Application of a custom haplotype caller to analyze sequence-based data of 56 microhaplotypes

Area of Science:

• Forensic Genetics
• Molecular Biology
• Bioinformatics

Background:

• Conventional forensic markers like STRs and SNPs have limitations.
• Massively parallel sequencing (MPS) shows promise for microhaplotypes (microhaps) in forensics.
• Simpler workflows are needed for practical MPS application in forensic science.

Purpose of the Study:

• To develop an in-house MPS panel for 56 microhaps.
• To create a user-friendly haplotype caller (Visual Microhap) for MPS data.
• To establish a streamlined workflow for microhap analysis in forensics.

Main Methods:

• Developed a 56-plex MPS panel for simultaneous microhap amplification.
• Created Visual Microhap, a web-based tool for SNP-based haplotype extraction.
• Analyzed 286 samples from four populations using the panel and workflow.

Main Results:

• The 56-plex microhap panel demonstrated higher forensic identification power than autosomal STRs.
• Average effective number of alleles (Ae) was 3.45, with a range of 1.74–6.98.
• Cluster analysis revealed distinct population groupings based on the microhap data.

Conclusions:

• The developed MPS panel and open-source workflow enhance microhap utility in forensic research.
• This approach offers a more powerful tool for human identification in forensic applications.
• The streamlined workflow facilitates broader adoption of MPS for microhap analysis.