title: High-resolution melt analysis of DNA methylation to discriminate semen in biological stains.
abstract: The goal of this study was to develop a method for the detection of semen in biological stains using high-resolution melt (HRM) analysis and DNA methylation. To perform this task, we used an epigenetic locus that targets a tissue-specific differentially methylated region for semen. This specific locus, ZC3H12D, contains methylated CpG sites that are hypomethylated in semen and hypermethylated in blood and saliva. Using this procedure, DNA from forensic stains can be isolated, processed using bisulfite-modified polymerase chain reaction (PCR), and detected by real-time PCR with HRM capability. The method described in this article is robust; we were able to obtain results from samples with as little as 1 ng of genomic DNA. Samples inhibited by humic acid still produced reliable results. Furthermore, the procedure is specific and will not amplify non-bisulfite-modified DNA. Because this process can be performed using real-time PCR and is quantitative, it fits nicely within the workflow of current forensic DNA laboratories. As a result, it should prove to be a useful technique for processing trace evidence samples for serological analysis.
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA.
authors.forename: Joana
authors.initials: J
authors.lastname: Antunes
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA; Faculty of Biosciences, Laboratory of Human and Molecular Genetics, PUCRS, 90619-900 Porto Alegre, Brazil.
authors.forename: Deborah S B S
authors.initials: DS
authors.lastname: Silva
authors.affiliation: School of Criminal Justice, University of Southern Mississippi, Hattiesburg, MS 39406, USA.
authors.forename: Kuppareddi
authors.initials: K
authors.lastname: Balamurugan
authors.affiliation: DNA Laboratory, Broward County Sheriff's Office, Fort Lauderdale, FL 33301, USA.
authors.forename: George
authors.initials: G
authors.lastname: Duncan
authors.affiliation: Faculty of Biosciences, Laboratory of Human and Molecular Genetics, PUCRS, 90619-900 Porto Alegre, Brazil.
authors.forename: Clarice S
authors.initials: CS
authors.lastname: Alho
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA. Electronic address: mccordb@fiu.edu.
authors.forename: Bruce
authors.initials: B
authors.lastname: McCord
mesh_terms: D001826:Body Fluids / Q000378:metabolism; D004247:DNA / Q000032:analysis* / Q000097:blood; D019175:DNA Methylation; D057890:Epigenomics; D053803:Forensic Genetics / Q000379:methods*; D006801:Humans; D006812:Humic Substances / Q000032:analysis; D008297:Male; D044367:Phase Transition; D060888:Real-Time Polymerase Chain Reaction; D012463:Saliva / Q000378:metabolism; D012661:Semen / Q000378:metabolism*; D013447:Sulfites / Q000737:chemistry; D044366:Transition Temperature
chemical_list: D006812:Humic Substances; D013447:Sulfites; D004247:DNA; C042345:hydrogen sulfite
keywords: Body fluid identification; DNA methylation; Epigenetic; Forensic; High-resolution melt; Real-time PCR