title: A method to determine the 5' end of the binding site of primers included in a commercially available forensic human identification kit.
abstract: Analysis for short tandem repeat (STR) loci is widely performed in forensic laboratories for human identification that utilizes commercially available kits that employ fluorescently labeled primers and capillary electrophoresis. With only a few exceptions, the sequences of the primers included in a kit are not disclosed by the kit manufacturers. Therefore, we developed a simple method to determine the 5' end of the binding site of the primers included in commercial kits. Our method requires only custom primers and human genome sequence data and routinely used equipment and consumables. One or two custom primers are added to the PCR reaction mixture containing kit primers and input human DNA prior to amplification, and PCR products are separated by capillary electrophoresis after amplification. With this method we can determine which primer of the pair is fluorescently labeled and the 5' end of the binding site of primers based on the changes in an electropherogram that are caused by the addition of the custom primer(s), and the human genome sequence data. This method is also useful for the determination of the shortest possible lengths of labeled kit primers.
authors: First Department of Forensic Science, National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa 277-0882, Japan. Electronic address: mizuno@nrips.go.jp.
authors: Natsuko
authors: N
authors: Mizuno
authors: First Department of Forensic Science, National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa 277-0882, Japan.
authors: Shota
authors: S
authors: Inokuchi
authors: First Department of Forensic Science, National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa 277-0882, Japan.
authors: Tetsushi
authors: T
authors: Kitayama
authors: First Department of Forensic Science, National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa 277-0882, Japan.
authors: Koji
authors: K
authors: Fujii
authors: First Department of Forensic Science, National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa 277-0882, Japan.
authors: Kentaro
authors: K
authors: Kasai
authors: First Department of Forensic Science, National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa 277-0882, Japan.
authors: Kazumasa
authors: K
authors: Sekiguchi
mesh_terms: D024506:5' Flanking Region; D001665:Binding Sites; D016172:DNA Fingerprinting; D017931:DNA Primers; D019075:Electrophoresis, Capillary; D019793:Fluorescein; D005456:Fluorescent Dyes; D006801:Humans; D018895:Microsatellite Repeats; D060885:Multiplex Polymerase Chain Reaction
chemical_list: D017931:DNA Primers; D005456:Fluorescent Dyes; D019793:Fluorescein
keywords: 3′ end of the binding site of a primer; AmpFlSTR(®) 5′ end of the binding site of a primer; Forensic; PCR primer; PowerPlex(®)