Somatic Genome-Engineered Mouse Models Using In Vivo Microinjection and Electroporation

Overview

This protocol describes a flexible method for CRISPR-mediated genome editing in the mouse oviduct, allowing for targeted mutations in a specific organ. This technique facilitates the study of cancer initiation by creating sporadically mutated cells within an immunocompetent environment.

Key Study Components

Area of Science

• Neuroscience
• Genetics
• Oncology

Background

• CRISPR technology enables precise genome editing.
• Targeting specific organs or cell types is crucial for studying disease mechanisms.
• The mouse oviduct serves as a model for cancer research.
• Electroporation enhances the efficiency of gene delivery.

Purpose of Study

• To develop a method for targeted genome editing in the mouse oviduct.
• To investigate the initiation of cancer in a controlled environment.
• To allow for flexible targeting of various cell types without the need for specific mouse lines.

Methods Used

• Microinjection of CRISPR components into the oviduct.
• Electroporation to facilitate gene editing.
• Use of fluorescent markers to identify edited cells.
• Isolation of cells for DNA sequencing to confirm editing.

Main Results

• Successful injection and electroporation of the oviduct.
• Creation of sporadically mutated cells among unedited cells.
• Electroporated cells were restricted to the mucosal epithelium.
• Fluorescent sorting confirmed CRISPR-mediated genome editing.

Conclusions

• This method provides a robust tool for studying gene function in vivo.
• It allows for the examination of cancer initiation mechanisms.
• The flexibility of the technique can be adapted for various research needs.

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