Exploring STR sequencing for forensic DNA intelligence databasing using the Austrian National DNA Database as an example

Abstract

Here, we present the results from a population study that evaluated the performance of massively parallel sequencing (MPS) of short tandem repeats (STRs) with a particular focus on DNA intelligence databasing purposes. To meet this objective, 247 randomly selected reference samples, earlier being processed with conventional capillary electrophoretic (CE) STR sizing from the Austrian National DNA Database, were reanalyzed with the PowerSeq 46Y kit (Promega). This sample set provides MPS-based population data valid for the Austrian population to increase the body of sequence-based STR variation. The study addressed forensically relevant parameters, such as concordance and backward compatibility to extant amplicon-based genotypes, sequence-based stutter ratios, and relative marker performance. Of the 22 autosomal STR loci included in the PowerSeq 46GY panel, 99.98% of the allele calls were concordant between MPS and CE. Moreover, 25 new sequence variants from 15 markers were found in the Austrian dataset that are yet undescribed in the STRSeq online catalogue and were submitted for inclusion. Despite the high degree of concordance between MPS and CE derived genotypes, our results demonstrate the need for a harmonized allele nomenclature system that is equally applicable to both technologies, but at the same time can take advantage of the increased information content of MPS. This appears to be particularly important with regard to database applications in order to prevent false exclusions due to varying allele naming based on different analysis platforms and ensures backward compatibility.

Accurate profiling of forensic autosomal STRs using the Oxford Nanopore Technologies MinION device

“…Despite the use of different amplification kits, these observations are consistent with other ONT-based STR studies [30]. Additionally, the loci identified as artifact-prone herein, namely D18S51 and D22S1045, were also noted in both of these studies [30,45].…”

“…We observed higher levels of both amplification bias and stutter compared to PowerSeq 46GY amplicon sequencing data produced on the Illumina MiSeq FGx [45]. Despite the use of different amplification kits, these observations are consistent with other ONT-based STR studies [30].…”

USAT: a bioinformatic toolkit to facilitate interpretation and comparative visualization of tandem repeat sequences

“…Further analysis within TR haplotypes could provide additional understanding and interpretation of TR variation and TR’s role in organisms. For example, 25 new sequence variants from 15 CODIS loci were found in an Austrian massive parallel sequencing dataset of 247 reference human samples via sequencing targeted STR loci compared with via a length-based CE method [ 26 ]. Such variants were undetectable with traditional CE methods, which do not accurately reflect the underlying sequence genotypes.…”

USAT: a Bioinformatic Toolkit to Facilitate Interpretation and Comparative Visualization of Tandem Repeat Sequences

“…Further analysis within TR haplotypes could provide additional understanding and interpretation of TR variation and TR’s role in organisms. For example, 25 new sequence variants from 15 CODIS loci were found in an Austrian MPS dataset of 247 reference human samples via sequencing targeted STR loci compared with a length-based CE method (Hölzl-Müller et al 2021). Such variants were undetectable with traditional CE methods which do not accurately reflect the underlying sequence genotypes.…”