MeltDoctor™ HRM Master Mix
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Applied Biosystems™

MeltDoctor™ HRM Master Mix

Applied Biosystems™ MeltDoctor™ HRM Master Mix contains all components (excluding template and primers) formulated for superior HRM performance across aRead more
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Catalog NumberQuantity
44154405 mL
440953550 mL
441545010 x 5 mL
44154525 x 5 mL
Catalog number 4415440
Price (USD)
664.00
Each
Add to cart
Quantity:
5 mL
Price (USD)
664.00
Each
Add to cart
Applied Biosystems™ MeltDoctor™ HRM Master Mix contains all components (excluding template and primers) formulated for superior HRM performance across a wide range of genomic targets. High Resolution Melting (HRM) analysis is a post-PCR analysis method used to identify genetic variation in nucleic acid sequences. Unlike some competitive mixes, the MeltDoctor™ HRM Master Mix does not require additional mixing prior to use, and was developed and optimized solely for HRM applications.
• Low background fluorescence
• High brightness in the presence of double-stranded DNA
• Minimal temperature shift of DNA melting due to dye binding
• Thermal stability to tolerate PCR cycling conditions
• No inhibition of polymerase activity, resulting in high PCR efficiency
• A dNTP blend including dUTP, which minimizes carryover contamination by allowing amplicon degradation by uracil DNA glycosylase (UDG) in subsequent PCR reactions

Perform Mutation Scanning
HRM analysis can be used to scan for mutations in target genes for the identification of variant samples prior to sequencing analysis. As a mutation scanning technique, HRM offers significant advantages over conventional methods such as Denaturing High- Performance Liquid Chromatography (DHPLC) and Denaturing Gradient Gel Electrophoresis (DGGE). Specifically, the advantages of HRM for mutation scanning include:
• Low reagent consumption, with little waste: HRM requires only a 20 μL PCR reaction for analysis of each sample, eliminating the need for HPLC solvents or DGGE gels
• Simple, fast workflow: no additional instrumentation is required after PCR amplification. A high-resolution melt step can be simply added to the end of the PCR profile for immediate analysis
• Fast optimization: unlike DHPLC, thermal optimization is not required
• Low sample consumption: following HRM analysis, the PCR product can be used directly in a Sanger sequencing reaction

Quality Components
• Magnesium salts and other buffer components, precisely formulated to obtain optimal HRM results.
• AmpliTaq Gold™ 360 DNA Polymerase, a highly purified DNA polymerase that provides hot-start performance, minimizing non-specific product formation and enabling reactions to be set up at room temperature
• MeltDoctor™ HRM Dye, a stabilized form of the fluorescent SYTO™ 9 double-stranded nucleic acid stain developed by Molecular Probes.
For Research Use Only. Not for use in diagnostic procedures.
Specifications
Detection MethodHRM
FormLiquid
GC-Rich PCR PerformanceHigh
PCR MethodqPCR
PolymeraseAmpliTaq Gold 360 DNA Polymerase
Reaction SpeedFast
TechniqueHRM (High Resolution Melting) Analysis, Mutation Scanning
For Use With (Equipment)7500 Fast System, 7900HT Fast System, QuantStudio™ 12k Flex, QuantStudio™ 3, QuantStudio™ 5, QuantStudio™ 6 Flex, QuantStudio™ 7, StepOne™, StepOnePlus™, ViiA™ 7 System
Genotyping TargetSNPs (Unknown or Numerous), Epigenetic Targets
Label or DyeHigh Resolution Melting (HRM), SYTO 9
Product LineMeltDoctor™
Product TypeHRM Master Mix
Quantity5 mL
Shipping ConditionWet Ice
For Use With (Application)Genotyping
No. of Reactions500 Reactions
Sample TypeDNA (Genomic), cDNA
Volume5 mL
Unit SizeEach
Contents & Storage
Components of the MeltDoctor™ HRM Master Mix, provided in a convenient 2X mix, include: AmpliTaq Gold™ 360 DNA Polymerase, MeltDoctor*trade; HRM Dye, a dNTP blend including dUTP, Magnesium salts and other buffer components, precisely formulated to obtain optimal HRM results.

Store at -15 °C to -25 °C

Guaranteed minimum shelf life is 60 days (exact expiry date printed on product and CofA).

Frequently asked questions (FAQs)

The HRM software will not accept my calibration file. What is wrong?

Check that there are no outliers on the plate. You cannot omit any wells on the HRM calibration plate.

I have a 7900HT Fast Real-Time PCR System, and cannot open the data file in HRM software. What could be causing this?

There are a few possibilities. First, make sure the HRM Software version is v2.0.1, and the 7900HT Fast Real-Time PCR System software version is v2.3 or above. Second, check that the run method used was as recommended in the HRM protocol; make sure the ramp rate for the dissociation stage is 1%. Then try to open the calibration file from the HRM Software; if it does not open, the calibration file is defective. The defects could be due to a bad calibration plate or instrument uniformity issue.

My 7500 Fast Real-Time PCR System has been upgraded to v2.0.6, so why can't I analyze my files with HRM Software v2.0.1?

For the 7500 Fast Real-Time PCR System, if the software version is below v2.0.4, the compatible HRM Software is v2.0.1. If the 7500 Fast Real-Time PCR System has been upgraded to software version 2.0.4 or above, then you will need to get HRM Software version 3.0.1.

How can I get positive controls for the different genotypes for my HRM experiment?

You can order a synthetic positive control plasmid using our GeneArt Gene Synthesis services. The final product is sequence verified and provided as 5 µg of lyophilized plasmid. Alternatively, you may be able to find a positive control sample from Coriell.

How do I perform the HRM calibration on my real-time PCR instrument?

The HRM calibration is a 3-step procedure. In some instrument software, the calibration process has been streamlined to a single-step procedure (applies to the ViiA7 Real-Time PCR System and QuantStudio Real-Time PCR Systems).

For the HRM calibration, the first step is a PCR reaction to generate the HRM dye plate. The reaction plate contains a DNA template, a primer pair, and the PCR mix with HRM dye. The second step is dye calibration using the PCR plate from step 1. The last step is a melt curve run, again using the same plate. For details, please refer to the HRM guides for each instrument:

- 7500 Fast Real-Time PCR System (with software version 1.4 or 2.0.4 and below) (https://tools.thermofisher.com/content/sfs/manuals/cms_050347.pdf)
- 7500 Fast Real-Time PCR System (with software version 2.0.5 and above) (https://tools.thermofisher.com/content/sfs/manuals/4457847.pdf)
- 7900HT Real-Time PCR System (https://tools.thermofisher.com/content/sfs/manuals/cms_050347.pdf)
- StepOne and StepOnePlus Real-Time PCR Systems (https://tools.thermofisher.com/content/sfs/manuals/4457847.pdf)
- ViiA 7 Real-Time PCR System (https://tools.thermofisher.com/content/sfs/manuals/4443531.pdf)
- QuantStudio 6 and 7 Real-Time PCR Systems (https://tools.thermofisher.com/content/sfs/manuals/4489823.pdf)
- QuantStudio 6 and 7 Pro Real-Time PCR Systems (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0018981_HighResolutionMelt_DA_SW_UG.pdf)
- QuantStudio 3 and 5 Real Time PCR Systems (https://tools.thermofisher.com/content/sfs/manuals/MAN0014394_HighResMeltExperiment_GSG.pdf)
- QuantStudio 12K Flex Real-Time PCR System (https://tools.thermofisher.com/content/sfs/manuals/4470692A.pdf)

Find additional tips, troubleshooting help, and resources within our High Resolution Melt Support Center.