Shotgun DNA sequencing for human identification: Dynamic SNP selection and likelihood ratio calculations accounting for errors

TL;DR

This paper introduces a statistical framework and an open-source R package for calculating likelihood ratios in forensic DNA analysis using shotgun sequencing, effectively handling low-quality samples and genotyping errors.

Contribution

It presents a novel model for shotgun sequencing data analysis, dynamic SNP selection, and error-aware likelihood ratio calculation for forensic identification.

Findings

Accurate error probability estimation improves LR calculations.

Method effectively analyzes low-quality and degraded DNA samples.

Open-source R package wgsLR facilitates practical application.

Abstract

In forensic genetics, short tandem repeats (STRs) are used for human identification (HID). Degraded biological trace samples with low amounts of short DNA fragments (low-quality DNA samples) pose a challenge for STR typing. Predefined single nucleotide polymorphisms (SNPs) can be amplified on short PCR fragments and used to generate SNP profiles from low-quality DNA samples. However, the stochastic results from low-quality DNA samples may result in frequent locus drop-outs and insufficient numbers of SNP genotypes for convincing identification of individuals. Shotgun DNA sequencing potentially analyses all DNA fragments in a sample in contrast to the targeted PCR-based sequencing methods and may be applied to DNA samples of very low quality, like heavily compromised crime-scene samples and ancient DNA samples. Here, we developed a statistical model for shotgun sequencing, sequence alignment, and genotype calling. Results from replicated shotgun sequencing of buccal swab (high-quality samples) and hair samples (low-quality samples) were arranged in a genotype-call confusion matrix to estimate the calling error probability by maximum likelihood and Bayesian inference. We developed formulas for calculating the evidential weight as a likelihood ratio (LR) based on data from dynamically selected SNPs from shotgun DNA sequencing. The method accounts for potential genotyping errors. Different genotype quality filters may be applied to account for genotyping errors. An error probability of zero resulted in the forensically commonly used LR formula. When considering a single SNP marker's contribution to the LR, error probabilities larger than zero reduced the LR contribution of matching genotypes and increased the LR in the case of a mismatch. We developed an open-source R package, wgsLR, which implements the method, including estimating the calling error probability and calculating LR values.