High-throughput sequencing of core STR loci for forensic genetic investigations using the Roche Genome Sequencer FLX platform.

The analysis and profiling of short tandem repeat (STR) loci is routinely used in forensic genetics. Current methods to investigate STR loci, including PCR-based standard fragment analyses and capillary electrophoresis, only provide amplicon lengths that are used to estimate the number of STR repeat units. These methods do not allow for the full resolution of STR base composition that sequencing approaches could provide. Here we present an STR profiling method based on the use of the Roche Genome Sequencer (GS) FLX to simultaneously sequence multiple core STR loci. Using this method in combination with a bioinformatic tool designed specifically to analyze sequence lengths and frequencies, we found that GS FLX STR sequence data are comparable to conventional capillary electrophoresis-based STR typing. Furthermore, we found DNA base substitutions and repeat sequence variations that would not have been identified using conventional STR typing.

[1]  Charlotte L. Oskam,et al.  Identification of microsatellites from an extinct moa species using high-throughput (454) sequence data. , 2009, BioTechniques.

[2]  M. Wingfield,et al.  Microsatellite discovery by deep sequencing of enriched genomic libraries. , 2009, BioTechniques.

[3]  P. Jarne,et al.  Microsatellites, from molecules to populations and back. , 1996, Trends in ecology & evolution.

[4]  Bruce Budowle,et al.  Enhancing resolution and statistical power by utilizing mass spectrometry for detection of SNPs within the short tandem repeats , 2009 .

[5]  Marcus Droege,et al.  The Genome Sequencer FLX System--longer reads, more applications, straight forward bioinformatics and more complete data sets. , 2008, Journal of biotechnology.

[6]  N. Gemmell,et al.  Fast, cost-effective development of species-specific microsatellite markers by genomic sequencing. , 2009, BioTechniques.

[7]  N. Morling,et al.  Automated extraction of DNA and PCR setup using a Tecan Freedom EVO® liquid handler , 2009 .

[8]  A. Morales-Valverde,et al.  Characterisation of 12 new alleles in the STR system D18S51 , 2009 .

[9]  Soong-Deok Lee,et al.  Alterations of Length Heteroplasmy in Mitochondrial DNA Under Various Amplification Conditions * , 2010, Journal of forensic sciences.

[10]  Charlotte L. Oskam,et al.  Profiling the Dead: Generating Microsatellite Data from Fossil Bones of Extinct Megafauna—Protocols, Problems, and Prospects , 2011, PloS one.

[11]  M. Allen,et al.  Forensic analysis of autosomal STR markers using Pyrosequencing. , 2010, Forensic science international. Genetics.

[12]  James R. Knight,et al.  Genome sequencing in microfabricated high-density picolitre reactors , 2005, Nature.

[13]  Herbert Oberacher,et al.  Increasing the discrimination power of forensic STR testing by employing high-performance mass spectrometry, as illustrated in indigenous South African and Central Asian populations , 2010, International Journal of Legal Medicine.

[14]  W. Mayr,et al.  Unusual FGA and D19S433 off-ladder alleles and other allelic variants at the STR loci D8S1132, vWA, D18S51 and ACTBP2 (SE33) , 2008 .

[15]  J. Butler,et al.  STR sequence analysis for characterizing normal, variant, and null alleles. , 2011, Forensic science international. Genetics.

[16]  W. Mayr,et al.  Further allelic variation at the STR-loci ACTBP2 (SE33), D3S1358, D8S1132, D18S51 and D21S11 , 2009 .

[17]  Eric Buel,et al.  Forensic DNA typing by capillary electrophoresis using the ABI Prism 310 and 3100 genetic analyzers for STR analysis , 2004, Electrophoresis.