Rapid screening method for male DNA by using the loop-mediated isothermal amplification assay

Area of Science:

• Forensic Science
• Molecular Biology
• Genetics

Background:

• Detecting male DNA in forensic samples is crucial for criminal investigations, especially in sexual assault cases.
• Current methods can be time-consuming and require specialized equipment.
• A need exists for rapid, on-site screening tools for male-derived biological material.

Purpose of the Study:

• To develop and validate a loop-mediated isothermal amplification (LAMP) assay for sensitive and specific detection of male DNA.
• To assess the assay's performance under various conditions, including sensitivity, specificity, and speed.
• To evaluate a simplified protocol for on-site forensic screening.

Main Methods:

• A loop-mediated isothermal amplification (LAMP) assay targeting multi-repeat Y-chromosome sequences was developed.
• The assay was optimized for temperature (61–67°C) and time (10–20 min).
• Sensitivity was tested down to picogram levels of male DNA, and specificity was confirmed against female DNA.

Main Results:

• The LAMP assay successfully amplified male DNA (0.5 ng) with no amplification of female DNA (up to 10 ng).
• The assay demonstrated high sensitivity, detecting as little as 1 pg of male DNA.
• A simplified protocol with DNA extraction and colorimetric detection processed samples in approximately 30 minutes.

Conclusions:

• The developed LAMP assay is a sensitive, specific, and rapid method for detecting male DNA.
• The simplified protocol allows for on-site screening of male-derived biological samples.
• This assay offers a promising tool for forensic investigations, requiring minimal hands-on time and equipment.