title: Short-read, high-throughput sequencing technology for STR genotyping.
abstract: DNA-based methods for human identification principally rely upon genotyping of short tandem repeat (STR) loci. Electrophoretic-based techniques for variable-length classification of STRs are universally utilized, but are limited in that they have relatively low throughput and do not yield nucleotide sequence information. High-throughput sequencing technology may provide a more powerful instrument for human identification, but is not currently validated for forensic casework. Here, we present a systematic method to perform high-throughput genotyping analysis of the Combined DNA Index System (CODIS) STR loci using short-read (150 bp) massively parallel sequencing technology. Open source reference alignment tools were optimized to evaluate PCR-amplified STR loci using a custom designed STR genome reference. Evaluation of this approach demonstrated that the 13 CODIS STR loci and amelogenin (AMEL) locus could be accurately called from individual and mixture samples. Sensitivity analysis showed that as few as 18,500 reads, aligned to an in silico referenced genome, were required to genotype an individual (>99% confidence) for the CODIS loci. The power of this technology was further demonstrated by identification of variant alleles containing single nucleotide polymorphisms (SNPs) and the development of quantitative measurements (reads) for resolving mixed samples.
authors.affiliation: Battelle Memorial Institute, Columbus, OH, USA.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Bornman
authors.affiliation: Battelle Memorial Institute, Charlottesville, VA, USA.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Hester
authors.affiliation: Human Cancer Genetics Program, The Ohio State University Comprehensive Cancer Center, Columbus, OH, USA.
authors.forename: Jared M
authors.initials: JM
authors.lastname: Schuetter
authors.affiliation: Department of Physics and Biochemistry, Center for RNA Biology, The Ohio State University, Columbus, OH, USA.
authors.forename: Manjula D
authors.initials: MD
authors.lastname: Kasoji
authors.forename: Angela
authors.initials: A
authors.lastname: Minard-Smith
authors.forename: Curt A
authors.initials: CA
authors.lastname: Barden
authors.forename: Scott C
authors.initials: SC
authors.lastname: Nelson
authors.forename: Gene D
authors.initials: GD
authors.lastname: Godbold
authors.forename: Christine H
authors.initials: CH
authors.lastname: Baker
authors.forename: Boyu
authors.initials: B
authors.lastname: Yang
authors.forename: Jacquelyn E
authors.initials: JE
authors.lastname: Walther
authors.forename: Ivan E
authors.initials: IE
authors.lastname: Tornes
authors.forename: Pearlly S
authors.initials: PS
authors.lastname: Yan
authors.forename: Benjamin
authors.initials: B
authors.lastname: Rodriguez
authors.forename: Ralf
authors.initials: R
authors.lastname: Bundschuh
authors.forename: Michael L
authors.initials: ML
authors.lastname: Dickens
authors.forename: Brian A
authors.initials: BA
authors.lastname: Young
authors.forename: Seth A
authors.initials: SA
authors.lastname: Faith
keywords: Bridge PCR; Illumina; SNP; STR; forensic; genotyping; high-throughput sequencing; next-generation sequencing