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M.J. Tolentino, S.J. Reich, J. Fosnot, A. Kuroki, W.X. Tang, A.M. Maguire, J. Bennett; In Vivo RNA Interference in the Murine RPE . Invest. Ophthalmol. Vis. Sci. 2003;44(13):2331.
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Purpose: RNA interference mediated by small interfering RNAs (siRNAs) is a powerful technology capable of silencing mamalian genes with great specificity and potency. The purpose of this study is to demonstrate RNA interference mediated by siRNA in the murine retina in vivo. Methods:Mice received bi-lateral subretinal injections with adenovirus expressing GFP driven by the CMV promoter. In the Ad-CMV-GFP infected animals, siRNA directed at GFP was injected in one eye and in the contrateral eye an unrelated siRNA was injected as a control. Indirect ophthalmoscopy was performed 48hours post injection . Mice were sacrificed at 48 hours and 60 hours postinjection and either choroidal flat mounts were made or the eyes were serial sectioned . Standard fluorescence microscopy was used to analyze both the choroidal flat mounts and the 10um sections. Results: Ophthalmoscopically silencing of GFP was obvious in 80% of mice eyes injected with siRNA directed at GFP. On choroidal flat mounts and in serial sections attenuation of GFP expression was demonstrated in all eyes that received siRNA directed at GFP. No silencing was noted in eyes injected with non specific control siRNA. Conclusions: siRNA can be delivered to the murine retina in vivo, are specific for the target RNA and are capable of silencing gene expression in these cells.
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