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J Yu, AJ Mears, R Farjo, S Yoshida, MI Othman, D Ghosh, A Swaroop; Retinal Gene Expression Profiling of Wildtype and Nrl Knockout Mice Utilizing I-Gene Microarrays . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1378.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To generate gene expression profiles of wildtype and Nrl knockout mice at different developmental time-points and to identify differentially expressed genes. Methods: RNA was isolated from both Nrl+/+ and Nrl-/- mice retina at four stages of development, postnatal day 0 (P0), P2, P10 and P21. Five microgram of total RNA, either from retina or a common reference RNA source, was labeled with Cy5 or Cy3 dye respectively using 3DNA® SubmicroTM Oligo Expression Array Detection Kit (Genisphere). These labeled targets were simultaneously hybridized to I-gene microarray slides produced in the Kellogg Eye Center, University of Michigan. The hybridized slides were then washed, and scanned using an Affymetrix 428 scanner to generate images of fluorescence intensity, which were then converted into expression data by GleamsTM (NuTec). Five replicate experiments were performed for each time-point to control for experimental variability and to enable statistical inference. Microarray data was then normalized, analyzed and clustered using appropriate statistical models. Results: Gene expression profiles for both Nrl+/+ and Nrl-/- mice retina during four critical developmental ages have been generated relative to a common reference RNA. A comparison between Nrl-/- and Nrl+/+ mice at P21 has identified a number of differentially expressed genes such as S-opsin, Rhodopsin, Gnat1 and several novel transcripts. Further analysis and results will be presented. Conclusion: Microarray analysis has been utilized to generate retinal gene expression profiles in wildtype and Nrl knockout mice. Up-regulated genes in Nrl-/- mice may contribute to cone development and function, while down-regulated genes may involve in rod development. These expression profiles will also assist in the identification of novel retinal disease genes and the study of cellular pathways of photoreceptor development and cone survival.
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