Nuclear-Targeted Material Enabled Intranuclear MicroRNA Imaging for Tracking Gene Editing Process.

Area of Science:

• Biotechnology
• Molecular Biology
• Cancer Research

Background:

• Clustered regularly interspaced short palindromic repeats/associated protein (CRISPR/Cas) systems are vital for cancer therapy.
• Current methods for tracking gene editing use lysed cells, failing to capture dynamic changes in living cells.
• There is a critical need for in situ, real-time imaging to monitor gene editing at the single-cell level.

Purpose of the Study:

• To develop a novel nuclear-targeted material for efficient gene editing plasmid delivery and tracking.
• To achieve in situ and real-time imaging of microRNA gene editing processes in living cells.
• To enable sensitive monitoring of gene editing in vitro and in vivo and facilitate screening of effective gene-editing plasmids.

Main Methods:

• A phenylboronic acid modified linear polyethyleneimine (LPBA) material was synthesized for nuclear targeting.
• LPBA was used to load gene editing plasmids and fluorescent probes for microRNA imaging.
• The LPBA system's efficiency was compared to the linear polyethyleneimine (LPEI) delivery system in living cells and in vivo.

Main Results:

• LPBA demonstrated highly efficient nuclear targeting and loading of gene editing plasmids and probes.
• Intranuclear microRNA imaging in living cells was achieved with LPBA, showing a 32.4-fold increase compared to LPEI.
• The LPBA system facilitated sensitive, real-time monitoring of gene editing processes in both living cells and in vivo.

Conclusions:

• The developed LPBA material enables efficient intranuclear imaging of microRNA gene editing in living cells.
• This technique provides a powerful tool for real-time monitoring of gene editing and screening of gene-editing plasmids.
• The LPBA-based imaging technology offers new insights into gene editing and nucleic acid detection for cancer therapy.