Precise, highly reproducible and fast genotyping of sequence variants through high resolution melting.
Product Highlights
Accurate – clear discrimination of even the most challenging sequence differences
Reproducible – unparalleled consistency between replicate difference plots for increased confidence in sample characterization
Sensitive – efficient detection from even very limiting amounts of sample
Flexible – enables reliable characterization of a broad range of sequence differences
Fast – rapid PCR amplification prior to high resolution melting, enabling higher throughput
Product Description
SensiFAST HRM Kit has been developed for detailed characterization of samples according to their base composition, length and GC content by high resolution melting. The latest advances in buffer chemistry and enhancers, together with an antibody-mediated hot-start DNA polymerase system, ensure the SensiFAST HRM Kit delivers reproducible, accurate HRM analysis.
SensiFAST HRM Kit contains EvaGreen®, a third generation, saturating fluorescent dye which selectively binds to double-stranded DNA. In contrast to dyes such as SYBR® Green I, EvaGreen can be used at higher concentrations without inhibiting PCR and shows equal binding affinity for GC-rich and AT-rich regions. The combination of our SensiFAST DNA Polymerase, unique buffer system and EvaGreen dye enables amplification and discrimination of even the most challenging sequence differences, such as class IV SNPs without sequence preference. Since it does not require expensive labelled oligonucleotide probes, SensiFAST HRM Kit is also a cost effective alternative to traditional probe based genotyping methods.
Applications
SNP genotyping
Characterization of haplotype blocks
DNA mapping
Detection of insertions, deletions and translocations
DNA fingerprinting
DNA methylation analysis
Gene scanning
Species identification
Allelic prevalence population studies
Screening for loss of heterozygosity
Fig. 1 Highly successful genotyping of a Class 4 (A/T) SNP using SensiFAST HRM
Class 4 SNPs are the most challenging to differentiate; having Tm changes typically less than 0.2oC. The three different genotypes of an A>T SNP were analysed on the Rotor-Gene 6000, using SensiFAST HRM Kit. A difference plot of this data illustrates the effect of subtracting an average heterozygous curve from all curves and plotting the results.
Introduction to SensiFAST
Overview, features and benefits of the SensiFAST product family
A discussion of the pros and cons of each detection strategy.
We used the Meridian SensiFAST HRM mix to discriminate changes at a locus with substitutions on both strands, yielding AT heteroduplexes as well as either A or T monoduplexes. We easily achieved calls on the mono duplexes over heteroduplexes with more than 99% confidence to provide extremely sensitive genotyping of an AT class 4 SNP.
Dr Simon Baker, Senior Director of R&D, Meridian, London
Instrument Compatibility
Please refer to the Real-Time PCR Selection Chart to confirm the SensiFAST™ HRM Kit is compatible with your instrument.
IO-32005: 500 x 20 µL Reactions: 5 x 1 mL BIO-32020: 2000 x 20 µL Reactions: 20 x 1 mL
Storage & Stability
All kit components should be stored at -20°C upon receipt. When stored under the recommended conditions and handled correctly, full activity of the kit is retained until the expiry date on the outer box label. Avoid exposure of the EvaGreen to light.