Identification of STR/Y-STR alleles with tolerance for variants and stutter detection using GeneMarker®HTS software

Abstract

GeneMarker®HTS is a rapid, user-friendly software for forensic mtDNA and STR analysis with high throughput sequencing (HTS)/ next generation sequencing (NGS)/ massively parallel sequencing (MPS) data. Compared to the traditional capillary electrophoresis (CE) allele separation, HTS data provides a precise description of the repeat allele structure for each STR locus including the variants in the flanking areas for iso-allele genotype reporting and for future use in probabilistic genotyping analyses. The variants in the repeat region or the flanking area may in some cases cause failure for identification of STR/Y-STR alleles. To increase the accuracy, an iterated sequence alignment method is used to detect alleles. First, we align sequences to GRCh38 to determine the repeat sequence. Then, using the repeat sequence, we perform a second alignment that minimizes alignment errors and improves accuracy for repeat number. Stutters are also identified by stutter filters. To validate the method, concordance study is made and the concordance with the CE allele calls for 22 Autosomal STR Loci, 22 Chr Y STR Loci and Amelogenin is 100 % for NIST-2391d samples and 99.93 % for a 651 sample NIST-Promega dataset. The identification error due to sequence variants coupled with STR repeats is solved.