High Resolution Melting Method in Molecular Diagnostics and Their Clinical Importance

Abstract

Along with the development of technology in the field of molecular biology, many new molecular analyses are being developed each day. In this review, we aim to explain high-resolution melting (HRM) analysis, which has been shown to be important in molecular diagnostics, its applications, and its importance in the areas of clinical practice. The HRM system in a closed tube is used to determine post-polymerase chain reaction (PCR)based genetic variations as a new method. The working principle of HRM is based on the melting behavior of nucleic acid samples. The denaturation of double-stranded DNA is determined by detecting the fluorescence change caused by increased melting temperature. The differences between wildtype and heterozygous samples may be easily detected in melting graphics. Using this method, melting curve analysis can be performed with more precision. In HRM analysis, the samples can be distinguished according to the guanine cytosine (GC) content and sequence length. Thus, the detection of common single-nucleotide polymorphisms (SNPs) in the population, scanning of gene mutations associated with diseases, and analysis of DNA methylation can be performed quickly and reliably using the HRM method. Nucleotide sequence variations and several variations in the PCR products can be detected with the DNA melting curve shape using the HRM method. In addition to being low-cost and easily implementable, the real-time HRM method enables powerful analysis because of the combined new-generation DNA dyes and developed gene scanning software, thereby standing out in many clinical applications.