STR melting curve analysis as a genetic screening tool for crime scene samples

Area of Science:

• Forensic Genetics
• Molecular Biology
• Analytical Chemistry

Background:

• Accurate human DNA profiling is crucial in forensic science.
• Short tandem repeat (STR) analysis is a standard method for DNA profiling.
• Efficient screening methods are needed to assess sample quality and potential value prior to full genotyping.

Purpose of the Study:

• To evaluate high-resolution melt curve (HRMC) analysis as a post-quantification screening tool.
• To determine the ability of HRMC to discriminate between different human CSF1PO and THO1 genotypes.
• To assess HRMC's utility in differentiating single-source and mixed DNA samples.

Main Methods:

• Amplification of CSF1PO and HUMTHO1 loci using mini-STR primers.
• Analysis of amplified products using high-resolution melt curve analysis with SYBR Green or LCGreen Plus dyes.
• Utilized LightScanner HR96 and LS-32 systems for melt curve acquisition and analysis.

Main Results:

• Successfully differentiated 12 CSF1PO and 11 HUMTHO1 genotypes based on unique melt profiles.
• STR amplicon melt curves demonstrated sensitivity to repeat number variations.
• Heteroduplex formation enabled differentiation between single-source and mixed DNA samples.
• Reproducible and unique melting curves were obtained from 20 to 0.4 ng of DNA.
• Identical and non-identical genotypes from various sample types were distinguished.

Conclusions:

• High-resolution melt curve analysis is a viable post-quantification screening tool for human STR genotyping.
• HRMC can efficiently discriminate between different CSF1PO and THO1 genotypes.
• This method provides a means to assess sample quantity and probative value, potentially reducing the need for full genotyping in certain scenarios.