ProbeMasterMix (2X) Low ROX for qPCR

ProbeMasterMix (2X) Low ROX für die qPCR


Order number: M3031.0100

Shipping: Shipment: on wet ice. Store at -20°C. For laboratory usage only!

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ProbeMasterMix low ROX optimised for realtime PCR assays in block systems that contains all... more
Product information "ProbeMasterMix (2X) Low ROX for qPCR"

ProbeMasterMix low ROX optimised for realtime PCR assays in block systems that contains all components to perform quantitative PCR with the exception of primer and template DNA. This 2X master mix is ready-to-use and contains optimised amounts of all ingredients.

Multiplex PCR: Applications at Genaxxon and at customers site have shown that the qPCR Probe Mastermix can be used for the simultaneous detection of up to four DNA targets in the same PCR reaction. For further details please refer to the product manual or contact us: info@genaxxon.com.

Advantages of the Genaxxon ProbeMasterMix:

  • Hotstart technology enables setup of PCR mixture at room temperatur
  • No pipetting on ice necessary anymore
  • No immediate further processing (PCR) necessary. The pipetted pre-PCR mixtures can be left at RT for up to 3 days.
  • Amplification of GC-rich templates
  • High yields
  • No Primer dimers

The Genaxxon SuperHot Taq DNA polymerase offers convenience and practicality, without compromising efficiency or specificity:

1. The chemical modification inhibits the polymerase, which prevents false priming at all temperatures tested, allowing for PCR reactions to be set up at RT (20°C to 25°C), and 
2. "High mix stability allows pre-assemby of the plate in advance", which means PCR reactions can be prepared and stored at RT for up to three days before they are run, without affecting the results.

The Probe qPCR master mix without ROX contains all the necessary components in an optimized composition to carry out quantitative PCR.

  • chemically modified Taq DNA Polymerase
  • dATP, dCTP, dGTP, dTTP
  • 50nM ROX
  • optimized reaction buffer
  • stabilizers and enhancers to enable even amplification of low copy number targets
  • the small aliquote size of 1.25mL simplifies handling and storage (less freeze-thaw cycles per aliquote)

Stability at +2°C to +8°C (refrigerator): minimum of 8 months.

This Mastermix is specially suited for the following instruments: Applied Biosystems® 7500, 7500 Fast and ViiA™ 7, QuantStudio™ instruments, Agilent Mx3000P™, Mx3005P™, Mx4000™ and AriaMx.

Other realtime PCR master mixes from Genaxxon are: M3023 - GreenMastermix No ROX >, M3045 - ProbeMastermix No ROX >, M3011 - GreenMastermix Low ROX >, M3031 - ProbeMastermix Low ROX >, M3052 - GreenMastermix High ROX >, M3010 - ProbeMastermix High ROX >.

Test sample available! The test sample price will be refunded on the first official order of the product.

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Our comment on "ProbeMasterMix (2X) Low ROX for qPCR"
High Sensitivity and Specifity, Optimised buffer system, Reproducibility over a wide range of template amounts, Broad dynamic range. Aliquot size: 1.25mL. This master mix is stable for at least 8 months at +2°C to +8°C.
2-time ready-to-use realtime PCR mastermix with 50nM ROX™ as reference dye. Aliquot size:... more
 

Technical Data:

2-time ready-to-use realtime PCR mastermix with 50nM ROX™ as reference dye.

Aliquot size: 1.25mL. This master mix is stable for at least 8 months at +2°C to +8°C.

application:

PCR Primer extension Multiplex PCR Low-copy targets PCR Real-time PCR Quantitative PCR

Unit Definition:

One unit is defined as the amount of enzyme which will convert 10 nmoles of dNTPs to an acid-insoluble form in 30 min at 72°C under the assay conditions (25 mM TAPS (tris-(hydroxymethyl)-methyl-amino-propanesulfonic acid, sodium salt) pH 9.3 (25°C); 50 mM KCl; 2 mM MgCl2; 1 mM b-mercaptoethanol; and activated calf thymus DNA as substrate.

Sicherheits Hinweise / Safety

Klassifizierungen / Classification

eclass-Nr: 34-16-04-90
Dokumente - Protokolle - Downloads more


Dokumente - Protokolle - Downloads

Here you will find information and further literature on ProbeMasterMix (2X) Low ROX for qPCR. For further documents (certificates with additional lot numbers, safety data sheets in other languages, further product information) please contact Genaxxon biosience at: info@genaxxon.com or phone: +49 731 3608 123.

 
 
 
Dokumente - Protokolle - Downloads more

Dokumente - Protokolle - Downloads

Fast PCR:

When performing fast real-time PCR, it is of high importance to consider primer design and target size. For efficient amplification during fast cycling conditions, target size between 70 bp and 200 bp is recommended. The shorter the target length,  the faster the total run time. For other targets and other primer sets, the annealing step has to be determined experimentally.

