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Digital High Resolution Melting (dHRM) is an advanced molecular technique used for the analysis of genetic variations, such as single nucleotide polymorphisms (SNPs), mutations, and methylations, by monitoring the melting behavior of double-stranded DNA. It is a post-PCR method that involves the gradual heating of PCR-amplified DNA in the presence of intercalating dyes that fluoresce when bound to double-stranded DNA. As the DNA melts, the fluorescence decreases, and the changes in fluorescence are monitored in real-time. The resulting melting curves are then analyzed to detect genetic differences based on the melting temperatures of the DNA fragments.

dHRM is enabled by the use of sensitive DNA-binding dyes and real-time PCR instrumentation, which allows for the collection of high-density data points to generate detailed melt profiles. These profiles can be used to identify even subtle differences in nucleic acid sequences, making dHRM a powerful tool for genotyping, mutation scanning, and methylation analysis

The technique has been further advanced by its application on digital microfluidics platforms, which can facilitate the analysis of single-nucleotide polymorphisms (SNPs) with high accuracy and sensitivity. Additionally, massively parallel dHRM has been developed to enable rapid and absolutely quantitative sequence profiling, which can be particularly useful in clinical and industrial settings where accurate quantification of nucleic acids is critical.

dHRM is also used in conjunction with digital PCR (dPCR) to improve quantitative power by providing additional information on the melting behavior of the amplified DNA, which can help in distinguishing between different genetic variants and in ensuring the accuracy of the quantification.

digital High Resolution Melting (dHRM) is a sophisticated technique for the detailed analysis of genetic variations, leveraging the precision of melting curve analysis in a digital or high-throughput format to provide accurate genotyping and mutation detection.