Advanced microinjection protocol for gene manipulation using the model newt Pleurodeles waltl

Area of Science:

• Amphibian research
• Developmental biology
• Regeneration biology

Background:

• Urodele amphibians, like newts, are established models in developmental and regeneration biology.
• The Spanish newt (Pleurodeles waltl) serves as a model for reverse/molecular genetics via gene manipulation.
• Microinjection is a fundamental technique for gene manipulation in newts.

Purpose of the Study:

• To optimize the microinjection protocol for Pleurodeles waltl.
• To improve efficiency and reduce costs of newt microinjection.
• To establish a versatile protocol for various gene manipulation applications.

Main Methods:

• Systematic examination of egg preparation techniques.
• Evaluation of de-jelly solutions for optimal egg handling.
• Formulation and testing of improved injection media.

Main Results:

• Successfully optimized microinjection protocol for P. waltl.
• Achieved higher efficiency and lower cost compared to previous methods.
• Validated protocol for plasmid DNA, mRNA, and CRISPR-Cas9 genome editing.

Conclusions:

• The optimized protocol significantly enhances gene manipulation capabilities in newts.
• This advancement facilitates broader research applications in developmental and regeneration biology.
• The improved method lowers barriers for genetic studies in P. waltl.