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Off-Target Retinal Pigment Epithelium (RPE) Expression of Cre in the Rhodopsin-iCre75 Mouse Line
David G Ball1,2, Eleanor Ostafin1,3, William J Spencer1,4
1Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, New York, United States.
Investigative Ophthalmology & Visual Science
|August 11, 2026
Summary
The rhodopsin-iCre75 mouse line, widely used for targeting rod photoreceptors, shows unintended Cre expression in retinal pigment epithelium (RPE) cells. This off-target recombination impacts the interpretation of studies using this Cre mouse line.
Area of Science:
- Genetics and Genomics
- Developmental Biology
- Ophthalmology
Background:
- Transgenic Cre mouse lines are crucial for cell-specific gene manipulation.
- Off-target Cre expression can compromise study validity.
- The rhodopsin-iCre75 line has been a standard for rod photoreceptor research.
Purpose of the Study:
- To re-evaluate the cell-type specificity of the rhodopsin-iCre75 mouse line.
- To determine if Cre expression is restricted to rod photoreceptors.
- To assess potential off-target recombination in other ocular cell types.
Main Methods:
- Crossed rhodopsin-iCre75 mice with the Ai14 Cre reporter strain (tdTomato expression).
- Analyzed Cre recombination in ocular tissues using confocal microscopy.
- Validated Cre protein presence via western blotting.
Main Results:
- Cre expression was not specific to rod photoreceptors in the rhodopsin-iCre75 line.
- Significant Cre recombination observed in retinal pigment epithelium (RPE) cells.
- Approximately 6% of RPE cells showed Cre recombination by postnatal day 4, with higher central concentrations.
Conclusions:
- The rhodopsin-iCre75 mouse line exhibits unintended Cre recombination in RPE cells.
- These findings necessitate re-interpretation of previous studies using this line.
- Future research must account for this RPE-specific off-target activity.
