title: Detection of a G>C single nucleotide polymorphism within a repetitive DNA sequence by high-resolution DNA melting.
abstract: In standard forensic DNA analysis, single base mutations within short tandem repeats (STR) mostly escape detection. In this study, high-resolution DNA melting (HRM) is compared to minisequencing and Sanger sequencing as to determine the most suitable method for detection of a G to C mutation within a repetitive DNA sequence, the STR system DXS10161. It shows an ATG/ATC polymorphism surrounded by a variable number of (TATC) and (ATCT) motifs. Neutral base changes like G:C to C:G result in very low differences in the melting temperature (T m) of the PCR amplicons. By enhanced resolution of fluorescence vs. temperature in HRM, the technique showed to be suitable for detecting a G to C transversion in this repetitive DNA sequence context. Compared to minisequencing, HRM is more time- and cost-effective. Results were confirmed by Sanger sequencing.
authors.affiliation: Institute of Legal Medicine, Freiburg University Medical Center, Albertstr. 9, 79104, Freiburg, Germany. ulrike.schmidt@uniklinik-freiburg.de.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Schmidt
authors.affiliation: Institute of Legal Medicine, Freiburg University Medical Center, Albertstr. 9, 79104, Freiburg, Germany.|Department of Biology, University of Freiburg, Schänzlestr. 1, 79104, Freiburg, Germany.
authors.forename: Johannes
authors.initials: J
authors.lastname: Hulkkonen
authors.affiliation: Institute of Legal Medicine, Freiburg University Medical Center, Albertstr. 9, 79104, Freiburg, Germany.|Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH, Amsterdam, Netherlands.
authors.forename: Jana
authors.initials: J
authors.lastname: Naue
mesh_terms: D005260:Female; D053803:Forensic Genetics; D005838:Genotype; D006801:Humans; D008297:Male; D018895:Microsatellite Repeats; D009691:Nucleic Acid Denaturation; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods*
keywords: Forensic genotyping; High-resolution melting (HRM); Minisequencing; Sanger sequencing; Short tandem repeat (STR); Single nucleotide polymorphism (SNP)