Postmortem CYP2D6 Genotyping and Copy Number Determinations Using DNA Extracted from Archived FTA Bloodstains

Area of Science:

• Forensic Science
• Pharmacogenetics
• Molecular Biology

Background:

• Post-mortem genetic characterization, specifically of the CYP2D6 gene, is crucial for investigating drug involvement in the cause of death.
• Dried blood spot samples archived on FTA cards at autopsy present a valuable resource for retrospective genetic analysis.

Purpose of the Study:

• To describe and validate methods for DNA extraction, CYP2D6 genotyping, and copy number variation (CNV) testing from post-mortem blood samples stored on FTA cards.
• To assess the feasibility and success rate of commercially available assays for this purpose.

Main Methods:

• DNA extraction from 3mm punches of bloodstained FTA cards obtained from autopsy cases.
• Utilizing the iPLEX® genotyping assay and MassARRAY platform (Agena Bioscience) for CYP2D6 allele detection.
• Performing copy number variation (CNV) analysis to quantify CYP2D6 gene copies.

Main Results:

• Successful DNA extraction yielded 9-100 ng/μL with acceptable purity (260/280 ratio 1.2-2.0).
• Identified various CYP2D6 alleles, including *2A, *3, *4, *5, *6, *10, *29, *35, and *41.
• Detected increased CYP2D6 copy numbers in 11 out of 75 samples, with CNV analysis failing in six.

Conclusions:

• Commercially available methods are effective for CYP2D6 genotyping and CNV testing using post-mortem blood archived on FTA cards.
• These validated methods can reliably support forensic investigations by providing critical genetic information related to drug metabolism.