Easy and efficient production of completely embryonic-stem-cell-derived mice using a micro-aggregation device

Area of Science:

• Developmental Biology
• Genetics
• Bioengineering

Background:

• Genetically modified mice are crucial for organismal-level phenotypic screening.
• Current methods for generating completely embryonic-stem-cell (ESC)-derived chimeric mice, such as microinjection or conventional aggregation, require specialized skills and can be inefficient.
• There is a need for more accessible and efficient techniques for producing chimeric mice without crossing.

Purpose of the Study:

• To develop a novel micro-aggregation device to simplify and improve the precision of generating completely ESC-derived chimeric mice.
• To provide a high-throughput, user-friendly alternative to existing methods for producing chimeric mice.

Main Methods:

• Fabrication of a polystyrene funnel-like micro-aggregation device for controlled ESC aggregation.
• Utilizing surface tension at a liquid-air interface to hold aggregated ESCs within a 300-μm diameter area.
• Integration of the device with standard 96-well plates for compatibility with multichannel pipettes and automated systems.
• Transplantation of aggregated ESCs into embryos to generate chimeric mice.

Main Results:

• Successful generation of chimeric blastocysts using the micro-aggregation device.
• High efficiency in producing completely ESC-derived chimeric mice post-transplantation.
• Identification of an optimal range of 90-120 ESCs per embryo for efficient chimeric mouse generation.
• Achieved efficiency comparable to or better than the microinjection method.

Conclusions:

• The developed micro-aggregation device offers an easy, precise, and efficient method for producing completely ESC-derived chimeric mice.
• This technology is a promising alternative to conventional microinjection and aggregation techniques, particularly for high-throughput applications.
• The device facilitates rapid phenotypic screening by simplifying the generation of genetically modified organisms.