA Polymerase enzyme and dNTPs in an optimized PCR buffer, except the template, primers and probes. TAQuest™ FAST qPCR Master Mix for TaqMan Probes has been designed to be used for duplex reactions using internal positive controls with superior performance. The master mix ensures PCR specificity and sensitivity with all sample types such as genomic, plasmid, viral and cDNA templates. This master mix contains a high amount of ROX reference dye.
Example protocol
SAMPLE EXPERIMENTAL PROTOCOL
The following protocol can be used as a guideline.
Note Thaw the TAQuest™ FAST qPCR Master Mix for TaqMan Probes *High ROX* at room temperature. Vortex qPCR Master Mix thoroughly before use.
Table 2.Thermal cycling parameters
Note Thaw the TAQuest™ FAST qPCR Master Mix for TaqMan Probes *High ROX* at room temperature. Vortex qPCR Master Mix thoroughly before use.
- Prepare one of the following reaction mixes as indicated in Table 1.
- Carefully mix the reagents with a gentle vortex followed by a brief centrifuge.
- Set up the plate in the qPCR instrument and run as indicated in Table 2.
| Components | Volume (25 µL/reaction) | Volume (50 µL/reaction) | Final Conc. |
| TAQuest™ FAST qPCR Master Mix for TaqMan Probes *High ROX* | 12.5 µL | 25 µL | 1X |
| Upstream primer, 10 µM | 0.25-2.5 µL | 0.5-5.0 µL | 0.1-1.0 µM |
| Downstream primer, 10 µM | 0.25-2.5 µL | 0.5-5.0 µL | 0.1-1.0 µM |
| TaqMan Probes, 10 µM | 0.25-0.625 µL | 0.5-1.25 µL | 100-250 nM |
| DNA template | 1-5 µL | 1-5 µL | Optimized conc. |
| Nuclease-Free Water to | 25 µL | 50 µL |
| Parameter | Polymerase Activation | PCR (30-40 cycles) | |
| Hold | Denature | Anneal/Extend | |
| Temperature | 95 °C | 95 °C | 60 °C |
| Time (m:ss) | 0:10 | 0:20 | 0:30 |
References
View all 9 references: Citation Explorer
Resilient SARS-CoV-2 diagnostics workflows including viral heat inactivation.
Authors: Lista, Maria Jose and Matos, Pedro M and Maguire, Thomas J A and Poulton, Kate and Ortiz-Zapater, Elena and Page, Robert and Sertkaya, Helin and Ortega-Prieto, Ana M and Scourfield, Edward and O'Byrne, Aoife M and Bouton, Clement and Dickenson, Ruth E and Ficarelli, Mattia and Jimenez-Guardeño, Jose M and Howard, Mark and Betancor, Gilberto and Galao, Rui Pedro and Pickering, Suzanne and Signell, Adrian W and Wilson, Harry and Cliff, Penelope and Kia Ik, Mark Tan and Patel, Amita and MacMahon, Eithne and Cunningham, Emma and Doores, Katie and Agromayor, Monica and Martin-Serrano, Juan and Perucha, Esperanza and Mischo, Hannah E and Shankar-Hari, Manu and Batra, Rahul and Edgeworth, Jonathan and Zuckerman, Mark and Malim, Michael H and Neil, Stuart and Martinez-Nunez, Rocio Teresa
Journal: PloS one (2021): e0256813
Authors: Lista, Maria Jose and Matos, Pedro M and Maguire, Thomas J A and Poulton, Kate and Ortiz-Zapater, Elena and Page, Robert and Sertkaya, Helin and Ortega-Prieto, Ana M and Scourfield, Edward and O'Byrne, Aoife M and Bouton, Clement and Dickenson, Ruth E and Ficarelli, Mattia and Jimenez-Guardeño, Jose M and Howard, Mark and Betancor, Gilberto and Galao, Rui Pedro and Pickering, Suzanne and Signell, Adrian W and Wilson, Harry and Cliff, Penelope and Kia Ik, Mark Tan and Patel, Amita and MacMahon, Eithne and Cunningham, Emma and Doores, Katie and Agromayor, Monica and Martin-Serrano, Juan and Perucha, Esperanza and Mischo, Hannah E and Shankar-Hari, Manu and Batra, Rahul and Edgeworth, Jonathan and Zuckerman, Mark and Malim, Michael H and Neil, Stuart and Martinez-Nunez, Rocio Teresa
Journal: PloS one (2021): e0256813
Development of a multiplex TaqMan qPCR assay for simultaneous detection and differentiation of four DNA and RNA viruses from clinical samples of sheep and goats.
Authors: Xu, Xingang and Yang, Feng and Zhang, Qi and Xu, Ying and Huang, Jiali and Fu, Mingzhe and Zhang, Weimin
Journal: Journal of virological methods (2019): 58-64
Authors: Xu, Xingang and Yang, Feng and Zhang, Qi and Xu, Ying and Huang, Jiali and Fu, Mingzhe and Zhang, Weimin
Journal: Journal of virological methods (2019): 58-64
Evaluation and utilization of preassembled frozen commercial fast real-time qPCR master mixes for detection of cytomegalovirus and BK virus.
Authors: Glover, William A and Atienza, Ederlyn E and Nesbitt, Shannon and Kim, Woo J and Castor, Jared and Cook, Linda and Jerome, Keith R
Journal: Journal of medical virology (2016): 115-9
Authors: Glover, William A and Atienza, Ederlyn E and Nesbitt, Shannon and Kim, Woo J and Castor, Jared and Cook, Linda and Jerome, Keith R
Journal: Journal of medical virology (2016): 115-9
Real-time stability testing of air-dried primers and fluorogenic hydrolysis probes stabilized by trehalose and xanthan.
Authors: Rombach, Markus and Kosse, Dominique and Faltin, Bernd and Wadle, Simon and Roth, Günter and Zengerle, Roland and von Stetten, Felix
Journal: BioTechniques (2014): 151-5
Authors: Rombach, Markus and Kosse, Dominique and Faltin, Bernd and Wadle, Simon and Roth, Günter and Zengerle, Roland and von Stetten, Felix
Journal: BioTechniques (2014): 151-5
Frequency-encoded laser-induced fluorescence for multiplexed detection in infrared-mediated quantitative PCR.
Authors: Schrell, Adrian M and Roper, Michael G
Journal: The Analyst (2014): 2695-701
Authors: Schrell, Adrian M and Roper, Michael G
Journal: The Analyst (2014): 2695-701