repliQa HiFi ToughMix
- High Fidelity – >90x wild type Taq
- Extreme Speed – up to 3x faster PCR results with extension rates as fast as 1 kb/sec*
- Tough Tested – tolerant to a wide range of PCR inhibitors
- Superior Sensitivity – higher yields from lower inputs as little as 2 pg
- Long Range – amplify +24 kb gDNA, +40 kb λDNA
*for fragments less than 1 kb in size
repliQa HiFi ToughMix is intended for molecular biology applications. This product is not intended for the diagnosis, prevention or treatment of a disease.
repliQa HiFi ToughMix
Description
The repliQa HiFi ToughMix is a 2x, ready-to-use master mix that contains all the components for high fidelity PCR amplification, including a genetically modified DNA polymerase coupled with hot start antibodies. This unique, next generation master mix provides 90x higher fidelity compared to Taq, while reducing time to PCR results by 2-3x. The extreme speed is enabled by extension times as fast as 1-10 kb/sec depending on target length. The enzyme is coupled with the industry leading ToughMix which is tolerant to a wide variety of inhibitors making it suitable for routine PCR, cloning, amplicon sequencing and site directed mutagenesis.
Details
2x reaction buffer containing optimized concentrations of MgCl2, dNTP’s and proprietarily formulated HiFi polymerase, hot start antibodies and ToughMix chemistry
Performance Data
Extreme Speed: Up to 3x faster results
Comparison of speed. A 2 kb fragment was amplified in 50 μl reaction volumes according to the recommended protocol. Following a 30 s activation at 98°C; 30 cycles of PCR were performed: 98°C, 10 s; 60°C, 10 s; 68°C, 5–30 s. The thermal cycler had a ramp rate of 5°C/s.
Tough Tested: Tolerant to a wide range of PCR inhibitors
Strong Inhibitor Resistance. A 2 kb λ DNA template was amplified using each manufacturer’s recommended cycling conditions with different amounts io inhibitors. This experiment was run in duplicate.
Superior Yield and Sensitivity
Comparison of yield. A gDNA template was amplified with varying GC-content and length targets using each manufacturer’s recommended cycling conditions. The experiment was run using 8 different targets, in duplicate.
Long Amplification and Consistent GC Tolerance
A Long Range Capabilities. A range from 0.25 to 24 kb of gDNA templates were amplified with varying GC-content and lengths using recommended cycling
conditions. B/C Wide GC-content tolerance range. gDNA templates were amplified with varying GC-content and lengths using recommended cycling conditions.
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Customer Product Reviews
| 5 star | 64% | |
| 4 star | 29% | |
| 3 star | 4% | |
| 2 star | 2% | |
| 1 star | 0% |
repliQa HiFi ToughMix
Worked very well for high GC target where other polymerases, even with high GC buffers or modifications failed.
This product works very well for us. It helps us amplify a large amount of gDNA (5-10ug), for which we had a lot of challenges with other polymerases.
I used the protocoll with very short time amplification and got at least the same amount of product, without mutations, compare to Q5 HiFi Polymerase from NEB . I ‘m part of the UNIKLINIK Mainz, so I #m not shure that your offer is better than the discount for the UNI-customers. I plane to change the Firma and to by your enzymes by the next order, but I have still to compare the prices.
The repliQa HiFi ToughMix was chosen for evaluation as a master mix for wildlife species identification, specifically for use with the 3500 Genetic Analyzer platform. The initial setup process for the repliQa HiFi ToughMix was notably straightforward and easy to implement in the laboratory workflow, which was a significant advantage.
The primary objective of this evaluation was to determine the suitability of the repliQa HiFi ToughMix for reliable species identification across a diverse panel of wildlife species and various challenging sample types commonly encountered in forensic and conservation case work. The testing was rigorously designed to assess three critical performance metrics: sensitivity, robustness, and overall sequence quality. The repliQa HiFi ToughMix was applied to a matrix of samples, including those with low DNA quantity, degraded DNA, and samples containing potential inhibitors.
Sensitivity and Setup: While the ease of setup was confirmed, and the mix demonstrated adequate sensitivity for high-quality, pristine DNA samples, performance declined significantly with compromised samples.
Robustness: The mix’s purported “tough” nature did not fully translate to the required robustness for the most challenging sample types. Specifically, a high rate of PCR inhibition or inconsistent amplification was observed when testing highly degraded or inhibited samples.
Sequence Quality: The most critical finding related to the quality of the resulting sequences. Across several tested species and difficult sample types, the reverse sequence often failed to meet the stringent quality control thresholds required for casework. Poor reverse sequence quality—manifesting as low signal-to-noise ratio, numerous ambiguous base calls, or outright sequence failure—renders the data unusable for reliable and legally defensible species identification in forensic or regulatory contexts.
In summary, the repliQa HiFi ToughMix proved to be an accessible reagent for initial laboratory setup. However, despite its user-friendly interface, its performance regarding sensitivity and, more critically, robustness and overall sequence quality—particularly for the reverse sequencing read—was inconsistent and often inadequate for the specific requirements of wildlife case work analysis. The failure to generate high-quality, bidirectional sequence data consistently, especially from challenging samples, means this particular master mix cannot be confidently recommended for routine use where absolute reliability and high sequence quality are paramount for species identification using the 3500 Genetic Analyzer.
Thanks for sharing your feedback. We are glad the AccuStart II PCR ToughMix worked. If you’d like – we have an expert FAS team that will be reaching out to help on repliQa.