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Updated: May 23, 2026
Ex Vivo Organoid Model of Adenovirus-Cre Mediated Gene Deletions in Mouse Urothelial Cells
Published on: May 5, 2022
A BAC-based transgenic mouse specifically expresses an inducible Cre in the urothelium
Tian Huai Shen1, Nataliya Gladoun, Mireia Castillo-Martin
1Herbert Irving Comprehensive Cancer Center, Columbia University Medical Center, New York, New York, United States of America.
Abstract:
Cre-loxp mediated conditional knockout strategy has played critical roles for revealing functions of many genes essential for development, as well as the causal relationships between gene mutations and diseases in the postnatal adult mice. One key factor of this strategy is the availability of mice with tissue- or cell type-specific Cre expression. However, the success of the traditional molecular cloning approach to generate mice with tissue specific Cre expression often depends on luck. Here we provide a better alternative by using bacterial artificial chromosome (BAC)-based recombineering to insert iCreERT2 cDNA at the ATG start of the Upk2 gene. The BAC-based transgenic mice express the inducible Cre specifically in the urothelium as demonstrated by mRNA expression and staining for LacZ expression after crossing with a Rosa26 reporter mouse. Taking into consideration the size of the gene of interest and neighboring genes included in a BAC, this method should be widely applicable for generation of mice with tissue specific gene expression or deletions in a more specific manner than previously reported.
Insights
We developed a new method for creating genetically modified mice with specific Cre expression in the urothelium. This bacterial artificial chromosome (BAC)-based recombineering approach improves tissue-specific gene targeting for research.
Area of Science:
- Genetics
- Developmental Biology
- Molecular Biology
Background:
- Cre-loxp recombination is crucial for studying gene function and disease models in mice.
- Generating mice with specific Cre expression is essential but challenging with traditional methods.
- The availability of reliable tissue-specific Cre driver lines is a key limitation.
Purpose of the Study:
- To develop a more efficient and reliable method for generating tissue-specific Cre-expressing mice.
- To create a novel bacterial artificial chromosome (BAC)-based system for inducible Cre expression in the urothelium.
- To validate the specificity and efficacy of the new transgenic mouse model.
Main Methods:
- Utilized bacterial artificial chromosome (BAC)-based recombineering to insert iCreERT2 cDNA into the Upk2 gene locus.
- Generated BAC-transgenic mice carrying the modified Upk2 locus.
- Crossed the generated mice with a Rosa26 reporter mouse line to assess Cre activity.
- Confirmed urothelial-specific Cre expression via mRNA analysis and LacZ staining.
Main Results:
- Successfully generated BAC-transgenic mice with inducible Cre recombinase expression.
- Demonstrated highly specific Cre expression within the urothelium.
- Reporter gene activation confirmed functional Cre activity in the target tissue.
- The BAC-based recombineering approach proved efficient and reproducible.
Conclusions:
- BAC-based recombineering offers a superior alternative to traditional cloning for generating tissue-specific Cre driver mice.
- This method enables precise control over gene expression and deletion in specific cell types, like the urothelium.
- The developed Upk2-iCreERT2 BAC transgenic line is a valuable tool for studying urothelial biology and disease.
