Comparisons of aged samples and modern references provide algorithm for mtDNA analysis in challenging material.

Area of Science:

• Forensic Genetics
• Human Identification
• Ancient DNA Analysis

Background:

• Analysis of low-quality DNA, particularly from aged bone samples, presents significant challenges in human identification.
• Existing genetic analysis methods often struggle with degraded samples, leading to discrepancies in haplotype comparisons.

Purpose of the Study:

• To develop and validate a new variant calling algorithm for improved analysis of low-quality mitochondrial DNA (mtDNA).
• To assess the efficacy of the new algorithm in human identification cases using historical bone samples.

Main Methods:

• Comparative analysis of haplotypes from 70-80 year old bone samples against modern references.
• Whole mitochondrial genome sequencing using Thermo Fisher Scientific's Precision ID and Ion GeneStudio S5.
• Development of a novel variant calling algorithm by altering parameters of Converge 2.2 and IGV 2.12.3.

Main Results:

• Standard analysis settings were insufficient for poor quality DNA data, with minimal improvement from threshold changes or contaminant read removal.
• The developed algorithm effectively aids in accepting genetic calls from difficult, low-quality samples.
• An additional step was introduced for samples with low region coverage, enhancing data interpretation.

Conclusions:

• Standard genetic analysis protocols are rarely suitable for degraded DNA samples.
• The new variant calling algorithm reduces manual labor and improves the reliability of human identification from challenging genetic materials.
• The developed protocol is applicable to various fields requiring DNA analysis with low quantity and quality.