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AJ Mears, R Farjo, J Yu, S Hiriyanna, SP MacNee, AS Maheshwary, A Swaroop; Identification of Novel Cone Photoreceptor Genes From Retinal Libraries Generated From the Rod-Less Nrl Knockout Mouse . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1429.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:The Nrl knockout mouse (Nrl-/-) was previously generated by gene targeting. Extensive characterization of this mouse by histology, immunohistochemistry, ERG and retinal gene expression analysis revealed a retina lacking rod photoreceptors but having an increased number of S-cone photoreceptors (Mears et al., 2001). Due to the functional switching of photoreceptor cell-types in the retina of these mice there is an enrichment of cone-specific transcripts. The goal is to identify novel cone expressed genes from the retina of these mice. Methods:We have constructed cDNA libaries from Nrl-/- retina from three different time-points (P0.5, P2.5 and P10.5) using the Superscript Plasmid System (Invitrogen). Results:All three libraries showed high fidelity with ≷90% of clones having inserts with an average size of 1.2-1.5 kb. Using bioinformatics tools, clone sequences were analyzed, annotated and compared with SAGE analysis in which rod photoreceptor-enriched transcripts have been identified (Blackshaw et al. 2001). To determine which of these transcripts are more highly expressed in the Nrl-/- mouse (i.e. cone photoreceptor-enriched), PCR-amplified inserts from the libraries are being spotted onto glass slides for microarray analysis. This data will be complementary to other gene expression studies on the Nrl-/- mouse that are utilizing custom mouse eye gene arrays and Affymetrix gene chips. Conclusion:We have generated and sequenced cDNA libraries from the retina of the rod-less Nrl-/- mouse. These clones are being currently utilized in microarray analysis to identify genes that demonstrate a cone photoreceptor enriched or specific expression. This information will lead to the further elucidation of pathways associated with cone photoreceptor development and function. Mears AJ, Kondo M, Swain PK, Takada Y, Bush RA, Saunders TL, Sieving PA and Swaroop A. (2001) Nrl is required for rod photoreceptor development. Nature Genet. 29, 447-452. Blackshaw S, Fraioli RE, Furukawa T, and Cepko CL. (2001) Comprehensive analysis of photoreceptor gene expression and the identification of candidate retinal disease genes. Cell 107, 579-589.
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