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Background  The PCR technique and its variations have been increasingly used in the clinical laboratory and recent advances in this field generated new higher resolution techniques based on nucleic acid denaturation dynamics. The principle of these new molecular tools is based on the comparison of melting profiles, after denaturation of a DNA double strand. Until now, the secondary structure of single-stranded nucleic acids has not been exploited to develop identification systems based on PCR. To test the potential of single-strand RNA denaturation as a new alternative to detect specific nucleic acid variations, sequences from viruses of the Totiviridae family were compared using a new  in silico  melting curve approach. This family comprises double-stranded RNA virus, with a genome constituted by two ORFs, ORF1 and ORF2, which encodes the capsid/RNA binding proteins and an RNA-dependent RNA polymerase (RdRp), respectively.    Results  A phylogenetic tree based on RdRp amino acid sequences was constructed, and eight monophyletic groups were defined. Alignments of RdRp RNA sequences from each group were screened to identify RNA regions with conserved secondary structure. One region in the second half of ORF2 was identified and individually modeled using the  RNAfold  tool. Afterwards, each DNA or RNA sequence was denatured  in silico  using the softwares  MELTSIM  and  RNAheat  that generate melting curves considering the denaturation of a double stranded DNA and single stranded RNA, respectively. The same groups identified in the RdRp phylogenetic tree were retrieved by a clustering analysis of the melting curves data obtained from  RNAheat . Moreover, the same approach was used to successfully discriminate different variants of  Trichomonas vaginalis  virus, which was not possible by the visual comparison of the double stranded melting curves generated by  MELTSIM .    Conclusion   In silico  analysis indicate that ssRNA melting curves are more informative than dsDNA melting curves. Furthermore, conserved RNA structures may be determined from analysis of individuals that are phylogenetically related, and these regions may be used to support the reconstitution of their phylogenetic groups. These findings are a robust basis for the development of  in vitro  systems to ssRNA melting curves detection.    Electronic supplementary material  The online version of this article (doi:10.1186/1471-2105-15-243) contains supplementary material, which is available to authorized users.

In silicosingle strand melting curve: a new approach to identify nucleic acid polymorphisms in Totiviridae