Routine Mitogenome MPS Analysis from 1 and 5 mm of Rootless Human Hair

Area of Science:

• Forensic Science
• Genetics
• Anthropology

Background:

• Hair shafts are common forensic evidence but often lack sufficient DNA for analysis.
• Mitochondrial DNA (mtDNA) sequencing is preferred for rootless hair due to high copy numbers and degraded nuclear DNA.

Purpose of the Study:

• To investigate the impact of hair age and physical characteristics on mtDNA recovery and sequencing quality using massively parallel sequencing (MPS).
• To assess the reliability of MPS for generating mitogenome sequences from small hair samples.

Main Methods:

• Microscopic characterization of hair samples (diameter, medullary structure, length) from 60 donors.
• Generation of mitogenome sequences using the Promega PowerSeq Whole Mito System and Illumina MiSeq.
• Analysis of mtDNA recovery, degradation, and MPS data quality in relation to hair age.

Main Results:

• Reportable mitogenome sequences were obtained from all hairs up to 27 years old.
• MPS yielded high-quality data (≥88% mitogenome coverage) for most samples, with concordant haplotypes from replicate segments.
• Hair age negatively correlated with mtDNA yield and data quality, while diameter and medullary structure had minimal impact.

Conclusions:

• MPS is a robust method for routine mitogenome sequencing from small hair shaft samples (1-5 mm) up to 27 years old.
• This technique offers significant potential for forensic investigations, biological anthropology, and medical genetics.
• The study validates MPS as a reliable tool for analyzing degraded DNA evidence from hair samples.