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Development of a DNA chip for the diagnosis of the most common corneal dystrophies caused by mutations in the βigh3 gene
  1. So Young Yoo1,
  2. Tae-Im Kim2,
  3. Sang Yup Lee3,
  4. Eung Kweon Kim2,
  5. Ki Chang Keum4,
  6. Nae Choon Yoo5,
  7. Won Min Yoo6
  1. 1Department of Chemical and Biomolecular Engineering (BK21 program), Korea Advanced Institute of Science and Technology, Daejeon, Korea; Medigenes, Seoul, Korea
  2. 2Corneal Dystrophy Research Institute, Department of Ophthalmology, Project Team of Nanobiomaterials for Cell-based Implants, Severance Hospital, Yonsei University, Seoul, Korea
  3. 3Department of Chemical and Biomolecular Engineering (BK21 program), Centre for Systems and Synthetic Biotechnology, Institute for the BioCentury, Korea Advanced Institute of Science and Technology, Daejon, Korea
  4. 4Department of Radiation Oncology, Yonsei University College of Medicine, Seoul, Korea
  5. 5Department of Oncology and Cancer Centre, Yonsei University College of Medicine, Seoul, Korea
  6. 6Department of Plastic and Reconstructive Surgery, Yongdong Severance Hospital, Seoul, Korea
  1. Correspondence to: Professor S Y Lee Department of Chemical & Biochemical Engineering, Korea Advanced Institute of Science and Technology (KAIST), 373–1 Guseong-dong, Yuseong-gu,Daejeon 305–701, Korea; leesy{at}kaist.ac.kr; Professor E K Kim, Department of Ophthalmology, Yousei University College of Medicine, Seodaemoongu Shinchondong 134, C PO Box 8044, Seoul, Korea; eungkkim{at}yumc.yonsei.ac.kr

Abstract

Aim: To develop a diagnostic DNA chip to detect mutations in the βigh3 gene causing the most common corneal dystrophies (CDs).

Methods: Samples from 98 people, including patients with βigh3-associated CDs (β-aCDs), were examined. Specific primer and probe sets were designed to examine exons 4 and 12 of the βigh3 gene, in order to identify mutant and wild-type alleles. Mutations were then identified by hybridisation signals of sequence-specific probes immobilised on the slide glass.

Results: Direct sequencing of exons 4 and 12 of the βigh3 gene in the patients’ genome showed that β-aCDs could be mainly classified into five types: homozygotic Avellino corneal dystrophy (ACD), heterozygotic ACD, heterozygotic lattice CD I, heterozygotic Reis–Bucklers CD and heterozygotic granular CD. Blind tests were performed by applying the target DNA amplified from the genomic DNA isolated from the peripheral blood of the participants onto a DNA chip. The results obtained by DNA chip hybridisation matched well with the direct DNA sequencing results.

Conclusions: The DNA chip developed in this study allowed successful detection of β-aCDs with a sensitivity of 100%. Mutational analysis of exons 4 and 12 of the βigh3 gene, which are the mutational hot spots causing β-aCDs, can be successfully performed with the DNA chip. Thus, this DNA chip-based method should allow a convenient, yet highly accurate, diagnosis of β-aCDs, and can be further applied to diagnose other types of CDs.

  • ACD, Avellino corneal dystrophy
  • β-aCD, βigh3-associated corneal dystrophy
  • CD, corneal dystrophy
  • FITC, fluorescein isothiocyanate
  • GCD, granular CD
  • LCD I, lattice CD type I
  • Ori, origin of replication
  • RBCD, Reis–Bucklers CD
  • SSPE, saline/sodium phosphate/EDTA

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Footnotes

  • Published Online First 10 January 2007

  • Competing interests: None.

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