Investigating the detection of two-person DNA mixtures using tri-allelic InDels.

Area of Science:

• Forensic Science
• Genetics
• Molecular Biology

Background:

• Analyzing complex DNA mixtures presents significant challenges in forensic science.
• Current methods struggle with unbalanced or degraded samples.
• Insertion/Deletion (InDel) markers offer advantages due to short amplicons and resistance to PCR slippage.

Purpose of the Study:

• To evaluate the performance of tri-allelic InDels for analyzing two-person DNA mixtures.
• To develop a multiplex assay for enhanced DNA mixture analysis.
• To assess the utility of tri-allelic InDels in forensic investigations.

Main Methods:

• Characterization of 38 tri-allelic InDels with specific length differences.
• Development of a multiplex amplification assay (FA-HID39plex) including the amelogenin sex identification marker.
• Evaluation of tri-allelic InDels in simulated two-person DNA mixtures of varying ratios and degradation levels.

Main Results:

• Tri-allelic InDels generated higher likelihood ratios (LRs) compared to biallelic markers for inclusion assessment.
• The FA-HID39plex assay detected minor contributors in non-degraded mixtures at ratios as low as 1:29.
• While allele dropout affected sensitivity in degraded mixtures, LRs still provided moderate support for inclusion.

Conclusions:

• Tri-allelic InDels are effective supplementary markers for analyzing unbalanced and degraded two-person DNA mixtures.
• The FA-HID39plex assay enhances the ability to identify individuals in complex forensic samples.
• LR calculations using tri-allelic InDels aid in the investigation of persons of interest (POI).