A novel biosensor for highly sensitive DNA damage detection using TdT and CRISPR-Cas12a

Area of Science:

• Biotechnology
• Molecular Biology
• Genetics

Background:

• Sperm DNA damage impacts male fertility and reproductive outcomes.
• Accurate and sensitive detection methods are crucial for clinical diagnostics and reproductive medicine.

Purpose of the Study:

• To develop a highly sensitive fluorescence biosensor for precise detection of sperm DNA damage.
• To integrate terminal deoxynucleotidyl transferase (TdT) enzyme with CRISPR-Cas12a technology for enhanced detection capabilities.

Main Methods:

• Development of a fluorescence biosensor system.
• Integration of TdT enzyme for labeling DNA strand breaks.
• Utilizing CRISPR-Cas12a collateral activity for signal amplification and detection.
• Characterization of biosensor sensitivity and linear detection range.

Main Results:

• Achieved a highly sensitive detection limit of 0.99 pM for sperm DNA damage.
• Established a linear detection range from 0.001 nM to 0.2 nM.
• Demonstrated exceptional sensitivity, simplicity, and versatility of the developed biosensor.

Conclusions:

• The TdT and CRISPR-Cas12a integrated biosensor is a transformative tool for sperm DNA damage detection.
• This method offers significant potential for advancing reproductive medicine and clinical diagnostics.
• The biosensor's high sensitivity and ease of use pave the way for improved fertility assessments.