Please use the following protocol for inspiration as you need to optimize instruments settings according to yourr target size and applied instruments.

Fast real-time PCR results using Genaxxon Master Mixes on Mx3005P
GreenMasterMix Low ROXTM and ProbeMasterMix Low ROXTM were tested on Mx3005P (Agilent Technologies) using fast cycling real-time 2-step protocols; Fast Pthr Green Mx and Fast Pthr Probe Mx, respectively.
qPCR Setup: For GreenMasterMix Low ROXTM and ProbeMasterMix Low ROXTM, primers targeting Pthr (75 bp) and 4 concentrations of gDNA (40 ng, 20 ng, 10 ng and 5 ng) were used. All DNA concentrations were tested in quadruple replicates. See fig. 3 and 4. The PCR reaction mix was run on MX3005P (Requiring Low ROX). Instrument settings, run times and quality criteria for the applied fast protocols; Fast Pthr Green Mx and Fast Pthr Probe Mx, were compared to the standard protocol; Standard Pthr Mx. See table 2.
Results: The standard Fast Pthr Probe Mx protocol has a run time of 83.4 minutes. Fast Pthr Green Mx and Fast Pthr Probe Mx protocols, resulted in run times of 50.1 and 46.8 minutes, respectively. All protocol times are inclusive ramping time and a15 minutes hot start. The determined criteria, as shown in table 2, for Fast Pthr Green Mx and Fast Pthr Probe Mx protocols were acceptable. Furthermore, the melt curve analysis using GreenMasterMix Low ROXTM with Fast Pthr Green Mx protocol, confirmed high specificity.


Figure 3. Amplification plot, standard curve and melt curve for GreenMasterMix run on Mx3005P using Fast Pthr Green Mx protocol. Primers targeting Pthr (75 bp) and 4 concentrations of gDNA (40 ng, 20 ng, 10 ng and 5 ng) were used. All DNA concentrations were tested in quadruple replicates.

Figure 4. Amplification plot and standard curve for ProbeMasterMix run on Mx3005P using Fast Pthr Probe Mx protocol. Primers targeting Pthr (75 bp) and 4 concentrations of gDNA (40 ng, 20 ng, 10 ng and 5 ng) were used.

 
 
 
FAQ
Fragen & Antworten zum Artikel ... mehr
Why do I need a Hot Start Polymerase?
The Hot Start-modification of the polymerase inhibits its enzymatic activity at low temperatures and prevents non-specific amplifications in the first PCR cycles, which can drastically affect the qPCR results.
When using SYBR Green as a fluorescent dye, my quantification is not accurate. What can I do?
SYBR Green has only a low specificity. It binds to double-stranded DNA during PCR cycles, even to potentially non-specific reaction products. This leads to inaccurate qPCR results with non-optimised protocols. To obtain more specific results, probe-based qPCR should be performed.
The qPCR MasterMix does not work with the specified protocol. What can I do?
The protocols provided by Genaxxon are standard protocols. In order to obtain optimal specificity and amplification, individual optimisation of the experimental conditions is recommended. If you have further questions about this, please feel free to contact our technical support: info@genaxxon.com.
With the qPCR MasterMix from Genaxxon I get a different Ct value than with my previous MasterMix. What can I do?
The MasterMixes from different manufacturers have different compositions, including the buffers. Some assays react sensitively to these variations, so that different results are possible. Therefore, a calibration is always necessary when changing the MasterMix. If you have further questions, please contact our technical support: info@genaxxon.com.
Can I run multiplex reactions with the Genaxxon qPCR mastermixes?
Yes, all qPCR mastermixes from Genaxxon can be used for multiplex applications. However, for easy and fast optimization of your multiplex reaction, we recommend using one of our specially designed multiplex mastermixes, e.g., 5X qPCR Multiplex MasterMix (M3024), Multiplex HS MasterMix (2X) (M3013) or the lyophilized LyoPlex Multiplex PCR master mix (M3005).
Can I use a fast protocol of my instrument settings with Genaxxon’s qPCR mastermixes?
Genaxxon offers different qPCR mastermixes optimised for fast qPCR assays. Depending on your needs, Genaxxon offers mastermixes for fast qPCR assays with (GreenMasterMix FAST) or without (ProbeMasterMix FAST) a fluorescence dye and with no (No ROX™) or a passive reference dye with different concentrations (Low ROX™ or High ROX™). If you have any problems choosing the right mastermix for your application, please contact our technical support: info@genaxxon.com.
Do I need to add ROX™ when using Genaxxon’s qPCR Mastermixes?
Depending on the qPCR instrument you’re using, you may need to use a passive reference dye, typically ROX, to overcome variations between wells caused by the machine limitations. Therefore, Genaxxon offers qPCR mastermixes with either no or low and high amounts of the passive reference dye ROX™ to ensure compatibility with a variety of qPCR instruments. If you have any problems choosing the right mastermix for your application, please contact our technical support: info@genaxxon.com.