Enhanced mixture interpretation with macrohaplotypes based on long-read DNA sequencing

Area of Science:

• Forensic Genetics
• Genomics
• Molecular Biology

Background:

• DNA mixture interpretation is a significant challenge in forensic science.
• Current methods using Short Tandem Repeats (STRs) have limitations with complex mixtures.
• Novel markers like Single Nucleotide Variants (SNVs) and microhaplotypes lack discrimination power and CODIS compatibility.

Purpose of the Study:

• To develop a novel CODIS-compatible forensic marker for enhanced DNA mixture interpretation.
• To leverage Long-Read Sequencing (LRS) technologies for improved forensic analysis.
• To design a panel of highly discriminative macrohaplotypes.

Main Methods:

• Development of macrohaplotypes by combining CODIS STRs with flanking variants.
• Utilizing Long-Read Sequencing (LRS) to obtain large DNA fragments with definitive phasing.
• Designing a panel of 20 macrohaplotypes (approx. 8 kbp) with high discrimination power using publicly available data.

Main Results:

• The developed macrohaplotypes demonstrate significantly higher discrimination power compared to CODIS STRs.
• Macrolhaplotypes show substantial improvement in interpreting complex DNA mixtures, particularly those with multiple contributors.
• Statistical evaluation confirms the superior performance of macrohaplotypes for forensic applications.

Conclusions:

• Macrolhaplotypes represent a significant advancement in forensic DNA analysis, particularly for challenging mixture samples.
• The study highlights the potential of LRS technologies in developing next-generation forensic markers.
• Further development of a complete wet-lab and bioinformatics workflow is recommended for macrohaplotype implementation.