title: Massively parallel sequencing of short tandem repeats-Population data and mixture analysis results for the PowerSeq™ system.
abstract: Current forensic DNA analysis predominantly involves identification of human donors by analysis of short tandem repeats (STRs) using Capillary Electrophoresis (CE). Recent developments in Massively Parallel Sequencing (MPS) technologies offer new possibilities in analysis of STRs since they might overcome some of the limitations of CE analysis. In this study 17 STRs and Amelogenin were sequenced in high coverage using a prototype version of the Promega PowerSeq™ system for 297 population samples from the Netherlands, Nepal, Bhutan and Central African Pygmies. In addition, 45 two-person mixtures with different minor contributions down to 1% were analysed to investigate the performance of this system for mixed samples. Regarding fragment length, complete concordance between the MPS and CE-based data was found, marking the reliability of MPS PowerSeq™ system. As expected, MPS presented a broader allele range and higher power of discrimination and exclusion rate. The high coverage sequencing data were used to determine stutter characteristics for all loci and stutter ratios were compared to CE data. The separation of alleles with the same length but exhibiting different stutter ratios lowers the overall variation in stutter ratio and helps in differentiation of stutters from genuine alleles in mixed samples. All alleles of the minor contributors were detected in the sequence reads even for the 1% contributions, but analysis of mixtures below 5% without prior information of the mixture ratio is complicated by PCR and sequencing artefacts.
authors.affiliation: Forensic Laboratory for DNA Research, Department of Human Genetics, Leiden University Medical Centre, Postzone S 05 P, P.O. Box 9600, 2300 RC Leiden, The Netherlands; Biological Traces, Netherlands Forensic Institute, Laan van Ypenburg 6, 2497GB The Hague, The Netherlands. Electronic address: k.van.der.gaag@nfi.minvenj.nl.
authors.forename: Kristiaan J
authors.initials: KJ
authors.lastname: van der Gaag
authors.affiliation: Forensic Laboratory for DNA Research, Department of Human Genetics, Leiden University Medical Centre, Postzone S 05 P, P.O. Box 9600, 2300 RC Leiden, The Netherlands. Electronic address: r.h.de_leeuw@lumc.nl.
authors.forename: Rick H
authors.initials: RH
authors.lastname: de Leeuw
authors.affiliation: Biological Traces, Netherlands Forensic Institute, Laan van Ypenburg 6, 2497GB The Hague, The Netherlands; Department of Human Genetics, Leiden University Medical Centre, 2300 RC Leiden, The Netherlands. Electronic address: j.hoogenboom@nfi.minvenj.nl.
authors.forename: Jerry
authors.initials: J
authors.lastname: Hoogenboom
authors.affiliation: Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA. Electronic address: Jaynish.Patel@promega.com.
authors.forename: Jaynish
authors.initials: J
authors.lastname: Patel
authors.affiliation: Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA. Electronic address: doug.storts@promega.com.
authors.forename: Douglas R
authors.initials: DR
authors.lastname: Storts
authors.affiliation: Department of Human Genetics, Leiden University Medical Centre, 2300 RC Leiden, The Netherlands; Leiden Genome Technology Centre, 2300 RC Leiden, The Netherlands; Netherlands Bioinformatics Centre, Leiden, The Netherlands. Electronic address: j.f.j.laros@lumc.nl.
authors.forename: Jeroen F J
authors.initials: JFJ
authors.lastname: Laros
authors.affiliation: Forensic Laboratory for DNA Research, Department of Human Genetics, Leiden University Medical Centre, Postzone S 05 P, P.O. Box 9600, 2300 RC Leiden, The Netherlands. Electronic address: p.de_knijff@lumc.nl.
authors.forename: Peter
authors.initials: P
authors.lastname: de Knijff
mesh_terms: D000350:Africa, Central; D053523:Amelogenin / Q000235:genetics; D001211:Asia, Western; D005828:Genetics, Population; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D018895:Microsatellite Repeats; D009426:Netherlands; D044469:Racial Groups / Q000235:genetics; D017422:Sequence Analysis, DNA
chemical_list: D053523:Amelogenin
keywords: Bioinformatics; Forensic science; Illumina; Massively Parallel Sequencing (MPS); MiSeq; Mixture analysis; Next Generation Sequencing (NGS); PowerSeq; Promega; STR stutter; Sequence variants; Short tandem repeat (STR); TSSV; fdstools