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Forensic analysis of autosomal STR markers using Pyrosequencing
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medicinsk genetik och genomik.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medicinsk genetik och genomik.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medicinsk genetik och genomik.
2010 (English)In: Forensic Science International: Genetics, ISSN 1872-4973, E-ISSN 1878-0326, Vol. 4, no 2, 122-129 p.Article in journal (Refereed) Published
Abstract [en]

Short tandem repeats (STRs) are highly variable, and therefore routinely used in forensic investigations for a DNA-based individual identification. The routine assay is commonly performed by size separation using capillary electrophoresis, but alternative technologies can also be used. In this study, a Pyrosequencing assay was developed for analysis of STR markers useful in forensic DNA analysis. The assay was evaluated for 10 different STR loci (CSF1PO, TH01, TPOX, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539 and Penta E) and a total of 114 Swedish individuals were genotyped. This genotyping strategy reveal the actual sequence and variant alleles were seen at several loci, providing additional information compared to fragment size analysis. At the D13S317 locus a T/A SNP located in the last repeat unit was observed in 92% of the genotypes. Moreover, an upstream flanking SNP at locus D7S820, a SNP within the repeats at D3S1358 and D8S1179 and a deletion in the flanking region at locus D5S818 were observed. The Pyrosequencing method was first developed for SNP typing and sequencing of shorter DNA fragments but the method also provides an alternative method for STR analysis of less complex repeats. This assay is suitable for investigation of new markers, a rapid compilation of population data and for confirmation of variant and new alleles.

Place, publisher, year, edition, pages
2010. Vol. 4, no 2, 122-129 p.
Keyword [en]
Forensic identification, Pyrosequencing, Sequence-directed nucleotide addition, STR
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-122745DOI: 10.1016/j.fsigen.2009.07.004ISI: 000274494800011PubMedID: 20129471OAI: oai:DiVA.org:uu-122745DiVA: diva2:310925
Available from: 2010-04-19 Created: 2010-04-19 Last updated: 2017-12-12
In thesis
1. Sensitive Identification Tools in Forensic DNA Analysis
Open this publication in new window or tab >>Sensitive Identification Tools in Forensic DNA Analysis
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

DNA as forensic evidence is valuable in criminal investigations. Implementation of new, sensitive and fast technologies is an important part of forensic genetic research. This thesis aims to evaluate new sensitive methods to apply in forensic DNA analysis including analysis of old skeletal remains.

In Paper I and II, two novel systems for analysis of STRs, based on the Pyrosequencing technology, are presented. In Paper I, Y chromosomal STRs are analysed. Markers on the male specific Y chromosome are especially useful in analysis of DNA mixtures. In Paper II, ten autosomal STRs are genotyped. The systems are based on sequencing of STR loci instead of size determination of STR fragments as in routine analysis. This provides a higher resolution since sequence variants within the repeats can be detected. Determination of alleles is based on a termination recognition base. This is the base in the template strand that is excluded from the dispensation order in the sequencing of the complementary strand and therefore terminates the reaction. Furthermore, skeletal remains are often difficult to analyse, due to damaging effects from the surrounding environment on the DNA and the high risk of exogenous contamination. Analysis of mitochondrial DNA is useful on degraded samples and in Paper III, mtDNA analysis of 700 years old skeletal remains is performed to investigate a maternal relationship. The quantity and quality of DNA are essential in forensic genetics. In Paper IV the efficiency of DNA isolation is investigated. Soaking skeletal remains in bleach is efficient for decontamination but result in a lower DNA yield, especially on pulverised skull samples.

In conclusion, this thesis presents novel sequencing systems for accurate and fast analysis of STR loci that can be useful in evaluation of new loci and database assembly as well as the utility of mtDNA in forensic genetics.

 

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2010. 49 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 606
Keyword
forensic genetics, STRs, Y-chromosome, Pyrosequencing, mitochondrial DNA, skeletal remains, DNA extraction
National Category
Medical and Health Sciences
Research subject
Medical Genetics
Identifiers
urn:nbn:se:uu:diva-131904 (URN)978-91-554-7915-2 (ISBN)
Public defence
2010-11-26, Rudbecksalen, Rudbeck Laboratory, Dag Hammarskjölds väg 20, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2010-11-04 Created: 2010-10-10 Last updated: 2011-01-13Bibliographically approved

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