title: Construction of a library of cloned short tandem repeat (STR) alleles as universal templates for allelic ladder preparation.
abstract: Short tandem repeat (STR) genotyping methods are widely used for human identity testing applications, including forensic DNA analysis. Samples of DNA containing the length-variant STR alleles are typically separated and genotyped by comparison to an allelic ladder. Here, we describe a newly devised library of cloned STR alleles. The library covers alleles X and Y for the sex-determining locus Amelogenin and 259 other alleles for 22 autosomal STR loci (TPOX, D3S1358, FGA, D5S818, CSF1PO, D7S820, D8S1179, TH01, vWA, D13S317, D16S539, D18S51, D21S11, D2S1338, D6S1043, D12S391, Penta E, D19S433, D11S4463, D17S974, D3S4529 and D12ATA63). New primers were designed for all these loci to construct recombinant plasmids so that the library retains core repeat elements of STR as well as 5'- and 3'-flanking sequences of ∼500 base pairs. Since amplicons of commercial STR genotyping kits and systems developed in laboratories are usually distributed from 50 to <500 base pairs, this library could provide universal templates for allelic ladder preparation. We prepared three different sets of allelic ladders for this locus TH01 and an updated version of an allelic ladder for the DNATyper(®)19 multiplex system using these plasmids to confirm the suitability of the library as a good source for allelic ladder preparation. Importantly, the authenticity of each construct was confirmed by bidirectional nucleotide sequencing and we report the repeat structures of the 259 STR alleles. The sequencing results showed all repeat structures we obtained for TPOX, CSF1PO, D7S820, TH01, D16S539, D18S51 and Penta E were the same as reported. However, we identified 102 unreported repeat structures from the other 15 STR loci, supplementing our current knowledge of repeat structures and leading to further understanding of these widely used loci.
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China. Electronic address: wangle_02@163.com.
authors.forename: Le
authors.initials: L
authors.lastname: Wang
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China. Electronic address: zhaoxchun@sina.com.
authors.forename: Xing-Chun
authors.initials: XC
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China. Electronic address: yejian77@126.com.
authors.forename: Jian
authors.initials: J
authors.lastname: Ye
authors.affiliation: School of Forensic Medicine, Shanxi Medical University, Taiyuan 030001, PR China.
authors.forename: Jin-Jie
authors.initials: JJ
authors.lastname: Liu
authors.affiliation: School of Forensic Medicine, Shanxi Medical University, Taiyuan 030001, PR China.
authors.forename: Ting
authors.initials: T
authors.lastname: Chen
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China.
authors.forename: Xue
authors.initials: X
authors.lastname: Bai
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China.
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Ou
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China.
authors.forename: Lan
authors.initials: L
authors.lastname: Hu
authors.affiliation: The First Research Institute of the Ministry of Public Security, Beijing 100048, PR China.
authors.forename: Bo-Wei
authors.initials: BW
authors.lastname: Jiang
authors.affiliation: Key Laboratory of Forensic Genetics, Institute of Forensic Science, Ministry of Public Security, Beijing 100038, PR China.
authors.forename: Feng
authors.initials: F
authors.lastname: Wang
mesh_terms: D000483:Alleles; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D053803:Forensic Genetics; D018895:Microsatellite Repeats; D016133:Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid
chemical_list: D017931:DNA Primers
keywords: Allelic ladder; Forensic science; Recombinant plasmid; Repeat structure; Short tandem repeat (STR)