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DNA chip for diagnosis of corneal dystrophy

A DNA chip, malnutrition technology, applied in the field of oligonucleotides, can solve problems such as visual loss

Inactive Publication Date: 2009-02-25
MEDIGENES CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Therefore, although an accurate diagnosis of Avellino corneal dystrophy is required to prevent Avellino corneal dystrophy through LASIK surgery, the diagnosis of Avellino corneal dystrophy is only made by microscopic observation of corneal transparency, so physicians often overlook the need for LASIK surgery in patients. Potential symptoms leading to vision loss

Method used

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  • DNA chip for diagnosis of corneal dystrophy
  • DNA chip for diagnosis of corneal dystrophy
  • DNA chip for diagnosis of corneal dystrophy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0102] Example 1: Determining the mutation type of BIGH3 protein

[0103] In order to construct a diagnostic probe for determining the gene mutation of BIGH3, one of the causes of ophthalmic diseases including Avellino corneal dystrophy, its mutation site was identified to generate a probe.

[0104] The mutation sites of the BIGH3 protein were identified, and their amino acid and nucleotide sequences were guaranteed through GenBank and OMIM (Online Medelian Inheritance in Man), NCBI Nucleotide Sequence Database (National Center for Biotechnology Information), and the equivalent of each disease bit genetic information is understood. The type of mutation to be examined was determined to test the efficacy of the DNA chip. Especially among hotspots for BIGH3 gene mutations, regions of mutations causing Avellino dystrophy, Lattice I dystrophy, and Ray-Brühl I were selected (Table 1).

[0105] Table 1. Ophthalmic disorders caused by mutations in the BIGH3 gene

[0106] ...

Embodiment 2

[0108] Example 2: Obtaining the search region by PCR

[0109] To retrieve all the mutations shown in Table 1, five pairs of primers including exon 4, exon 11, exon 12 and exon 14 were designed. Among these primers, two pairs of primers were used to amplify the mutated region of exon 4. One of the two pairs of primers (primer 1 and primer 2) was considered suitable for the experiment of diagnostic DNA chip, and the other pair was designed for direct DNA sequence analysis (primer 3 and primer 4). In addition, DNA probes and their complementary primers to be searched (reverse primers) are labeled with fluorescent chemicals on their 5' hydroxyl groups. Those primer pairs were thus effectively distinguished by Cy5 and Cy3 binding to primer 2 and primer 4, respectively.

[0110] Table 2: Primers used to amplify the region of the gene including the mutation site

[0111] Primer# serial number. sequence(5'->3') 1 1 agc cct acc act ctc aa 2 2 cag gcc tc...

Embodiment 3

[0114] Example 3: Manufacturing of probes for diagnosing BIGH3 gene mutations

[0115] To retrieve the corneal dystrophy mutations shown in Table 1, probes were designed containing sequences extending 5-8 more nucleotides, preferably 7 more nucleotides, from both sides of the hotspot. Probes were designed for the normal counterparts, comprising the same sequence as above but with the nucleotide sequence mutated in the center replaced by the normal.

[0116] Table 3: Constructed probes for the diagnosis of ophthalmic disease mutations

[0117]

serial number

genotype normal /

mutation

probe sequence

Exon 11 normal R124 acg gac cgc acg gag 4 12 Avellino Malnutrition R124H acg gac cac acg gag 4 13 Ray-Buddha (CDB1) R124L acg gac ctc acg gag 4

[0118] 14 normal R124 cac gga ccg cac gga 4 15 Lattice Type I R124C cac gga ctg cac gga 4 16 normal P501 gac ccc ccc aat ggg 11 ...

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Abstract

The present invention relates to oligonucleotides for diagnosis of corneal dystrophy. More particularly, the present invention relates to oligonucleotides for detecting mutation of BIGH3 gene for diagnosis of corneal dystrophy including Avelllino corneal dystrophy, which must be precisely diagnosed before vision correction surgery, and a DNA chip for diagnosis of corneal dystrophy, which has the oligonucleotides fixed thereon. According to the present invention, conventional microscopic diagnosis of corneal dystrophy can be replaced with a precise genetic method, which prevents a patient with corneal dystrophy from losing eyesight by eyesight correction surgery after erroneous diagnosis.

Description

technical field [0001] The present invention relates to oligonucleotides useful in the diagnosis of corneal dystrophies. In particular, the invention relates to oligonucleotides for the detection of mutations in the BIGH3 gene for the diagnosis of corneal dystrophies, including Avelllino corneal dystrophy, which must be accurately diagnosed prior to vision corrective surgery, and to A DNA chip for diagnosing corneal dystrophy, having oligonucleotides immobilized thereon. Background technique [0002] Corneal dystrophies are autosomal dominant disorders that begin with vague symptoms in the center of the cornea and spread gradually, so that eventually vision loss occurs as patients age. This disease includes Avellino corneal dystrophy, granular (Granular) corneal dystrophy, lattice (Lattice) type I corneal dystrophy, Reis-bucklers (Reis-bucklers) corneal dystrophy, etc., and is coded by βIG- Caused by mutations in the H3 gene. [0003] Heterozygous patients afflicted with ...

Claims

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Application Information

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IPC IPC(8): C07H19/00C12Q1/68
Inventor 李相烨刘昭永金应权琴基昌柳元敏柳来春
Owner MEDIGENES CO LTD
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