Sequence Variations of 31 Υ-Chromosomal Short Tandem Repeats Analyzed by Massively Parallel Sequencing in Three U.S. Population Groups and Korean Population.

BACKGROUND: Rapidly mutating (RM) Y-chromosomal short tandem repeats (Y-STRs) have been demonstrated to increase the possibility of distinguishing between male relatives due to a higher mutation rate than conventional Y-STRs. Massively parallel sequencing (MPS) can be useful for forensic DNA typing as it allows the detection of sequence variants of many forensic markers. Here, we present sequence variations of 31 Y-STRs including nine RM Y-STRs (DYF387S1, DYF399S1, DYF404S1, DYS449, DYS518, DYS570, DYS576, DYS612, and DYS627), their frequencies, distribution, and the gain in the number of alleles using MPS. METHODS: We constructed a multiplex MPS assay capable of simultaneously amplifying 32 Y-chromosomal markers, producing amplicons ranging from 85-274 bp. Barcoded libraries from 220 unrelated males from four populations-African Americans, Caucasians, Hispanics, and Koreans-were generated via two-step polymerase chain reaction and sequenced on a MiSeq system. Genotype concordance between the capillary electrophoresis (CE) and MPS method and sequence variation of Y-STRs were investigated. RESULTS: In total, 195 alleles were increased by MPS compared to CE-based alleles (261 to 456). The DYS518 marker showed the largest increase due to repeat region variation (a 3.69-fold increase). The highest increase in the number of alleles due to single nucleotide polymorphisms in the flanking region was found in DYF399S1. RM Y-STRs had more diverse sequences than conventional Y-STRs. Furthermore, null alleles were observed in DYS576 due to primer-binding site mutation, and allele drop-outs in DYS449 resulted from low marker coverage of less than the threshold. CONCLUSION: The results suggest that the expanded and discriminative MPS assay could provide more genetic information for Y-STRs, especially for RM Y-STRs, and could advance male individualization. Compiling sequence-based Y-STR data for worldwide populations would facilitate the application of MPS in the field of forensic genetics and could be applicable in solving male-related forensic cases.

Genetic investigation of 124 SNPs in a Myanmar population using the Precision ID Identity Panel and the Illumina MiSeq.

Single nucleotide polymorphisms (SNPs) have become popular in forensic genetics as an alternative to short tandem repeats (STRs). The Precision ID Identity Panel (Thermo Fisher Scientific), consisting of 90 autosomal SNPs and 34 Y-chromosomal SNPs, enabled human identification studies on global populations through next-generation sequencing (NGS). However, most previous studies on the panel have used the Ion Torrent platform, and there are few reports on the Southeast Asian population. Here, a total of 96 unrelated males from Myanmar (Yangon) were analyzed with the Precision ID Identity Panel on a MiSeq (Illumina) using an in-house TruSeq compatible universal adapter and a custom variant caller, Visual SNP. The sequencing performance evaluated by locus balance and heterozygote balance was comparable to that of the Ion Torrent platform. For 90 autosomal SNPs, the combined match probability (CMP) was 6.994 × 10-34, lower than that of 22 PowerPlex Fusion autosomal STRs (3.130 × 10-26). For 34 Y-SNPs, 14 Y-haplogroups (mostly O2 and O1b) were observed. We found 51 cryptic variations (42 haplotypes) around target SNPs, of which haplotypes corresponding to 33 autosomal SNPs decreased CMP. Interpopulation analysis revealed that the Myanmar population is genetically closer to the East and Southeast Asian populations. In conclusion, the Precision ID Identity Panel can be successfully analyzed on the Illumina MiSeq and provides high discrimination power for human identification in the Myanmar population. This study broadened the accessibility of the NGS-based SNP panel by expanding the available NGS platforms and adopting a robust NGS data analysis tool.