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For research use only.
Probe-based qPCR master mix that has been optimized for highly sensitive and accurate quantification of up to 4 targets in a single reaction. This master mix was developed for TaqMan® probes but is also suitable for other hydrolysis probe types
HOT FIREPol® Multiplex qPCR Mix (Purple) is optimized for amplifying multiple targets in a single reaction in real-time quantitative PCR assays. The qPCR Mix comprises all the components necessary (except primers, probes, and templates) to perform qPCR: HOT FIREPol® DNA Polymerase, optimized buffer components, ultrapure dNTPs, MgCl2, and Purple passive reference dye according to system requirements.
HOT FIREPol® Multiplex qPCR Mix (Purple) is optimized for DNA hydrolysis probes based on the 5' flap endonuclease activity.
HOT FIREPol® DNA Polymerase is activated by a 10 min incubation step at 95°C. This prevents the extension of non-specifically annealed primers and primer-dimers formed at low temperatures during qPCR setup.
Concentration: 5x
Hot-start: yes, initial activation in 10 min
Detection type: Probe-based
Reference dye: Purple
Suitable qPCR Cyclers: 7500 Fast Real-Time PCR System, 7500 Real-Time PCR System, ViiA™ 7 PCR System, and QuantStudio™ 6 Flex, 7 Flex, and 12K Flex Real-Time PCR systems. Check Your cycler!
HOT FIREPol® DNA Polymerase: chemically modified FIREPol® DNA Polymerase enabling hot-start
5x Multiplex qPCR buffer with 15 mM MgCl2: 1x PCR solution – 3mM MgCl2
dNTPs including dUTP: the mix allows UNG treatment to prevent carryover contamination from previous runs.IMPORTANT: UNG is not included in the HOT FIREPol® Multiplex qPCR Mix
Purple dye: If Cy5 or a similar dye is used as one of the fluorophores, Purple passive reference dye might interfere with the signal. A version without passive reference or with ROX reference dye is available.
Dye-based detection (e.g., EvaGreen®, SolisGreen®, SYBR® Green) is a cost-effective qPCR option, as it requires only addition of PCR primers. However, the intercalating dye will detect any dsDNA (non-specific amplicons, primer dimers) produced in the reaction. Melt curve analysis performed after the qPCR can be used to verify the specificity of amplification and to check for the presence of non-specific amplification products.
The probe-based qPCR system demonstrates higher specificity compared to dye-based qPCR, because probes only detect the gene of interest. Keep in mind that in probe-based assays, primer dimers and non-specific products will not be detected, however, they may compromise the PCR efficiency. Using probe-based qPCR system, it is possible to distinguish between sequences with high similarity (e.g. single-nucleotide variations). Additionally, probe-based qPCR assays allow for multiplex reactions in one tube, while only a single target can be amplified and measured in a dye-based qPCR.
Recommended final amount of cDNA sample in downstream PCR reaction is up to one tenth of the final reaction volume. Overload of cDNA sample may compromise the downstream PCR, because cDNA sample may contain reaction components that may inhibit your PCR reaction.
ROX (carboxy-X-rhodamine) is one of the passive reference dyes providing a constant fluorescent signal that is used for signal normalization during the amplification cycles. The emission recorded from a reference dye during the baseline cycles is used to normalize the emission recorded from the reporter (e.g. EvaGreen®, SolisGreen®, SYBR® Green, etc.) in later cycles in ROX-dependent real-time PCR systems (e.g. Applied Biosystems 5700, 7000, 7300, 7700, 7900HT, StepOne™, StepOnePlus™). Reference dye compensates for small fluorescent fluctuations and well-to-well variations that may occur.
In Solis BioDyne qPCR mixes we use technology, based on ROX, that allows us to use same mix for high- and low- ROX requiering cyclers.
Please check cycler-mix compatibility here if not sure.
Solis BioDyne products should be stored at -20°C.
Shipping and temporary storage for up to 1 month at room temperature (*15−25°C) has no detrimental effects on the quality of Solis BioDyne reagents.
Freeze-thaw stability is tested for each product. Most PCR and qPCR products have passed 30 freeze-thaw cycles with no changes in performance. Specific information is found in Storage and Shipping conditions of each product.
When stored and handled under the recommended conditions, full activity of the reagents is retained until the Expiry Date printed on the tube label.
*World Health Organization (2003). Guidelines for the Storage of Essential Medicines and Other Health Commodities.
Routine storage: -20°C
Temporary storage for up to 1 month at room temperature has no detrimental effects on the quality of HOT FIREPol® Multiplex qPCR Mix.
At room temperature
Hot-start DNA Polymerase with unique 30-day room temperature stability for your everyday PCR needs.
High performance hot-start Master Mix for efficient multiplex reaction with ready-to-load feature - analyse up to 18 targets in one reaction.
Robust and reliable hot-start Master Mix with higher fidelity and longer amplification range for more demanding reactions.
High-performing probe-based qPCR Mix for AT-rich, regular and GC-rich templates.
Sensitive and highly specific cDNA synthesis and probe-based qPCR in a single tube. Ideal solution for high throughput RNA analysis.
