Linking STRs/SNPs and DNA methylation using massively parallel sequencing for potential forensic applications

Area of Science:

• Forensic Science
• Genetics
• Epigenetics

Background:

• DNA methylation analysis is crucial for forensic trace analysis, particularly for linking individuals to body fluids.
• Combining tissue-specific differentially methylated DNA positions (tDMPs) with person-identifying single nucleotide polymorphisms (iSNPs) shows promise for mixture analysis.
• Short tandem repeats (STRs) are the established standard for individual identification in forensics.

Purpose of the Study:

• To develop a pipeline for STR analysis of bisulfite-converted DNA.
• To characterize DNA methylation sites near STRs for potential body fluid-specific variations.
• To evaluate the utility of tDMPs and iSNPs for linking mixture contributors to body fluids.

Main Methods:

• Analysis of forensically relevant STR regions in various body fluids (blood, semen, saliva, etc.) using amplicon-based bisulfite massively parallel sequencing.
• Evaluation of DNA methylation at known tDMPs and surrounding iSNPs in blood, semen, saliva, and vaginal fluid.
• Utilized FDSTools with a new library for STR naming and allele-linked DNA methylation analysis.

Main Results:

• Successful STR analysis from bisulfite-converted DNA was achieved for 18 out of 22 STRs.
• Identified STR flanking sites with body fluid-varying DNA methylation (e.g., D1S1677, FGA), though not sufficient for exclusive body fluid identification.
• Demonstrated the potential of STR analysis for contributor assignment in mixtures, especially those involving semen.

Conclusions:

• The developed pipeline enables STR analysis from bisulfite-converted DNA, facilitating allele-linked DNA methylation analysis.
• Confirmed tDMPs and linked iSNPs are valuable for contributor assignment in forensic mixtures.
• The methodology offers a flexible tool for diverse research applications beyond forensic science.