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Site-specific somatic mutagenesis in the retinal pigment epithelium
Mikiro Mori1, Daniel Metzger, Jean-Marie Garnier
1Institute of Genetics and Molecular and Cellular Biology (IGBMC), CNRS/INSERM/ULP/Collège de France, BP 163, 67404 Illkirch Cedex, CU de Strasbourg, France.
Purpose:
Generate site-specific somatic mutations selectively in the retinal pigment epithelium (RPE) in mice.
Methods:
A transgenic mouse line expressing the Cre recombinase under the control of the tyrosinase-related protein (TRP)-1 promoter was generated. The presence of Cre was determined by in situ hybridization, and Cre-mediated excision of DNA was analyzed by PCR and alkaline phosphatase (AP) histochemistry in reporter mice carrying a loxP-flanked (floxed) retinoid X receptor alpha (RXRa) gene and in Z/AP mice, respectively.
Results:
Cre was expressed in the RPE from embryonic day 10.5 to postnatal day 12, resulting in efficient floxed excision of DNA in the RPE from embryonic day 10.5 to adulthood in TRP1-Cre mice. Expressed Cre and excision of DNA were also detected in the ciliary margin of the retina and in some cells in the neural retina, but not in the embryonic periocular mesenchyme or in the choroid.
Conclusions:
The TRP1-Cre mouse line, which induces efficient Cre-mediated excision of DNA selectively in the RPE, provides a new, powerful tool to study gene functions in the RPE in vivo.
Insights
A new TRP1-Cre mouse model enables targeted gene mutation in the retinal pigment epithelium (RPE). This powerful tool allows for precise in vivo study of RPE gene function.
Area of Science:
- Genetics
- Ophthalmology
- Developmental Biology
Background:
- The retinal pigment epithelium (RPE) is crucial for vision.
- Studying RPE-specific gene function in vivo is essential for understanding retinal diseases.
Purpose of the Study:
- To generate a transgenic mouse model for site-specific somatic mutations in the RPE.
- To establish a tool for investigating gene function within the RPE.
Main Methods:
- Generated a transgenic mouse line using Cre recombinase under the tyrosinase-related protein (TRP)-1 promoter.
- Confirmed Cre presence via in situ hybridization.
- Analyzed DNA excision using PCR and alkaline phosphatase (AP) histochemistry in reporter mice.
Main Results:
- TRP1-Cre mice demonstrated Cre expression in the RPE from embryonic day 10.5 to postnatal day 12.
- Efficient DNA excision was observed in the RPE from embryonic stages to adulthood.
- Cre expression and DNA excision were also noted in the ciliary margin and neural retina, but not in periocular mesenchyme or choroid.
Conclusions:
- The TRP1-Cre mouse line enables selective and efficient Cre-mediated DNA excision in the RPE.
- This model serves as a valuable new tool for in vivo studies of RPE gene function.