Some applications of this product may require a license which is not provided by the purchase of this product.
For research use only.
High-Resolution Melt (HRM) analysis is a fast and cost-effective method for post-PCR analysis to detect single nucleotide polymorphisms (SNP genotyping), genetic mutations, and DNA sequence variations.
5x HOT FIREPol® EvaGreen® HRM Mix is an optimized ready-to-use solution for HRM analysis, incorporating EvaGreen® dye.
It comprises all the components necessary to perform qPCR and HRM Analysis: HOT FIREPol® DNA Polymerase, ultrapure dNTPs, MgCl2, and EvaGreen® dye. The user simply needs to add water, template, and primers.
HOT FIREPol® DNA Polymerase is activated by a 12 min incubation step at 95°C. This prevents the extension of non-specifically annealed primers and primer-dimers.
Concentration: 5x
Hot-start: yes, initial activation in 12 min
Detection type: dye-based, includes EvaGreen® intercalating dye
Reference dye: none
Compatible real-time instruments: Cyclers that do not require ROX reference dye. Check Your cycler!
HOT FIREPol® DNA polymerase: chemically modified FIREPol® DNA Polymerase enabling hot-start
5x EvaGreen® qPCR buffer with 12. 5 mM MgCl2, 1x PCR solution – 2.5 mM MgCl2
dNTPs: dATP, dCTP, dGTP and dTTP
EvaGreen® dye
Bovine serum albumin (BSA) to enhance qPCR reaction
EvaGreen® is a DNA-binding dye with many features that make it a superior alternative to SYBR® Green I for qPCR. Apart from having similar spectra, EvaGreen® has three important features that set it apart from SYBR® Green I: EvaGreen® has much less PCR inhibition, is an extremely stable dye and has been shown to be non-mutagenic and non-cytotoxic. EvaGreen® is compatible with all common real-time PCR cyclers – simply select the standard settings for SYBR® Green or FAM!
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.
Both dyes are DNA intercalating agents that are used to stain dsDNA.
EvaGreen® dye is spectrally similar to FAM or SYBR® Green I, which means that no change in optical settings is required for using an EvaGreen®-based qPCR mix.
EvaGreen® was reported to have several benefits over SYBR® Green I:
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 oC
Temporary storage for up to 1 month at room temperature has no detrimental effects on the quality of the product
At room temperature
Ready-to -use solution for simple dye-based qPCR assays, incorporating EvaGreen® dye. Available for ROX, no ROX, and capillary cyclers.
Highly specific and reproducible dye-based qPCR Mix with blue visualization dye for easy pipetting. Suitable for ROX-dependent and ROX-independent qPCR cyclers.
High-performing probe-based qPCR Mix for AT-rich, regular and GC-rich templates.
All-inclusive and the most convenient product format for first-strand cDNA synthesis. A combined enzyme mix (FIREScript® and RiboGrip®) and a reaction premix in separate tubes allowing you to choose the most appropriate priming option.
