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Y.S. Rabinowitz, L. Don, J. Gao, P. Buchoff, G. Wistow; Gene expression profile of keratoconus cornea for NEIBank. . Invest. Ophthalmol. Vis. Sci. 2004;45(13):2883.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Previous attempts at making cDNA libraries from normal human cornea have yielded poor results, mainly attributable to difficulties in obtaining fresh tissue. Here we describe construction and analysis of a cDNA library from fresh corneal tissue from patients with keratoconus(KC) at the time of keratoplasty. Methods: RNA was harvested from 7 surgically removed KC corneas. cDNA was synthesized using Superscript reverse transcriptase and directionally cloned in the pCMVSport6 vector. 7680 clones were sequenced from the 5’ end for EST analysis and sequences were analyzed and organized using the program GRIST. Results: In total 4090 clusters of clones, potentially representing individual genes, were identified. Of these, 887 genes are represented by more than one clone. The five most abundant transcripts, represented by more than 60 clones each, are Keratin 12, BIGH3, decorin, ALDH3, and enolase 1, all known markers for cornea. Many other markers for epithelial, stromal and endothelial expressed genes are also present. One cluster of 6 clones comes from an apparently novel gene of unknown function located on chromosome 18p12.3. Conclusions: This is the most comprehensive cDNA library of genes expressed in the cornea described to date. It provides a great resource for new insights into both normal corneal function and the pathogenic pathways in KC.
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