DNA Robots for CRISPR/Cas12a Activity Management and Universal Platforms for Biosensing

Area of Science:

• Molecular Biology
• Biotechnology
• Genomics

Background:

• CRISPR/Cas12a is a powerful genome editing tool used in biosensing and diagnostics.
• Controlling Cas12a's trans-cleavage activity often requires complex methods.
• Simple modifications for precise Cas12a regulation are limited.

Purpose of the Study:

• To develop a novel, simple regulatory mechanism for CRISPR/Cas12a activity.
• To enhance the precision and selectivity of Cas12a in biosensing applications.
• To create a versatile platform for molecular diagnostics.

Main Methods:

• Introduced DNA robots (DREAM strategy) with AP site or nick activators to regulate Cas12a.
• Investigated Cas12a regulation mechanisms based on varying binding affinities.
• Developed a modular biosensor for base excision repair enzymes using DNA robots.
• Integrated multi-signal outputs (fluorescence, colorimetry, lateral flow) and logic sensing circuits.

Main Results:

• The DREAM strategy precisely regulated Cas12a activity through differential binding affinities.
• Enhanced Cas12a selectivity for identifying base mismatches.
• Successfully developed a modular biosensor for base excision repair enzymes.
• Constructed a multi-signal output platform and logic sensing circuits for simultaneous detection.

Conclusions:

• The DREAM strategy offers a new approach for programmable CRISPR/Cas12a biosensing.
• This method enables portable, specific, and user-friendly molecular diagnostics.
• The strategy holds significant potential for advancing molecular diagnostic tools.