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Generation of Genetically Modified Mice through the Microinjection of Oocytes
Published on: June 15, 2017
Rapid bacterial artificial chromosome modification for large-scale mouse transgenesis.
Shiaoching Gong1, Laura Kus, Nathaniel Heintz
1The Rockefeller University, New York, New York, USA. gongs@rockefeller.edu
Nature Protocols
|October 2, 2010
Summary
This study introduces a new two-plasmid method for modifying bacterial artificial chromosomes (BACs), enabling rapid generation of over 1,000 BAC transgenic mice for research.
Area of Science:
- Molecular Biology
- Genetics
- Transgenic Technology
Background:
- Bacterial artificial chromosomes (BACs) are crucial for genetic engineering and studying gene function.
- Efficient modification of BACs is essential for generating transgenic models.
- Current methods can be time-consuming and labor-intensive.
Purpose of the Study:
- To develop a high-throughput, efficient method for modifying BACs.
- To streamline the generation of BAC transgenic mice.
- To provide a robust protocol for genetic research.
Main Methods:
- A novel two-plasmid system was employed for BAC modification.
- One plasmid contains an R6Kγ origin and a recombination cassette for BAC insertion.
- A second plasmid, pSV1.RecA, provides temporal control via a temperature-sensitive origin and recombinase gene.
Main Results:
- Successfully modified over 2,000 BACs.
- Generated more than 1,000 BAC transgenic mice.
- The complete BAC to transgenic mouse generation cycle takes approximately 5 weeks.
- Introduced marker genes such as EGFP and EGFP-L10a.
Conclusions:
- The developed method offers a highly efficient and rapid approach for BAC modification.
- This protocol significantly accelerates the creation of BAC transgenic mouse models.
- The generated resources, including vectors and EGFP reporter mice, are available to the research community.
