Typing of semen-containing mixtures using ARMS-based semen-specific CpG-InDel/STR markers

Area of Science:

• Forensic genetics
• Epigenetics
• Molecular biology

Background:

• Identifying mixed biological traces at crime scenes is a significant forensic genetics challenge.
• DNA methylation offers a promising avenue for body fluid identification, complementing traditional length polymorphic markers for individual identification.

Purpose of the Study:

• To develop and validate a novel panel of CpG-InDel/STR compound markers for simultaneous body fluid and individual identification.
• To assess the panel's efficacy in identifying semen, including in mixed stains and degraded samples.

Main Methods:

• Tissue-specific CpG sites were analyzed using bisulfite conversion and amplification refractory mutation system-multiprimer-PCR.
• Linked insertion or deletion (InDel) or short tandem repeat (STR) markers were analyzed via capillary electrophoresis.
• A panel of 17 semen-specific CpG-InDel/STR markers, 3 body-fluid controls, and 1 gender locus was constructed.

Main Results:

• The panel achieved full genotyping with 50 ng DNA input.
• Efficient identification of semen was achieved from samples stored for 7 months and heat-degraded samples (up to 6 hours).
• The panel successfully identified semen in mixed stains with as low as 3.03% semen content, demonstrating a cumulative discrimination power of 0.9999998.

Conclusions:

• A robust panel of CpG-InDel/STR markers was successfully developed for semen identification and individual profiling.
• This panel is effective for validating tissue source and individual identity in semen, even in challenging mixed and degraded samples.
• The developed panel offers a valuable alternative solution for forensic case investigations.