HRM and SNaPshot as alternative forensic SNP genotyping methods.

Area of Science:

• Forensic genetics
• Molecular biology

Background:

• Single nucleotide polymorphisms (SNPs) are crucial for forensic applications like identity, biogeographical ancestry (BGA), and externally visible characteristics (EVCs).
• Single base extension (SBE) assays, such as SNaPshot®, are standard for forensic SNP genotyping using capillary electrophoresis.
• High Resolution Melt (HRM) analysis presents an alternative, rapid, single-tube method for SNP typing.

Purpose of the Study:

• To compare the performance of High Resolution Melt (HRM) analysis and SNaPshot® for forensic SNP genotyping.
• To evaluate the reliability, reproducibility, and limitations of each method.

Main Methods:

• Comparison of HRM and SNaPshot® assays for genotyping single nucleotide polymorphisms (SNPs).
• Assessment of genotyping reproducibility at different DNA input levels (e.g., 500 pg for HRM, 100 pg for SNaPshot®).
• Evaluation of assay performance concerning SNP characteristics like GC content and symmetry.

Main Results:

• HRM demonstrated reproducible genotyping at 500 pg DNA but faced challenges with high GC content and symmetrical SNPs.
• SNaPshot® showed reproducibility at 100 pg DNA and was less sensitive to SNP selection.
• HRM offered faster processing times and reduced post-PCR contamination risk compared to SNaPshot®.

Conclusions:

• Both HRM and SNaPshot® can be used for forensic SNP genotyping, with distinct advantages and limitations.
• SNaPshot® provides broader applicability across different SNP types.
• HRM shows potential as a rapid screening tool in forensic genetics due to its speed and reduced contamination risk.