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J. J. Kang, J. Zernant, J. Uksti, J. Merriam, E. Dhrami-Gavazi, L. Kleiman, A. Schrier, S. Trokel, R. E. Braunstein, R. Allikmets; Development of a Genotyping Microarray (Disease Chip) for the Screening of Corneal Dystrophies and Keratoconus. Invest. Ophthalmol. Vis. Sci. 2008;49(13):999. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To develop a screening tool for corneal dystrophies and keratoconus using genotyping microarray technology.
By systematic analysis of all published data, we identified and collected the most comprehensive database of candidate genes for corneal dystrophies and keratoconus for the corneal dystrophy gene chip. The genotyping microarray was designed and constructed by arrayed primer extension (APEX) technology, which can detect single-nucleotide polymorphisms, deletions, and insertions in heterozygous and homozygous patient samples. Each sequence change in the identified genes was included on the chip by synthesis and application of sequence-specific oligonucleotides. Intronic sequences were included only in cases with predicted or documented involvement in splicing.
Twelve candidate genes (CHST6, COL8A2, CYP4V2, GSN, KRT3, KRT12, SLC4A11, SOD1, TACSTD2, TCF8, TGFBI, VSX1) for keratoconus, Fuchs’ endothelial dystrophy, Meesmann’s juvenile epithelial dystrophy, Reis-Bucklers’ dystrophy, Thiel-Behnke dystrophy, granular dystrophy, lattice dystrophy, Avellino dystrophy, familial subepithelial amyloidosis, Bietti’s crystalline dystrophy, macular corneal dystrophy, posterior polymorphous dystrophy and congenital hereditary endothelial dystrophy type II were identified from the literature. A total of 288 disease-associated variants from the coding region and adjacent intronic sequences of the 12 corneal dystrophy and keratoconus candidate genes were included on the genotyping microarray. The resultant array allows simultaneous detection of all currently known disease-associated genetic variants in patients with keratoconus and corneal dystrophies. The genotyping microarray has been validated in 30 DNA samples. Screening in a population of clinically diagnosed corneal dystrophy and keratoconus patients is underway, followed by segregation analyses in families, if applicable.
The corneal dystrophy genotyping microarray represents the first similar screening tool for patients with genetically heterogeneous conditions of the anterior segment. Simultaneous screening for all known corneal dystrophy and keratoconus associated variants in patients and in the general population will allow systematic detection and analysis of genetic variation and identification of heterozygous carriers, facilitating prospective diagnosis and ultimately treatment options.
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