Background: High resolution melting curve analysis (HRM) is a technique that measures exactly the decreasing fluorescence of intercalating dye in the process of dissociation of double stranded DNA. The measurement is immediately following PCR in a one-step, closed-tube method. The shape of the melting curve depends on the GC content, length and sequence of the amplicon. Hence it is a powerful, fast and cheap method to detect Single Nucleotide Polymorphisms (SNPs) and other mutations.
Hellbori nigri rhizoma is a drug that is difficult to distinguish from other species of the genus Helleborus. In this communication we present a DNA-based identification by high-resolution melting analysis (HRM) that is able to differentiate between Helleborus niger and other species of the genus. HRM is a very specific, time- and labour-saving method for identifying DNA sequence variations and is ideally suitable for routine PCR analysis. The HRM assay developed is specific for the genus Helleborus. This method not only detects the presence of the target species H. niger but also, to a certain extent, identifies other Helleborus species by their different melting curve shapes. Markers were developed based on the trnL-trnF intergenic spacer and on the matK sequence. For an unambiguous identification of Helleborus niger, melting curves of both markers should be used.
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